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Sample records for los genes apc

  1. Nuclear APC.

    PubMed

    Neufeld, Kristi L

    2009-01-01

    Mutational inactivation of the tumor suppressor gene APC (Adenomatous polyposis coli) is thought to be an initiating step in the progression of the vast majority ofcolorectal cancers. Attempts to understand APC function have revealed more than a dozen binding partners as well as several subcellular localizations including at cell-cell junctions, associated with microtubules at the leading edge of migrating cells, at the apical membrane, in the cytoplasm and in the nucleus. The present chapter focuses on APC localization and functions in the nucleus. APC contains two classical nuclear localization signals, with a third domain that can enhance nuclear import. Along with two sets of nuclear export signals, the nuclear localization signals enable the large APC protein to shuttle between the nucleus and cytoplasm. Nuclear APC can oppose beta-catenin-mediated transcription. This down-regulation of nuclear beta-catenin activity by APC most likely involves nuclear sequestration of beta-catenin from the transcription complex as well as interaction of APC with transcription corepressor CtBP. Additional nuclear binding partners for APC include transcription factor activator protein AP-2alpha, nuclear export factor Crm1, protein tyrosine phosphatase PTP-BL and perhaps DNA itself. Interaction of APC with polymerase beta and PCNA, suggests a role for APC in DNA repair. The observation that increases in the cytoplasmic distribution of APC correlate with colon cancer progression suggests that disruption of these nuclear functions of APC plays an important role in cancer progression. APC prevalence in the cytoplasm of quiescent cells points to a potential function for nuclear APC in control of cell proliferation. Clear definition of APC's nuclear function(s) will expand the possibilities for early colorectal cancer diagnostics and therapeutics targeted to APC. PMID:19928349

  2. ApcMin, A Mutation in the Murine Apc Gene, Predisposes to Mammary Carcinomas and Focal Alveolar Hyperplasias

    NASA Astrophysics Data System (ADS)

    Moser, Amy Rapaich; Mattes, Ellen M.; Dove, William F.; Lindstrom, Mary J.; Haag, Jill D.; Gould, Michael N.

    1993-10-01

    ApcMin (Min, multiple intestinal neoplasia) is a point mutation in the murine homolog of the APC gene. Min/+ mice develop multiple intestinal adenomas, as do humans carrying germ-line mutations in APC. Female mice carrying Min are also prone to develop mammary tumors. Min/+ mammary glands are more sensitive to chemical carcinogenesis than are +/+ mammary glands. Transplantation of mammary cells from Min/+ or +/+ donors into +/+ hosts demonstrates that the propensity to develop mammary tumors is intrinsic to the Min/+ mammary cells. Long-term grafts of Min/+ mammary glands also gave rise to focal alveolar hyperplasias, indicating that the presence of the Min mutation also has a role in the development of these lesions.

  3. Methylation of the adenomatous polyposis coli (APC) gene in human placenta and hypermethylation in choriocarcinoma cells.

    PubMed

    Wong, N C; Novakovic, B; Weinrich, B; Dewi, C; Andronikos, R; Sibson, M; Macrae, F; Morley, R; Pertile, M D; Craig, J M; Saffery, R

    2008-09-01

    Methylation of the human APC gene promoter is associated with several different types of cancers and has also been documented in some pre-cancerous tissues. We have examined the methylation of APC gene promoters in human placenta and choriocarcinoma cells. This revealed a general hypomethylation of the APC-1b promoter and a pattern with monoallelic methylation of the APC-1a promoter in full term placental tissue. However, there was no evidence of a parent-of-origin effect, suggesting random post zygotic origin of methylation. Increased methylation of this promoter was observed in all choriocarcinoma-derived trophoblast cell lines, suggesting a trophoblastic origin of placental APC methylation and implicating APC hypermethylation in the development of this group of gestational tumours. Our demonstration of placental methylation of the APC-1a promoter represents the first observation of monoallelic methylation of this gene in early development, and provides further support for a role of canonical Wnt signalling in placental trophoblast invasiveness. This also implicates tumour suppressor gene silencing as an integral part of normal human placental development. PMID:18485586

  4. Germ-line mutations of the APC gene in 53 familial adenomatous polyposis patients.

    PubMed Central

    Miyoshi, Y; Ando, H; Nagase, H; Nishisho, I; Horii, A; Miki, Y; Mori, T; Utsunomiya, J; Baba, S; Petersen, G

    1992-01-01

    We searched for germ-line mutations of the APC gene in 79 unrelated patients with familial adenomatous polyposis using a ribonuclease protection analysis coupled with polymerase chain reaction amplifications of genomic DNA. Mutations were found in 53 patients (67%); 28 of the mutations were small deletions and 2 were 1- to 2-base-pair insertions; 19 were point mutations resulting in stop codons and only 4 were missense point mutations. Thus, 92% of the mutations were predicted to result in truncations of the APC protein. More than two-thirds (68%) of the mutations were clustered in the 5' half of the last exon, and nearly two-fifths of the total mutations occurred at one of five positions. This information has significant implications for understanding the role of APC mutation in inherited forms of colorectal neoplasia and for designing effective methods for genetic counseling and presymptomatic diagnosis. Images PMID:1316610

  5. Disruption of the APC gene by t(5;7) translocation in a Turcot family.

    PubMed

    Sahnane, Nora; Bernasconi, Barbara; Carnevali, Ileana; Furlan, Daniela; Viel, Alessandra; Sessa, Fausto; Tibiletti, Maria Grazia

    2016-03-01

    Turcot syndrome (TS) refers to the combination of colorectal polyps and primary tumours of the central nervous system. TS is a heterogeneous genetic condition due to APC and/or mismatch repair germline mutations. When APC is involved the vast majority of mutations are truncating, but in approximately 20%-30% of patients with familial polyposis no germline mutation can be found. A 30-year-old Caucasian woman with a positive pedigree for TS was referred to our Genetic Counselling Service. She was negative for APC and MUTYH but showed a reciprocal balanced translocation t(5;7)(q22;p15) at chromosome analysis. FISH analysis using specific BAC probes demonstrated that 5q22 breakpoint disrupted the APC gene. Transcript analysis by MLPA and digital PCR revealed that the cytogenetic rearrangement involving the 3' end of the APC gene caused a defective expression of a truncated transcript. This result allowed cytogenetic analysis to be offered to all the other family members and segregation analysis clearly demonstrated that all the carriers were affected, whereas non-carriers did not have the polyposis. A cytogenetic approach permitted the identification of the mutation-causing disease in this family, and the segregation analysis together with the transcript study supported the pathogenetic role of this mutation. Karyotype analysis was used as a predictive test in all members of this family. This family suggests that clinically positive TS and FAP cases, which test negative with standard molecular analysis, could be easily and cost-effectively resolved by a classical and molecular cytogenetic approach. PMID:26797314

  6. APC alterations are frequently involved in the pathogenesis of acinar cell carcinoma of the pancreas, mainly through gene loss and promoter hypermethylation.

    PubMed

    Furlan, Daniela; Sahnane, Nora; Bernasconi, Barbara; Frattini, Milo; Tibiletti, Maria Grazia; Molinari, Francesca; Marando, Alessandro; Zhang, Lizhi; Vanoli, Alessandro; Casnedi, Selenia; Adsay, Volkan; Notohara, Kenji; Albarello, Luca; Asioli, Sofia; Sessa, Fausto; Capella, Carlo; La Rosa, Stefano

    2014-05-01

    Genetic and epigenetic alterations involved in the pathogenesis of pancreatic acinar cell carcinomas (ACCs) are poorly characterized, including the frequency and role of gene-specific hypermethylation, chromosome aberrations, and copy number alterations (CNAs). A subset of ACCs is known to show alterations in the APC/β-catenin pathway which includes mutations of APC gene. However, it is not known whether, in addition to mutation, loss of APC gene function can occur through alternative genetic and epigenetic mechanisms such as gene loss or promoter methylation. We investigated the global methylation profile of 34 tumor suppressor genes, CNAs of 52 chromosomal regions, and APC gene alterations (mutation, methylation, and loss) together with APC mRNA level in 45 ACCs and related peritumoral pancreatic tissues using methylation-specific multiplex ligation probe amplification (MS-MLPA), fluorescence in situ hybridization (FISH), mutation analysis, and reverse transcription-droplet digital PCR. ACCs did not show an extensive global gene hypermethylation profile. RASSF1 and APC were the only two genes frequently methylated. APC mutations were found in only 7 % of cases, while APC loss and methylation were more frequently observed (48 and 56 % of ACCs, respectively). APC mRNA low levels were found in 58 % of cases and correlated with CNAs. In conclusion, ACCs do not show extensive global gene hypermethylation. APC alterations are frequently involved in the pathogenesis of ACCs mainly through gene loss and promoter hypermethylation, along with reduction of APC mRNA levels. PMID:24590585

  7. An AT-rich region in the APC gene may cause misinterpretation of familial adenomatous polyposis molecular screening.

    PubMed

    Palmirotta, Raffaele; De Marchis, Maria Laura; Ludovici, Giorgia; Leone, Barbara; Valente, Maria Giovanna; Alessandroni, Jhessica; Spila, Antonella; Della-Morte, David; Guadagni, Fiorella

    2012-05-01

    Familial adenomatous polyposis (FAP) is an autosomal-dominant condition mainly due to a mutation of the adenomatous polyposis coli (APC) gene. The present study reports evidence of a technical issue occurring during the mutational analysis of APC exon 4. Genetic conventional direct sequence analysis of a repetitive AT-rich region in the splice acceptor site of APC intron 3 could be misinterpreted as a pathogenetic frameshift result. However, this potential bias may be bypassed adopting a method for random mutagenesis of DNA based on the use of a triphosphate nucleoside analogues mixture. Using this method as a second-level analysis, we also demonstrated the nonpathogenic nature of the variant in the poly A trait in APC exon 4 region (c.423-4delA) that do not result in aberrant splicing of APC exons 3-4; conversely, we did not find a previously reported T deletion/insertion polymorphism. PMID:22447671

  8. Molecular genetic analysis of exons 1 to 6 of the APC gene in non-polyposis familial colorectal cancer.

    PubMed

    Joyce, J A; Froggatt, N J; Davies, R; Evans, D G; Trembath, R; Barton, D E; Maher, E R

    1995-12-01

    Familial adenomatous polyposis coli is caused by constitutional mutations in the APC gene. The hallmark of familial adenomatous polyposis coli is the presence of numerous (> 100) colorectal polyps, but mutations in the 5' end of the APC gene have been associated with familial colorectal cancer without florid polyposis. Although familial adenomatous polyposis coli accounts for only a minority of familial colorectal cancer cases, we hypothesised that APC mutations which were not associated with florid polyposis might make a significant contribution to nonpolyposis familial colorectal cancer. To investigate this possibility, we analysed 40 unrelated patients with familial colorectal cancer without classical familial adenomatous polyposis coli for mutations in exons 1 to 6 (codons 1 to 243) of the APC gene. No mutations were detected, but a C-->T polymorphism at nucleotide 333 (Arg-->Trp at codon 99) was identified. No 5' APC mutations were detected in two patients with desmoid tumours and a family history of colorectal cancer and polyps. We conclude that mutations in exons 1 to 6 of the APC gene are infrequent in patients with familial colorectal cancer who do not have many colorectal polyps. PMID:8835324

  9. Antitumor Molecular Mechanism of Chlorogenic Acid on Inducting Genes GSK-3 β and APC and Inhibiting Gene β -Catenin.

    PubMed

    Xu, Ruoshi; Kang, Qiumei; Ren, Jie; Li, Zukun; Xu, Xiaoping

    2013-01-01

    Objective. Inhibiting gene β -catenin and inducting genes GSK-3 β and APC, promoting the tumor cell apoptosis in Wnt pathway, by chlorogenic acid were discussed (CGA). Method. The different genes were scanned by the 4∗44K mouse microarray chips. The effect of the three genes was confirmed by RT-PCR technique with CGA dosage of 5, 10, and 20 mg/kg. Result. The expression of GSK-3 β and APC was upregulated in group of 20 mg/kg dosage (P < 0.05) and the expression of β -catenin was downregulated in the same dosage (P < 0.05). Conclusion. The results infer that the multimeric protein complex of β -catenin could be increased by CGA upregulated genes GSK-3 β and APC, which could inhibit the free β -catenin into the nucleus to connect with TCF. So the transcriptional expression of the target genes will be cut to abnormal cell proliferation. It is probably one of the ways that can stop the tumor increase by CGA. PMID:23844319

  10. Molecular analysis of the APC gene in 71 Israeli families: 17 novel mutations.

    PubMed

    Gavert, Nancy; Yaron, Yuval; Naiman, Tova; Bercovich, Dani; Rozen, Paul; Shomrat, Ruth; Legum, Cyril; Orr-Urtreger, Avi

    2002-06-01

    Familial adenomatous polyposis (FAP) is caused by germline mutations in the APC gene. This study included 71 Israeli families referred for molecular analysis of the APC gene. Analysis was performed by the protein truncation test (PTT) of exon 15, and if negative, by direct sequencing of exon 1 to 14. Mutations were found in 36 (50.7%) probands. Mutation detection rates depended on the pattern of referral, such that among the 40 probands referred from the Service for Hereditary Cancer the mutation detection rate was 70%, whereas among the 31 probands referred by other gastroenterologists detection rate was significantly lower (25.8%). Of the 36 mutations detected, 21 were within exon 15, 13 within exons 1 to 14 and 2 were newly-described splicing mutations in introns 9 and 14. A relatively high proportion of the mutations was detected in exon 9 (6/36), five of them newly described. Altogether, we describe here 17 new mutations. Within the two major ethnic groups in Israel, patients of Ashkenazi and non-Ashkenazi origin, there was no significant differences in the mutation detection rate or the distribution of mutations within the APC gene. No founder mutation was detected in any of these populations. Our data confirm that higher detection rates may be expected in patients referred by clinical services specializing in hereditary colon cancer. These results further underscore the importance of complete analysis of all exons and exon/intron boundaries, in order to achieve maximal detection rate in patients suspected of FAP. PMID:12007223

  11. Hereditary desmoid disease due to a frameshift mutation at codon 1924 of the APC gene.

    PubMed Central

    Eccles, D. M.; van der Luijt, R.; Breukel, C.; Bullman, H.; Bunyan, D.; Fisher, A.; Barber, J.; du Boulay, C.; Primrose, J.; Burn, J.; Fodde, R.

    1996-01-01

    Desmoid tumors are slowly growing fibrous tumors highly resistant to therapy and often fatal. Here, we report hereditary desmoid disease (HDD), a novel autosomal dominant trait with 100% penetrance affecting a three-generation kindred. Desmoid tumors are usually a complication of familial adenomatous polyposis, a predisposition to the early development of premalignant adenomatous polyps in the colorectum due to chain-terminating mutations of the APC gene. In general, one or more members in approximately 10% of the FAP families manifest desmoid tumors. Affected individuals from the HDD kindred are characterized by multifocal fibromatosis of the paraspinal muscles, breast, occiput, arms, lower ribs, abdominal wall, and mesentery. Osteomas, epidermal cysts, and other congenital features were also observed. We show that HDD segregates with an unusual germ-line chain-terminating mutation at the 3' end of the APC gene (codon 1924) with somatic loss of the wild-type allele leading to tumor development. Images Figure 2 Figure 3 Figure 4 Figure 5 PMID:8940264

  12. Promoter methylation of APC and RAR-β genes as prognostic markers in non-small cell lung cancer (NSCLC).

    PubMed

    Feng, Hongxiang; Zhang, Zhenrong; Qing, Xin; Wang, Xiaowei; Liang, Chaoyang; Liu, Deruo

    2016-02-01

    Aberrant promoter hypermethylations of tumor suppressor genes are promising markers for lung cancer diagnosis and prognosis. The purpose of this study was to determine methylation status at APC and RAR-β promoters in primary NSCLC, and whether they have any relationship with survival. APC and RAR-β promoter methylation status were determined in 41 NSCLC patients using methylation specific PCR. APC promoter methylation was detectable in 9 (22.0%) tumor samples and 6 (14.6%) corresponding non-tumor samples (P=0.391). RAR-β promoter methylation was detectable in 13 (31.7%) tumor samples and 4 (9.8%) corresponding non-tumor samples (P=0.049) in the NSCLC patients. APC promoter methylation was found to be associated with T stage (P=0.046) and nodal status (P=0.019) in non-tumor samples, and with smoking (P=0.004) in tumor samples. RAR-β promoter methylation was found associated with age (P=0.031) in non-tumor samples and with primary tumor site in tumor samples. Patients with APC promoter methylation in tumor samples showed significantly longer survival than patients without it (Log-rank P=0.014). In a multivariate analysis of prognostic factors, APC methylation in tumor samples was an independent prognostic factor (P=0.012), as were N1 positive lymph node number (P=0.025) and N2 positive lymph node number (P=0.06). Our study shows that RAR-β methylation detected in lung tissue may be used as a predictive marker for NSCLC diagnosis and that APC methylation in tumor sample may be a useful marker for superior survival in NSCLC patients. PMID:26681652

  13. Loss of heterozygosity of the APC and MCC genes in squamous cell carcinoma of the head and neck

    SciTech Connect

    Nogueira, C.P.; Afridi, N.A.; Licameli, G.

    1994-09-01

    Squamous cell carcinoma of the head and neck offers a unique opportunity to study genetic mechanisms in human tumorigenesis. Two types of premalignant lesions can be identified clinically, suggesting a multistep process. The majority of the lesions are easy to identify and access. Despite these advantages, possible genetic models for these cancers remain relatively unexplored. Loss of heterozygosity of the tumor suppressor genes APC (adenomatous polyposis coli) and MCC (mutated in colon cancer) was investigated in 24 squamous cell carcinomas of the head and neck. Three sites were analyzed by assays based on the polymerase chain reaction: two RFLPs in the APC gene (Rsa I on exon 11; Ssp I on the 3{prime} untranslated region) and an insertion/deletion in exon 10 of the MCC gene. Sixty three percent of the individuals were informative for at least one marker. Loss of heterozygosity (LOH) was observed in 33% of the tumors, either at the APC or MCC locus. Two out of 11 informative tumors (18%) showed LOH on the APC gene. Three out of seven informative tumors (43%) showed LOH on the MCC gene. These results suggest that inactivation of the APC, MCC and/or a linked gene on chromosome 5q plays a important role in squamous cell carcinoma of the head and neck. The data is in agreement with another study, which detected 25% of loss of heterozygosity in the chromosomal region 5q by using microsatellite markers. These values are much lower than the ones observed in esophageal cancer, a closely related type of tumor, with the same etiology. This may be attributed to differences in the biology of these two cancers.

  14. Expression and promoter methylation status of hMLH1, MGMT, APC, and CDH1 genes in patients with colon adenocarcinoma.

    PubMed

    Michailidi, Christina; Theocharis, Stamatios; Tsourouflis, Gerasimos; Pletsa, Vasiliki; Kouraklis, Gregorios; Patsouris, Efstratios; Papavassiliou, Athanasios G; Troungos, Constantinos

    2015-12-01

    Colorectal cancer (CRC) is the third most common cancer in men and the second in women worldwide. CRC development is the result of genetic and epigenetic alterations accumulation in the epithelial cells of colon mucosa. In the present study, DNA methylation, an epigenetic event, was evaluated in tumoral and matching normal epithelium in a cohort of 61 CRC patients. The results confirmed and expanded knowledge for the tumor suppressor genes hMLH1, MGMT, APC, and CDH1. Promoter methylation was observed for all the examined genes in different percentage. A total of 71% and 10% of the examined cases were found to be methylated in two or more and in all genes, respectively. mRNA and protein levels were also evaluated. Promoter methylation of hMLH1, MGMT, APC, and CDH1 genes was present at the early stages of tumor's formation and it could also be detected in the normal mucosa. Correlations of the methylated genes with patient's age and tumor's clinicopathological characteristics were also observed. Our findings suggest that DNA methylation is a useful marker for tumor progression monitoring and that promoter methylation in certain genes is associated with more advanced tumor stage, poor differentiation, and metastasis. PMID:25908636

  15. APC16 is a conserved subunit of the anaphase-promoting complex/cyclosome

    PubMed Central

    Kops, Geert J. P. L.; van der Voet, Monique; Manak, Michael S.; van Osch, Maria H. J.; Naini, Said M.; Brear, Andrea; McLeod, Ian X.; Hentschel, Dirk M.; Yates, John R.; van den Heuvel, Sander; Shah, Jagesh V.

    2010-01-01

    Error-free chromosome segregation depends on timely activation of the multi-subunit E3 ubiquitin ligase APC/C. Activation of the APC/C initiates chromosome segregation and mitotic exit by targeting critical cell-cycle regulators for destruction. The APC/C is the principle target of the mitotic checkpoint, which prevents segregation while chromosomes are unattached to spindle microtubules. We now report the identification and characterization of APC16, a conserved subunit of the APC/C. APC16 was found in association with tandem-affinity-purified mitotic checkpoint complex protein complexes. APC16 is a bona fide subunit of human APC/C: it is present in APC/C complexes throughout the cell cycle, the phenotype of APC16-depleted cells copies depletion of other APC/C subunits, and APC16 is important for APC/C activity towards mitotic substrates. APC16 sequence homologues can be identified in metazoans, but not fungi, by four conserved primary sequence stretches. We provide evidence that the C. elegans gene K10D2.4 and the D. rerio gene zgc:110659 are functional equivalents of human APC16. Our findings show that APC/C is composed of previously undescribed subunits, and raise the question of why metazoan APC/C is molecularly different from unicellular APC/C. PMID:20392738

  16. Putative direct and indirect Wnt targets identified through consistent gene expression changes in APC-mutant intestinal adenomas from humans and mice

    PubMed Central

    Segditsas, Stefania; Sieber, Oliver; Deheragoda, Maesha; East, Phil; Rowan, Andrew; Jeffery, Rosemary; Nye, Emma; Clark, Susan; Spencer-Dene, Bradley; Stamp, Gordon; Poulsom, Richard; Suraweera, Nirosha; Silver, Andrew; Ilyas, Mohammad; Tomlinson, Ian

    2008-01-01

    In order to identify new genes with differential expression in early intestinal tumours, we performed mRNA (messenger ribonucleic acid) expression profiling of 16 human and 63 mouse adenomas. All individuals had germline APC mutations to ensure that tumorigenesis was driven by ‘second hits’ at APC. Using stringent filtering to identify changes consistent between humans and mice, we identified 60 genes up-regulated and 151 down-regulated in tumours. For 22 selected genes—including known Wnt targets—expression differences were confirmed by qRT–PCR (quantitative reverse transcription polymerase chain reaction). Most, but not all, differences were also present in colorectal carcinomas. In situ analysis showed a complex picture. Expression of up-regulated genes in adenomas was usually uniform/diffuse (e.g. ITGA6) or prominent in the tumour core (e.g. LGR5); in normal tissue, these genes were expressed at crypt bases or the transit amplifying zone. Down-regulated genes were often undetectable in adenomas, but in normal tissue were expressed in mesenchyme (e.g. GREM1/2) or differentiated cells towards crypt tops (e.g. SGK1). In silico analysis of TCF4-binding motifs showed that some of our genes were probably direct Wnt targets. Previous studies, mostly focused on human tumours, showed partial overlap with our ‘expression signature’, but 37 genes were unique to our study, including TACSTD2, SEMA3F, HOXA9 and IER3 (up-regulated), and TAGLN, GREM1, GREM2, MAB21L2 and RARRES2 (down-regulated). Combined analysis of our and published human data identified additional genes differentially expressed in adenomas, including decreased BMPs (bone morphogenetic proteins) and increased BUB1/BUB1B. Several of the newly identified, differentially expressed genes represent potential diagnostic or therapeutic targets for intestinal tumours. PMID:18782851

  17. Attenuated APC alleles produce functional protein from internal translation initiation

    PubMed Central

    Heppner Goss, Kathleen; Trzepacz, Chris; Tuohy, Thérèse M. F.; Groden, Joanna

    2002-01-01

    Some truncating mutations of the APC tumor suppressor gene are associated with an attenuated phenotype of familial adenomatous polyposis coli (AAPC). This work demonstrates that APC alleles with 5′ mutations produce APC protein that down-regulates β-catenin, inhibits β-catenin/T cell factor-mediated transactivation, and induces cell-cycle arrest. Transfection studies demonstrate that cap-independent translation is initiated internally at an AUG at codon 184 of APC. Furthermore, APC coding sequence between AAPC mutations and AUG 184 permits internal ribosome entry in a bicistronic vector. These data suggest that AAPC alleles in vivo may produce functional APC by internal initiation and establish a functional correlation between 5′ APC mutations and their associated clinical phenotype. PMID:12034871

  18. Industrial strength lithography APC

    NASA Astrophysics Data System (ADS)

    Ausschnitt, Christopher P.; Barker, Brian; Muth, William A.; Postiglione, Marc; Walentosky, Thomas

    2003-06-01

    Fully automated semiconductor manufacturing, becoming a reality with the ramping of 300mm fabricators throughout the world, demands the integration of advanced process control (APC). APC is particularly critical for the lithography sector, whose performance correlates to yield and whose productivity often gates the line. We describe the implementation of a comprehensive lithography APC system at the IBM Center for Nanoelectronics, a 300mm manufacturing and development facility. The base lithography APC function encompasses closed-loop run-to-run control of exposure tool inputs to sustain the overlay and critical dimension outputs consistent with product specifications. Automation demands that no decision regarding the appropriate exposure tool run-time settings be left to human judgment. For each lot, the APC system provides optimum settings based on existing data derived from pertinent process streams. In the case where insufficient prior data exists, the APC system either invokes the appropriate combination of send ahead processing and/or pre-determined defaults. We give specific examples of the application of APC to stitched field and dose control, and quantify its technical benefits. Field matching < 0.1 ppm and critical dimension control < 2.5% is achieved among multiple exposure tools and masks.

  19. Analysis of APC mutation in human ameloblastoma and clinical significance.

    PubMed

    Li, Ning; Liu, Bing; Sui, Chengguang; Jiang, Youhong

    2016-01-01

    As a highly conserved signaling pathway, Wnt/β-catenin signal transduction pathway plays an important role in many processes. Either in the occurrence or development of tumor, activation of this pathway takes an important place. APC inhibits Wnt/β-catenin pathway to regulate cell proliferation and differentiation. This study aimed to investigate the function of cancer suppressor gene. PCR amplification and sequencing method was used to analyze APC mutations of human clinical specimens. The pathological specimens were collected for PCR and clear electrophoretic bands were obtained after electrophoresis. The gene sequence obtained after purification and sequencing analysis was compared with the known APC gene sequence (NM_000038.5). Base mutations at APC 1543 (T → C), APC-4564 (G → A), APC-5353 (T → G), APC-5550 (T → A) and APC-5969 (G → A) locus existed in 22 (27.5 %), 12 (15 %), 5 (6.25 %), 13 (16.25 %) and 12 patients (15 %), respectively. Gene mutations existed in ameloblastoma, and the mutation loci were 1543 locus (T → C), 4564 locus (G → A), 5353 locus (T → G), 5550 locus (T → A) and 5969 locus (G → A) 15 %, respectively. APC mutation plays a certain role in monitoring the tumor malignant degree as it may indicate the transition process of ameloblastoma malignant phenotype. PMID:27065015

  20. Comparison Study of MS-HRM and Pyrosequencing Techniques for Quantification of APC and CDKN2A Gene Methylation

    PubMed Central

    Migheli, Francesca; Stoccoro, Andrea; Coppedè, Fabio; Wan Omar, Wan Adnan; Failli, Alessandra; Consolini, Rita; Seccia, Massimo; Spisni, Roberto; Miccoli, Paolo; Mathers, John C.; Migliore, Lucia

    2013-01-01

    There is increasing interest in the development of cost-effective techniques for the quantification of DNA methylation biomarkers. We analyzed 90 samples of surgically resected colorectal cancer tissues for APC and CDKN2A promoter methylation using methylation sensitive-high resolution melting (MS-HRM) and pyrosequencing. MS-HRM is a less expensive technique compared with pyrosequencing but is usually more limited because it gives a range of methylation estimates rather than a single value. Here, we developed a method for deriving single estimates, rather than a range, of methylation using MS-HRM and compared the values obtained in this way with those obtained using the gold standard quantitative method of pyrosequencing. We derived an interpolation curve using standards of known methylated/unmethylated ratio (0%, 12.5%, 25%, 50%, 75%, and 100% of methylation) to obtain the best estimate of the extent of methylation for each of our samples. We observed similar profiles of methylation and a high correlation coefficient between the two techniques. Overall, our new approach allows MS-HRM to be used as a quantitative assay which provides results which are comparable with those obtained by pyrosequencing. PMID:23326336

  1. Methylated APC and GSTP1 genes in serum DNA correlate with the presence of circulating blood tumor cells and are associated with a more aggressive and advanced breast cancer disease

    PubMed Central

    2010-01-01

    Background Tumor-related methylated DNA and circulating tumor cells (CTC) in the peripheral blood might be of prognostic importance in breast cancer. Thus, the aim of our study was to examine free methylated DNA and CTC in the blood from breast cancer patients and to correlate it with clinicopathological features known to influence prognosis. Materials and methods We prospectively obtained serum samples from 85 patients with breast cancer and 22 healthy volunteers. Sera were analysed by methylation specific PCR (MethyLight PCR) for five genes: adenomatous polyposis coli (APC), ras association domain family protein 1A (RASSF1A), estrogen receptor 1 (ESR1), CDKN2A (p16) and glutathione s-transferase pi 1 (GSTP1). Beta actin (ACTB) served as control. In parallel matched peripheral blood of 63 patients was used to assay for circulating tumor cells in the peripheral blood by a modified immunomagnetic AdnaTest BreastCancerSelect with PCR detection for EPCAM, MUC1, MGB1 and SPDEF. Results We found a hypermethylation in the APC gene in 29% (25/85), in RASSF1A in 26% (22/85), in GSTP1 in 18% (14/76) and in ESR1 in 38% (32/85) of all breast cancer patients. No hypermethylation of CDKN2A was found (0/25). Blood samples of patients were defined CTC positive by detecting the EPCAM 13% (8/63), MUC1 16% (10/63), MGB 9% (5/55), SPDEF 12% (7/58) and in 27% detecting one or more genes (15/55). A significant difference was seen in methylated APC DNA between cancer patients and healthy volunteers. Moreover, methylated APC, RASSF1 and CTC were significantly different in metastatic versus non-metastatic disease. In addition, the presence of methylated APC, RASSF1A and CTC correlated significantly with AJCC-staging (p = 0.001, p = 0.031 and 0.002, respectively). High incidences of methylations were found for the genes RASSF1 and ESR1 in healthy individuals (both 23% 5/22). Methylated GSTP1 was predominantly found in the serum of patients with large primaries (p = 0.023) and was highly

  2. A multigene mutation classification of 468 colorectal cancers reveals a prognostic role for APC

    PubMed Central

    Schell, Michael J.; Yang, Mingli; Teer, Jamie K.; Lo, Fang Yin; Madan, Anup; Coppola, Domenico; Monteiro, Alvaro N. A.; Nebozhyn, Michael V.; Yue, Binglin; Loboda, Andrey; Bien-Willner, Gabriel A.; Greenawalt, Danielle M.; Yeatman, Timothy J.

    2016-01-01

    Colorectal cancer (CRC) is a highly heterogeneous disease, for which prognosis has been relegated to clinicopathologic staging for decades. There is a need to stratify subpopulations of CRC on a molecular basis to better predict outcome and assign therapies. Here we report targeted exome-sequencing of 1,321 cancer-related genes on 468 tumour specimens, which identified a subset of 17 genes that best classify CRC, with APC playing a central role in predicting overall survival. APC may assume 0, 1 or 2 truncating mutations, each with a striking differential impact on survival. Tumours lacking any APC mutation carry a worse prognosis than single APC mutation tumours; however, two APC mutation tumours with mutant KRAS and TP53 confer the poorest survival among all the subgroups examined. Our study demonstrates a prognostic role for APC and suggests that sequencing of APC may have clinical utility in the routine staging and potential therapeutic assignment for CRC. PMID:27302369

  3. A multigene mutation classification of 468 colorectal cancers reveals a prognostic role for APC.

    PubMed

    Schell, Michael J; Yang, Mingli; Teer, Jamie K; Lo, Fang Yin; Madan, Anup; Coppola, Domenico; Monteiro, Alvaro N A; Nebozhyn, Michael V; Yue, Binglin; Loboda, Andrey; Bien-Willner, Gabriel A; Greenawalt, Danielle M; Yeatman, Timothy J

    2016-01-01

    Colorectal cancer (CRC) is a highly heterogeneous disease, for which prognosis has been relegated to clinicopathologic staging for decades. There is a need to stratify subpopulations of CRC on a molecular basis to better predict outcome and assign therapies. Here we report targeted exome-sequencing of 1,321 cancer-related genes on 468 tumour specimens, which identified a subset of 17 genes that best classify CRC, with APC playing a central role in predicting overall survival. APC may assume 0, 1 or 2 truncating mutations, each with a striking differential impact on survival. Tumours lacking any APC mutation carry a worse prognosis than single APC mutation tumours; however, two APC mutation tumours with mutant KRAS and TP53 confer the poorest survival among all the subgroups examined. Our study demonstrates a prognostic role for APC and suggests that sequencing of APC may have clinical utility in the routine staging and potential therapeutic assignment for CRC. PMID:27302369

  4. Interaction between APC and Fen1 during breast carcinogenesis.

    PubMed

    Narayan, Satya; Jaiswal, Aruna S; Law, Brian K; Kamal, Mohammad A; Sharma, Arun K; Hromas, Robert A

    2016-05-01

    Aberrant DNA base excision repair (BER) contributes to malignant transformation. However, inter-individual variations in DNA repair capacity plays a key role in modifying breast cancer risk. We review here emerging evidence that two proteins involved in BER - adenomatous polyposis coli (APC) and flap endonuclease 1 (Fen1) - promote the development of breast cancer through novel mechanisms. APC and Fen1 expression and interaction is increased in breast tumors versus normal cells, APC interacts with and blocks Fen1 activity in Pol-β-directed LP-BER, and abrogation of LP-BER is linked with cigarette smoke condensate-induced transformation of normal breast epithelial cells. Carcinogens increase expression of APC and Fen1 in spontaneously immortalized human breast epithelial cells, human colon cancer cells, and mouse embryonic fibroblasts. Since APC and Fen1 are tumor suppressors, an increase in their levels could protect against carcinogenesis; however, this does not seem to be the case. Elevated Fen1 levels in breast and lung cancer cells may reflect the enhanced proliferation of cancer cells or increased DNA damage in cancer cells compared to normal cells. Inactivation of the tumor suppressor functions of APC and Fen1 is due to their interaction, which may act as a susceptibility factor for breast cancer. The increased interaction of APC and Fen1 may occur due to polypmorphic and/or mutational variation in these genes. Screening of APC and Fen1 polymorphic and/or mutational variations and APC/Fen1 interaction may permit assessment of individual DNA repair capability and the risk for breast cancer development. Such individuals might lower their breast cancer risk by reducing exposure to carcinogens. Stratifying individuals according to susceptibility would greatly assist epidemiologic studies of the impact of suspected environmental carcinogens. Additionally, a mechanistic understanding of the interaction of APC and Fen1 may provide the basis for developing new and

  5. The APC tumor suppressor is required for epithelial cell polarization and three-dimensional morphogenesis.

    PubMed

    Lesko, Alyssa C; Goss, Kathleen H; Yang, Frank F; Schwertner, Adam; Hulur, Imge; Onel, Kenan; Prosperi, Jenifer R

    2015-03-01

    The Adenomatous Polyposis Coli (APC) tumor suppressor has been previously implicated in the control of apical-basal polarity; yet, the consequence of APC loss-of-function in epithelial polarization and morphogenesis has not been characterized. To test the hypothesis that APC is required for the establishment of normal epithelial polarity and morphogenesis programs, we generated APC-knockdown epithelial cell lines. APC depletion resulted in loss of polarity and multi-layering on permeable supports, and enlarged, filled spheroids with disrupted polarity in 3D culture. Importantly, these effects of APC knockdown were independent of Wnt/β-catenin signaling, but were rescued with either full-length or a carboxy (c)-terminal segment of APC. Moreover, we identified a gene expression signature associated with APC knockdown that points to several candidates known to regulate cell-cell and cell-matrix communication. Analysis of epithelial tissues from mice and humans carrying heterozygous APC mutations further supports the importance of APC as a regulator of epithelial behavior and tissue architecture. These data also suggest that the initiation of epithelial-derived tumors as a result of APC mutation or gene silencing may be driven by loss of polarity and dysmorphogenesis. PMID:25578398

  6. Intestinal flora of FAP patients containing APC-like sequences.

    PubMed

    Hainova, K; Adamcikova, Z; Ciernikova, S; Stevurkova, V; Tyciakova, S; Zajac, V

    2014-01-01

    Colorectal cancer mortality is one of the most common cause of cancer-related mortality. A multiple risk factors are associated with colorectal cancer, including hereditary, enviromental and inflammatory syndromes affecting the gastrointestinal tract. Familial adenomatous polyposis (FAP) is characterized by the emergence of hundreds to thousands of colorectal adenomatous polyps and FAP syndrome is caused by mutations within the adenomatous polyposis coli (APC) tumor suppressor gene. We analyzed 21 rectal bacterial subclones isolated from FAP patient 41-1 with confirmed 5bp ACAAA deletion within codons 1060-1063 for the presence of APC-like sequences in longest exon 15. The studied section was defined by primers 15Efor-15Erev, what correlates with mutation cluster region (MCR) in which the 75% of all APC germline mutations were detected. More than 90% homology was showed by sequencing and subsequent software comparison. The expression of APC-like sequences was demostrated by Western blot analysis using monoclonal and polyclonal antibodies against APC protein. To study missing link between the DNA analysis (PCR, DNA sequencing) and protein expresion experiments (Western blotting) we analyzed bacterial transcripts containing the 15Efor-15Erev sequence of APC gene by reverse transcription-PCR, what indicated that an APC gene derived fragment may be produced. We observed 97-100 % homology after computer comparison of cDNA PCR products. Our results suggest that presence of APC-like sequences in intestinal/rectal bacteria is enrichment of bacterial genetic information in which horizontal gene transfer between humans and microflora play an important role. PMID:24824929

  7. Functional Comparison of Human Adenomatous Polyposis Coli (APC) and APC-Like in Targeting Beta-Catenin for Degradation

    PubMed Central

    Schneikert, Jean; Vijaya Chandra, Shree Harsha; Ruppert, Jan Gustav; Ray, Suparna; Wenzel, Eva Maria; Behrens, Jürgen

    2013-01-01

    Truncating mutations affect the adenomatous polyposis coli (APC) gene in most cases of colon cancer, resulting in the stabilization of β-catenin and uncontrolled cell proliferation. We show here that colon cancer cell lines express also the paralog APC-like (APCL or APC2). RNA interference revealed that it controls the level and/or the activity of β-catenin, but it is less efficient and binds less well to β-catenin than APC, thereby providing one explanation as to why the gene is not mutated in colon cancer. A further comparison indicates that APCL down-regulates the β-catenin level despite the lack of the 15R region known to be important in APC. To understand this discrepancy, we performed immunoprecipitation experiments that revealed that phosphorylated β-catenin displays a preference for binding to the 15 amino acid repeats (15R) rather than the first 20 amino acid repeat of APC. This suggests that the 15R region constitutes a gate connecting the steps of β-catenin phosphorylation and subsequent ubiquitination/degradation. Using RNA interference and domain swapping experiments, we show that APCL benefits from the 15R of truncated APC to target β-catenin for degradation, in a process likely involving heterodimerization of the two partners. Our data suggest that the functional complementation of APCL by APC constitutes a substantial facet of tumour development, because the truncating mutations of APC in colorectal tumours from familial adenomatous polyposis (FAP) patients are almost always selected for the retention of at least one 15R. PMID:23840886

  8. The Anaphase-Promoting Complex (APC) ubiquitin ligase affects chemosensory behavior in C. elegans

    PubMed Central

    Wang, Julia; Jennings, Alexandra K.

    2016-01-01

    The regulation of fundamental aspects of neurobiological function has been linked to the ubiquitin signaling system (USS), which regulates the degradation and activity of proteins and is catalyzed by E1, E2, and E3 enzymes. The Anaphase-Promoting Complex (APC) is a multi-subunit E3 ubiquitin ligase that controls diverse developmental and signaling processes in post-mitotic neurons; however, potential roles for the APC in sensory function have yet to be explored. In this study, we examined the effect of the APC ubiquitin ligase on chemosensation in Caenorhabditis elegans by testing chemotaxis to the volatile odorants, diacetyl, pyrazine, and isoamyl alcohol, to which wild-type worms are attracted. Animals with loss of function mutations in either of two alleles (g48 and ye143) of the gene encoding the APC subunit EMB-27 APC6 showed increased chemotaxis towards diacetyl and pyrazine, odorants sensed by AWA neurons, but exhibited normal chemotaxis to isoamyl alcohol, which is sensed by AWC neurons. The statistically significant increase in chemotaxis in the emb-27 APC6 mutants suggests that the APC inhibits AWA-mediated chemosensation in C. elegans. Increased chemotaxis to pyrazine was also seen with mutants lacking another essential APC subunit, MAT-2 APC1; however, mat-2 APC1 mutants exhibited wild type responses to diacetyl. The difference in responsiveness of these two APC subunit mutants may be due to differential strength of these hypomorphic alleles or may indicate the presence of functional sub-complexes of the APC at work in this process. These findings are the first evidence for APC-mediated regulation of chemosensation and lay the groundwork for further studies aimed at identifying the expression levels, function, and targets of the APC in specific sensory neurons. Because of the similarity between human and C. elegans nervous systems, the role of the APC in sensory neurons may also advance our understanding of human sensory function and disease. PMID

  9. The Anaphase-Promoting Complex (APC) ubiquitin ligase affects chemosensory behavior in C. elegans.

    PubMed

    Wang, Julia; Jennings, Alexandra K; Kowalski, Jennifer R

    2016-01-01

    The regulation of fundamental aspects of neurobiological function has been linked to the ubiquitin signaling system (USS), which regulates the degradation and activity of proteins and is catalyzed by E1, E2, and E3 enzymes. The Anaphase-Promoting Complex (APC) is a multi-subunit E3 ubiquitin ligase that controls diverse developmental and signaling processes in post-mitotic neurons; however, potential roles for the APC in sensory function have yet to be explored. In this study, we examined the effect of the APC ubiquitin ligase on chemosensation in Caenorhabditis elegans by testing chemotaxis to the volatile odorants, diacetyl, pyrazine, and isoamyl alcohol, to which wild-type worms are attracted. Animals with loss of function mutations in either of two alleles (g48 and ye143) of the gene encoding the APC subunit EMB-27 APC6 showed increased chemotaxis towards diacetyl and pyrazine, odorants sensed by AWA neurons, but exhibited normal chemotaxis to isoamyl alcohol, which is sensed by AWC neurons. The statistically significant increase in chemotaxis in the emb-27 APC6 mutants suggests that the APC inhibits AWA-mediated chemosensation in C. elegans. Increased chemotaxis to pyrazine was also seen with mutants lacking another essential APC subunit, MAT-2 APC1; however, mat-2 APC1 mutants exhibited wild type responses to diacetyl. The difference in responsiveness of these two APC subunit mutants may be due to differential strength of these hypomorphic alleles or may indicate the presence of functional sub-complexes of the APC at work in this process. These findings are the first evidence for APC-mediated regulation of chemosensation and lay the groundwork for further studies aimed at identifying the expression levels, function, and targets of the APC in specific sensory neurons. Because of the similarity between human and C. elegans nervous systems, the role of the APC in sensory neurons may also advance our understanding of human sensory function and disease. PMID

  10. Nuclear-cytoplasmic shuttling of APC regulates beta-catenin subcellular localization and turnover.

    PubMed

    Henderson, B R

    2000-09-01

    Mutational inactivation of the APC gene is a key early event in the development of familial adenomatous polyposis and colon cancer. APC suppresses tumour progression by promoting degradation of the oncogenic transcriptional activator beta-catenin. APC gene mutations can lead to abnormally high levels of beta-catenin in the nucleus, and the consequent activation of transforming genes. Here, we show that APC is a nuclear-cytoplasmic shuttling protein, and that it can function as a beta-catenin chaperone. APC contains two active nuclear export sequences (NES) at the amino terminus, and mutagenesis of these conserved motifs blocks nuclear export dependent on the CRM1 export receptor. Treatment of cells with the CRM1-specific export inhibitor leptomycin B shifts APC from cytoplasm to nucleus. beta-catenin localization is also regulated by CRM1, but in an APC-dependent manner. Transient expression of wild-type APC in SW480 (APCmut/mut) colon cancer cells enhances nuclear export and degradation of beta-catenin, and these effects can be blocked by mutagenesis of the APC NES. These findings suggest that wild-type APC controls the nuclear accumulation of beta-catenin by a combination of nuclear export and cytoplasmic degradation. PMID:10980707

  11. The effect of 1,25 dihydroxyvitamin D3 treatment on the mRNA levels of β catenin target genes in mice with colonic inactivation of both APC alleles

    PubMed Central

    DeWitt, Marsha; Johnson, Robert L.; Snyder, Paul; Fleet, James C.

    2015-01-01

    In colon cancer, adenomatous polyposis coli (APC) inactivating gene mutations increase nuclear β-catenin levels and stimulate proliferation. In vitro, 1,25 dihydroxyvitamin D (1,25(OH)2D), suppresses β-catenin-mediated gene transcription by inducing vitamin D receptor (VDR)-β-catenin interactions. We examined whether acute treatment with 1,25(OH)2D could suppress β-catenin-mediated gene transcription in the hyperplastic colonic lesions ofmice with colon-specific deletion of both APC gene alleles (CAC; APCΔ580/Δ580). At four weeks of age, CAC; APCΔ580/Δ580 and control mice were injected with vehicle or 1,25(OH)2D (1 μg/kg body weight) once a day for three days and then killed six hours after the last injection. mRNA levels of β-catenin target genes were elevated in the colon of CAC; APCΔ580/Δ580 mice. 1,25(OH)2D increased 25 hydroxyvitamin D-24 hydroxylase mRNA levels in the colon of CAC; APCΔ580/Δ580 and control mice indicating the treatments activated the VDR. However, 1,25(OH)2D had no effect on either β-catenin target gene mRNA levels or the proliferation index in CAC; APCΔ580/Δ580 or control mice. VDR mRNA and protein levels were lower (−65% and −90%) in the colon of CAC; APCΔ580/Δ580 mice compared to control mice, suggesting loss of colon responsiveness to vitamin D. Consistent with this, vitamin D-induced expression of Transient Receptor Potential cation channel, subfamily V, member 6 mRNA was reduced in the colon of CAC; APCΔ580/Δ580 mice. Our data show that short term exposure to 1,25(OH)2D does not suppress colonic β-catenin signaling in vivo. PMID:25597951

  12. A knock-in mouse model reveals roles for nuclear Apc in cell proliferation, Wnt signal inhibition and tumor suppression.

    PubMed

    Zeineldin, M; Cunningham, J; McGuinness, W; Alltizer, P; Cowley, B; Blanchat, B; Xu, W; Pinson, D; Neufeld, K L

    2012-05-10

    Mutation of the tumor suppressor adenomatous polyposis coli (APC) is considered an initiating step in the genesis of the vast majority of colorectal cancers. APC inhibits the Wnt-signaling pathway by targeting the proto-oncogene β-catenin for destruction by cytoplasmic proteasomes. In the presence of a Wnt signal, or in the absence of functional APC, β-catenin can serve as a transcription cofactor for genes required for cell proliferation such as cyclin-D1 and c-Myc. In cultured cells, APC shuttles between the nucleus and the cytoplasm, with nuclear APC implicated in the inhibition of Wnt target gene expression. Adopting a genetic approach to evaluate the functions of nuclear APC in the context of a whole organism, we generated a mouse model with mutations that inactivate the nuclear localization signals (NLSs) of Apc (Apc(mNLS)). Apc(mNLS/mNLS) mice are viable and fractionation of mouse embryonic fibroblasts (MEFs) isolated from these mice revealed a significant reduction in nuclear Apc as compared with Apc(+/+) MEFs. The levels of Apc and β-catenin protein were not significantly altered in small intestinal epithelia from Apc(mNLS/mNLS) mice. Compared with Apc(+/+) mice, Apc(mNLS/mNLS) mice showed increased proliferation in epithelial cells from the jejunum, ileum and colon. These same tissues from Apc(mNLS/mNLS) mice showed more mRNA from three genes upregulated in response to canonical Wnt signal, c-Myc, axin-2 and cyclin-D1, and less mRNA from Hath-1, which is downregulated in response to Wnt. These observations suggest a role for nuclear Apc in the inhibition of canonical Wnt signaling and the control of epithelial proliferation in intestinal tissue. Furthermore, we found Apc(Min/+) mice, which harbor a mutation that truncates Apc, to have an increased polyp size and multiplicity if they also carry the Apc(mNLS) allele. Taken together, this analysis of the novel Apc(mNLS) mouse model supports a role for nuclear Apc in the control of Wnt target genes

  13. Identification of APC mutations and evaluation of their expression level using a functional screening assay

    SciTech Connect

    Varesco, L.; Gismondi, V.; Bafico, A.

    1994-09-01

    A functional screen for chain-terminating mutations in the APC gene recently has been developed. It is based on the PCR and cloning of a segment of the gene in-frame with a colorimetric marker gene (lacz) followed by screening for the level of activity of the marker polypeptide (beta-galactosidase). This method scores colony number with different blue colors that are produced by bacteria containing normal and mutant APC segments. In the present work this method was used to screen the entire APC coding region by using eight primer pairs. DNA segments with known APC mutations at different positions in the gene were used as controls and were clearly identifiable with this assay. In addition, the entire APC coding region has been examined in 21 APC patients in whom PCR-SSCP did not identify an APC mutation. Novel mutations (n=14) were identified by the blue/white assay and were all confirmed by sequence analysis. This method also was used to quantitate the expression of paternal and maternal APC alleles taking advantage of an RsaI site polymorphism at position 1458 in a small number of informative individuals. Differential expression of some known mutant APC mRNAs was observed.

  14. Extensive metabolic disorders are present in APC(min) tumorigenesis mice.

    PubMed

    Liu, Zhenzhen; Xiao, Yi; Zhou, Zhengxiang; Mao, Xiaoxiao; Cai, Jinxing; Xiong, Lu; Liao, Chaonan; Huang, Fulian; Liu, Zehao; Ali Sheikh, Md Sayed; Plutzky, Jorge; Huang, He; Yang, Tianlun; Duan, Qiong

    2016-05-15

    Wnt signaling plays essential role in mesenchymal stem cell (MSC) differentiation. Activation of Wnt signaling suppresses adipogenesis, but promotes osteogenesis in MSC. Adenomatous polyposis coli (APC) is a negative regulator of β-catenin and Wnt signaling activity. The mutation of APC gene leads to the activation of Wnt signaling and is responsible for tumorigenesis in APC(min) mouse; however, very few studies focused on its metabolic abnormalities. The present study reports a widespread metabolic disorder phenotype in APC(min) mice. The old APC(min) mice have decreased body weight and impaired adipogenesis, but severe hyperlipidemia, which mimic the phenotypes of Familial Adenomatous Polyposis (FAP), an inherited disease also caused by APC gene mutation in human. We found that the expression of lipid metabolism and free fat acids (FA) use genes in the white adipose tissue (WAT) of the APC(min) mice is much lower than those of control. The changed gene expression pattern may lead to the disability of circulatory lipid transportation and storage at WAT. Moreover, the APC(min) mice could not maintain the core body temperature in cold condition. PET-CT determination revealed that the BAT of APC(min) mice has significantly impaired ability to take up (18)FDG from the blood. Morphological studies identified that the brown adipocytes of APC(min) mice were filled with lipid droplets but fewer mitochondria. These results matched with the findings of impaired BAT function in APC(min) mice. Collectively, our study explores a new mechanism that explains abnormal metabolism in APC(min) mice and provides insights into studying the metabolic disorders of FAP patients. PMID:26948948

  15. Reduced expression of APC-1B but not APC-1A by the deletion of promoter 1B is responsible for familial adenomatous polyposis.

    PubMed

    Yamaguchi, Kiyoshi; Nagayama, Satoshi; Shimizu, Eigo; Komura, Mitsuhiro; Yamaguchi, Rui; Shibuya, Tetsuo; Arai, Masami; Hatakeyama, Seira; Ikenoue, Tsuneo; Ueno, Masashi; Miyano, Satoru; Imoto, Seiya; Furukawa, Yoichi

    2016-01-01

    Germline mutations in the tumor suppressor gene APC are associated with familial adenomatous polyposis (FAP). Here we applied whole-genome sequencing (WGS) to the DNA of a sporadic FAP patient in which we did not find any pathological APC mutations by direct sequencing. WGS identified a promoter deletion of approximately 10 kb encompassing promoter 1B and exon1B of APC. Additional allele-specific expression analysis by deep cDNA sequencing revealed that the deletion reduced the expression of the mutated APC allele to as low as 11.2% in the total APC transcripts, suggesting that the residual mutant transcripts were driven by other promoter(s). Furthermore, cap analysis of gene expression (CAGE) demonstrated that the deleted promoter 1B region is responsible for the great majority of APC transcription in many tissues except the brain. The deletion decreased the transcripts of APC-1B to 39-45% in the patient compared to the healthy controls, but it did not decrease those of APC-1A. Different deletions including promoter 1B have been reported in FAP patients. Taken together, our results strengthen the evidence that analysis of structural variations in promoter 1B should be considered for the FAP patients whose pathological mutations are not identified by conventional direct sequencing. PMID:27217144

  16. Reduced expression of APC-1B but not APC-1A by the deletion of promoter 1B is responsible for familial adenomatous polyposis

    PubMed Central

    Yamaguchi, Kiyoshi; Nagayama, Satoshi; Shimizu, Eigo; Komura, Mitsuhiro; Yamaguchi, Rui; Shibuya, Tetsuo; Arai, Masami; Hatakeyama, Seira; Ikenoue, Tsuneo; Ueno, Masashi; Miyano, Satoru; Imoto, Seiya; Furukawa, Yoichi

    2016-01-01

    Germline mutations in the tumor suppressor gene APC are associated with familial adenomatous polyposis (FAP). Here we applied whole-genome sequencing (WGS) to the DNA of a sporadic FAP patient in which we did not find any pathological APC mutations by direct sequencing. WGS identified a promoter deletion of approximately 10 kb encompassing promoter 1B and exon1B of APC. Additional allele-specific expression analysis by deep cDNA sequencing revealed that the deletion reduced the expression of the mutated APC allele to as low as 11.2% in the total APC transcripts, suggesting that the residual mutant transcripts were driven by other promoter(s). Furthermore, cap analysis of gene expression (CAGE) demonstrated that the deleted promoter 1B region is responsible for the great majority of APC transcription in many tissues except the brain. The deletion decreased the transcripts of APC-1B to 39–45% in the patient compared to the healthy controls, but it did not decrease those of APC-1A. Different deletions including promoter 1B have been reported in FAP patients. Taken together, our results strengthen the evidence that analysis of structural variations in promoter 1B should be considered for the FAP patients whose pathological mutations are not identified by conventional direct sequencing. PMID:27217144

  17. Atomic-Resolution Structures of the APC/C Subunits Apc4 and the Apc5 N-Terminal Domain

    PubMed Central

    Cronin, Nora B.; Yang, Jing; Zhang, Ziguo; Kulkarni, Kiran; Chang, Leifu; Yamano, Hiroyuki; Barford, David

    2015-01-01

    Many essential biological processes are mediated by complex molecular machines comprising multiple subunits. Knowledge on the architecture of individual subunits and their positions within the overall multimeric complex is key to understanding the molecular mechanisms of macromolecular assemblies. The anaphase-promoting complex/cyclosome (APC/C) is a large multisubunit complex that regulates cell cycle progression by ubiquitinating cell cycle proteins for proteolysis by the proteasome. The holo-complex is composed of 15 different proteins that assemble to generate a complex of 20 subunits. Here, we describe the crystal structures of Apc4 and the N-terminal domain of Apc5 (Apc5N). Apc4 comprises a WD40 domain split by a long α-helical domain, whereas Apc5N has an α-helical fold. In a separate study, we had fitted these atomic models to a 3.6-Å-resolution cryo-electron microscopy map of the APC/C. We describe how, in the context of the APC/C, regions of Apc4 disordered in the crystal assume order through contacts to Apc5, whereas Apc5N shows small conformational changes relative to its crystal structure. We discuss the complementary approaches of high-resolution electron microscopy and protein crystallography to the structure determination of subunits of multimeric complexes. PMID:26343760

  18. APC15 mediates CDC20 auto-ubiquitylation by APC/CMCC and MCC disassembly

    PubMed Central

    Uzunova, Kristina; Dye, Billy T.; Schutz, Hannelore; Ladurner, Rene; Petzold, Georg; Toyoda, Yusuke; Jarvis, Marc A.; Brown, Nicholas G.; Poser, Ina; Novatchkova, Maria; Mechtler, Karl; Hyman, Anthony A.; Stark, Holger; Schulman, Brenda A.; Peters, Jan-Michael

    2012-01-01

    The anaphase-promoting complex/cyclosome bound to CDC20 (APC/CCDC20) initiates anaphase by ubiquitylating B-type cyclins and securin. During chromosome bi-orientation, CDC20 assembles with MAD2, BUBR1 and BUB3 into a mitotic checkpoint complex (MCC) which inhibits substrate recruitment to the APC/C. APC/C activation depends on MCC disassembly, which has been proposed to require CDC20 auto-ubiquitylation. Here we characterized APC15, a human APC/C subunit related to yeast Mnd2. APC15 is located near APC/C’s MCC binding site, is required for APC/CMCC-dependent CDC20 auto-ubiquitylation and degradation, and for timely anaphase initiation, but is dispensable for substrate ubiquitylation by APC/CCDC20 and APC/CCDH1. Our results support the view that MCC is continuously assembled and disassembled to enable rapid activation of APC/CCDC20 and that CDC20 auto-ubiquitylation promotes MCC disassembly. We propose that APC15 and Mnd2 negatively regulate APC/C coactivators, and report the first generation of recombinant human APC/C. PMID:23007861

  19. Structure of an APC3-APC16 complex: Insights into assembly of the Anaphase Promoting Complex/Cyclosome

    PubMed Central

    Yamaguchi, Masaya; Yu, Shanshan; Qiao, Renping; Weissmann, Florian; Miller, Darcie J.; VanderLinden, Ryan; Brown, Nicholas G.; Frye, Jeremiah J.; Peters, Jan-Michael; Schulman, Brenda A.

    2015-01-01

    The Anaphase Promoting Complex/Cyclosome (APC/C) is a massive E3 ligase that controls mitosis by catalyzing ubiquitination of key cell cycle regulatory proteins. The APC/C assembly contains two subcomplexes: the “Platform” centers around a cullin-RING-like E3 ligase catalytic core; the “Arc Lamp” is a hub that mediates transient association with regulators and ubiquitination substrates. The Arc Lamp contains the small subunits APC16, CDC26, and APC13, and tetratricopeptide repeat (TPR) proteins (APC7, APC3, APC6, and APC8) that homodimerize and stack with quasi-twofold symmetry. Within the APC/C complex, APC3 serves as center for regulation. APC3’s TPR motifs recruit substrate-binding coactivators, CDC20 and CDH1, via their C-terminal conserved Ile-Arg (IR) tail sequences. Human APC3 also binds APC16 and APC7, and contains a >200-residue loop that is heavily phosphorylated during mitosis, although the basis for APC3 interactions and whether loop phosphorylation is required for ubiquitination are unclear. Here, we map the basis for human APC3 assembly with APC16 and APC7, report crystal structures of APC3Δloop alone and in complex with the C-terminal domain of APC16, and test roles of APC3’s loop and IR-tail binding surfaces in APC/C-catalyzed ubiquitination. The structures show how one APC16 binds asymmetrically to the symmetric APC3 dimer, and together with biochemistry and prior data explain how APC16 recruits APC7 to APC3, show how APC3’s C-terminal domain is rearranged in the full APC/C assembly, and visualize residues in the IR-tail binding cleft important for coactivator-dependent ubiquitination. Overall, the results provide insights into assembly, regulation, and interactions of TPR proteins and the APC/C. PMID:25490258

  20. Loss of Rassf1a co-operates with ApcMin to accelerate intestinal tumourigenesis

    PubMed Central

    van der Weyden, L.; Arends, M.J.; Dovey, O.M.; Harrison, H. L.; Lefebvre, G.; Conte, N.; Gergely, F.V.; Bradley, A.; Adams, D.J.

    2013-01-01

    Promoter methylation of the RAS-association domain family 1, isoform A gene (RASSF1A) is one of the most frequent events found in human tumours. In this study we set out to test the hypothesis that loss of Rassf1a can co-operate with inactivation of the adenomatous polyposis coli (Apc) gene to accelerate intestinal tumourigenesis using the Apc-Min (ApcMin/+) mouse model, as mutational or deletional inactivation of APC is a frequent early event in the genesis of intestinal cancer. Further, loss of RASSF1A has also been reported to occur in premalignant adenomas of the bowel. RASSF1A has been implicated in an array of pivotal cellular processes, including regulation of the cell cycle, apoptosis, microtubule stability and most recently in the β-catenin signalling pathway. By interbreeding isoform specific Rassf1a knockout mice with Apc+/Min mice we showed that loss of Rassf1a results in a significant increase in adenomas of the small intestine and accelerated intestinal tumourigenesis leading to the earlier death of adenocarcinoma-bearing mice and decreased overall survival. Comparative genomic hybridization of adenomas from Rassf1a−/−; Apc+/Min mice revealed no evidence of aneuploidy or gross chromosomal instability (no difference to adenomas from Rassf1a+/+; Apc+/Min mice). Immunohistochemical analysis of adenomas revealed increased nuclear β-catenin accumulation in adenomas from Rassf1a−/−; Apc+/Min mice, compared to those from Rassf1a+/+; Apc+/Min mice, but no differences in proliferation marker (Ki67) staining patterns. Collectively these data demonstrate co-operation between inactivation of Rassf1a and Apc resulting in accelerated intestinal tumourigenesis, with adenomas showing increased nuclear accumulation of β-catenin, supporting a mechanistic link via loss of the known interaction of Rassf1 with β-TrCP that usually mediates degradation of β-catenin. PMID:18391979

  1. APC mutations in colorectal tumors with mismatch repair deficiency.

    PubMed Central

    Huang, J; Papadopoulos, N; McKinley, A J; Farrington, S M; Curtis, L J; Wyllie, A H; Zheng, S; Willson, J K; Markowitz, S D; Morin, P; Kinzler, K W; Vogelstein, B; Dunlop, M G

    1996-01-01

    We have investigated the influence of genetic instability [replication error (RER) phenotype] on APC (adenomatous polyposis coli), a gene thought to initiate colorectal tumorigenesis. The prevalence of APC mutations was similar in RER and non-RER tumors, indicating that both tumor types share this step in neoplastic transformation. However, in a total of 101 sequenced mutations, we noted a substantial excess of APC frameshift mutations in the RER cases (70% in RER tumors versus 47% in non-RER tumors, P < 0.04). These frameshifts were characteristic of mutations arising in cells deficient in DNA mismatch repair, with a predilection for mononucleotide repeats in the RER tumors (P < 0.0002), particularly (A)n tracts (P < 0.00007). These findings suggest that the genetic instability that is reflected by the RER phenotype precedes, and is responsible for, APC mutation in RER large bowel tumors and have important implications for understanding the very earliest stages of neoplasia in patients with tumors deficient in mismatch repair. Images Fig. 2 PMID:8799152

  2. Prostate Cancer Induced by Loss of Apc Is Restrained by TGFβ Signaling

    PubMed Central

    Bjerke, Glen A.; Pietrzak, Karolina; Melhuish, Tiffany A.; Frierson Jr., Henry F.; Paschal, Bryce M.; Wotton, David

    2014-01-01

    Recent work with mouse models of prostate cancer (CaP) has shown that inactivation of TGFβ signaling in prostate epithelium can cooperate with deletion of the Pten tumor suppressor to drive locally aggressive cancer and metastatic disease. Here, we show that inactivating the TGFβ pathway by deleting the gene encoding the TGFβ type II receptor (Tgfbr2) in combination with a deletion of the Apc tumor suppressor gene specifically in mouse prostate epithelium, results in the rapid onset of invasive CaP. Micro-metastases were observed in the lymph nodes and lungs of a proportion of the double mutant mice, whereas no metastases were observed in Apc single mutant mice. Prostate-specific Apc;Tgfbr2 mutants had a lower frequency of metastasis and survived significantly longer than Pten;Tgfbr2 double mutants. However, all Apc;Tgfbr2 mutants developed invasive cancer by 30 weeks of age, whereas invasive cancer was rarely observed in Apc single mutant animals, even by one year of age. Further comparison of the Pten and Apc models of CaP revealed additional differences, including adenosquamous carcinoma in the Apc;Tgfbr2 mutants that was not seen in the Pten model, and a lack of robust induction of the TGFβ pathway in Apc null prostate. In addition to causing high-grade prostate intra-epithelial neoplasia (HGPIN), deletion of either Pten or Apc induced senescence in affected prostate ducts, and this restraint was overcome by loss of Tgfbr2. In summary, this work demonstrates that TGFβ signaling restrains the progression of CaP induced by different tumor suppressor mutations, suggesting that TGFβ signaling exerts a general tumor suppressive effect in prostate. PMID:24651496

  3. Parafibromin and APC as screening markers for malignant potential in atypical parathyroid adenomas.

    PubMed

    Juhlin, C Christofer; Nilsson, Inga-Lena; Johansson, Kenth; Haglund, Felix; Villablanca, Andrea; Höög, Anders; Larsson, Catharina

    2010-09-01

    The identification of parathyroid carcinomas is based upon histopathological criteria in which an invasive growth pattern or distant metastasis is demonstrated. A dilemma arises when tumours present with atypical histopathological features but lack direct evidence of malignancy. Recently, reduced expression or loss of the tumour suppressor proteins parafibromin and adenomatous polyposis coli (APC) has been associated with parathyroid malignancy. We report results from APC and parafibromin expression analyses by immunohistochemistry and Western blot in five cases of atypical adenoma, a single case of carcinoma and 54 adenomas without atypical features. Complete loss of APC immunoreactivity and reduced expression of parafibromin was evident in two of the atypical adenomas and in the parathyroid carcinoma. By contrast, all adenomas displayed APC expression, including two cases with hyperparathyroidism 2 gene (HRPT2) mutations and loss of parafibromin expression. We conclude that loss of APC is a frequent molecular event in atypical adenomas and carcinomas, but not in adenomas. Following verification in an independent material, APC could become a valuable tool when assessing parathyroid tumours in the clinical setting. Furthermore, the molecular resemblance of atypical adenomas with carcinoma concerning parafibromin and APC expression indicates that atypical adenomas should be subjects to watchful follow-up. PMID:20473645

  4. Targeting the DNA replication checkpoint by pharmacologic inhibition of Chk1 kinase: a strategy to sensitize APC mutant colon cancer cells to 5-fluorouracil chemotherapy

    PubMed Central

    Martino-Echarri, Estefania

    2014-01-01

    5-fluorouracil (5-FU) is the first line component used in colorectal cancer (CRC) therapy however even in combination with other chemotherapeutic drugs recurrence is common. Mutations of the adenomatous polyposis coli (APC) gene are considered as the initiating step of transformation in familial and sporadic CRCs. We have previously shown that APC regulates the cellular response to DNA replication stress and recently hypothesized that APC mutations might therefore influence 5-FU resistance. To test this, we compared CRC cell lines and show that those expressing truncated APC exhibit a limited response to 5-FU and arrest in G1/S-phase without undergoing lethal damage, unlike cells expressing wild-type APC. In SW480 APC-mutant CRC cells, 5-FU-dependent apoptosis was restored after transient expression of full length APC, indicating a direct link between APC and drug response. Furthermore, we could increase sensitivity of APC truncated cells to 5-FU by inactivating the Chk1 kinase using drug treatment or siRNA-mediated knockdown. Our findings identify mutant APC as a potential tumor biomarker of resistance to 5-FU, and importantly we show that APC-mutant CRC cells can be made more sensitive to 5-FU by use of Chk1 inhibitors. PMID:25301724

  5. Adenomatous Polyposis Coli (APC) Is Required for Normal Development of Skin and Thymus

    PubMed Central

    Kuraguchi, Mari; Wang, Xiu-Ping; Bronson, Roderick T; Rothenberg, Rebecca; Ohene-Baah, Nana Yaw; Lund, Jennifer J; Kucherlapati, Melanie; Maas, Richard L; Kucherlapati, Raju

    2006-01-01

    The tumor suppressor gene Apc (adenomatous polyposis coli) is a member of the Wnt signaling pathway that is involved in development and tumorigenesis. Heterozygous knockout mice for Apc have a tumor predisposition phenotype and homozygosity leads to embryonic lethality. To understand the role of Apc in development we generated a floxed allele. These mice were mated with a strain carrying Cre recombinase under the control of the human Keratin 14 (K14) promoter, which is active in basal cells of epidermis and other stratified epithelia. Mice homozygous for the floxed allele that also carry the K14-cre transgene were viable but had stunted growth and died before weaning. Histological and immunochemical examinations revealed that K14-cre–mediated Apc loss resulted in aberrant growth in many ectodermally derived squamous epithelia, including hair follicles, teeth, and oral and corneal epithelia. In addition, squamous metaplasia was observed in various epithelial-derived tissues, including the thymus. The aberrant growth of hair follicles and other appendages as well as the thymic abnormalities in K14-cre; ApcCKO/CKO mice suggest the Apc gene is crucial in embryonic cells to specify epithelial cell fates in organs that require epithelial–mesenchymal interactions for their development. PMID:17002498

  6. Mechanism of APC/CCDC20 activation by mitotic phosphorylation

    PubMed Central

    Qiao, Renping; Weissmann, Florian; Yamaguchi, Masaya; Brown, Nicholas G.; VanderLinden, Ryan; Imre, Richard; Jarvis, Marc A.; Brunner, Michael R.; Davidson, Iain F.; Litos, Gabriele; Haselbach, David; Mechtler, Karl; Stark, Holger; Schulman, Brenda A.; Peters, Jan-Michael

    2016-01-01

    Chromosome segregation and mitotic exit are initiated by the 1.2-MDa ubiquitin ligase APC/C (anaphase-promoting complex/cyclosome) and its coactivator CDC20 (cell division cycle 20). To avoid chromosome missegregation, APC/CCDC20 activation is tightly controlled. CDC20 only associates with APC/C in mitosis when APC/C has become phosphorylated and is further inhibited by a mitotic checkpoint complex until all chromosomes are bioriented on the spindle. APC/C contains 14 different types of subunits, most of which are phosphorylated in mitosis on multiple sites. However, it is unknown which of these phospho-sites enable APC/CCDC20 activation and by which mechanism. Here we have identified 68 evolutionarily conserved mitotic phospho-sites on human APC/C bound to CDC20 and have used the biGBac technique to generate 47 APC/C mutants in which either all 68 sites or subsets of them were replaced by nonphosphorylatable or phospho-mimicking residues. The characterization of these complexes in substrate ubiquitination and degradation assays indicates that phosphorylation of an N-terminal loop region in APC1 is sufficient for binding and activation of APC/C by CDC20. Deletion of the N-terminal APC1 loop enables APC/CCDC20 activation in the absence of mitotic phosphorylation or phospho-mimicking mutations. These results indicate that binding of CDC20 to APC/C is normally prevented by an autoinhibitory loop in APC1 and that its mitotic phosphorylation relieves this inhibition. The predicted location of the N-terminal APC1 loop implies that this loop controls interactions between the N-terminal domain of CDC20 and APC1 and APC8. These results reveal how APC/C phosphorylation enables CDC20 to bind and activate the APC/C in mitosis. PMID:27114510

  7. Mechanism of APC/CCDC20 activation by mitotic phosphorylation.

    PubMed

    Qiao, Renping; Weissmann, Florian; Yamaguchi, Masaya; Brown, Nicholas G; VanderLinden, Ryan; Imre, Richard; Jarvis, Marc A; Brunner, Michael R; Davidson, Iain F; Litos, Gabriele; Haselbach, David; Mechtler, Karl; Stark, Holger; Schulman, Brenda A; Peters, Jan-Michael

    2016-05-10

    Chromosome segregation and mitotic exit are initiated by the 1.2-MDa ubiquitin ligase APC/C (anaphase-promoting complex/cyclosome) and its coactivator CDC20 (cell division cycle 20). To avoid chromosome missegregation, APC/C(CDC20) activation is tightly controlled. CDC20 only associates with APC/C in mitosis when APC/C has become phosphorylated and is further inhibited by a mitotic checkpoint complex until all chromosomes are bioriented on the spindle. APC/C contains 14 different types of subunits, most of which are phosphorylated in mitosis on multiple sites. However, it is unknown which of these phospho-sites enable APC/C(CDC20) activation and by which mechanism. Here we have identified 68 evolutionarily conserved mitotic phospho-sites on human APC/C bound to CDC20 and have used the biGBac technique to generate 47 APC/C mutants in which either all 68 sites or subsets of them were replaced by nonphosphorylatable or phospho-mimicking residues. The characterization of these complexes in substrate ubiquitination and degradation assays indicates that phosphorylation of an N-terminal loop region in APC1 is sufficient for binding and activation of APC/C by CDC20. Deletion of the N-terminal APC1 loop enables APC/C(CDC20) activation in the absence of mitotic phosphorylation or phospho-mimicking mutations. These results indicate that binding of CDC20 to APC/C is normally prevented by an autoinhibitory loop in APC1 and that its mitotic phosphorylation relieves this inhibition. The predicted location of the N-terminal APC1 loop implies that this loop controls interactions between the N-terminal domain of CDC20 and APC1 and APC8. These results reveal how APC/C phosphorylation enables CDC20 to bind and activate the APC/C in mitosis. PMID:27114510

  8. Oncogenic mutations in adenomatous polyposis coli (Apc) activate mechanistic target of rapamycin complex 1 (mTORC1) in mice and zebrafish

    PubMed Central

    Valvezan, Alexander J.; Huang, Jian; Lengner, Christopher J.; Pack, Michael; Klein, Peter S.

    2014-01-01

    Truncating mutations in adenomatous polyposis coli (APC) are strongly linked to colorectal cancers. APC is a negative regulator of the Wnt pathway and constitutive Wnt activation mediated by enhanced Wnt–β-catenin target gene activation is believed to be the predominant mechanism responsible for APC mutant phenotypes. However, recent evidence suggests that additional downstream effectors contribute to APC mutant phenotypes. We previously identified a mechanism in cultured human cells by which APC, acting through glycogen synthase kinase-3 (GSK-3), suppresses mTORC1, a nutrient sensor that regulates cell growth and proliferation. We hypothesized that truncating Apc mutations should activate mTORC1 in vivo and that mTORC1 plays an important role in Apc mutant phenotypes. We find that mTORC1 is strongly activated in apc mutant zebrafish and in intestinal polyps in Apc mutant mice. Furthermore, mTORC1 activation is essential downstream of APC as mTORC1 inhibition partially rescues Apc mutant phenotypes including early lethality, reduced circulation and liver hyperplasia. Importantly, combining mTORC1 and Wnt inhibition rescues defects in morphogenesis of the anterior-posterior axis that are not rescued by inhibition of either pathway alone. These data establish mTORC1 as a crucial, β-catenin independent effector of oncogenic Apc mutations and highlight the importance of mTORC1 regulation by APC during embryonic development. Our findings also suggest a new model of colorectal cancer pathogenesis in which mTORC1 is activated in parallel with Wnt/β-catenin signaling. PMID:24092877

  9. The CDK-APC/C Oscillator Predominantly Entrains Periodic Cell-Cycle Transcription.

    PubMed

    Rahi, Sahand Jamal; Pecani, Kresti; Ondracka, Andrej; Oikonomou, Catherine; Cross, Frederick R

    2016-04-01

    Throughout cell-cycle progression, the expression of multiple transcripts oscillate, and whether these are under the centralized control of the CDK-APC/C proteins or can be driven by a de-centralized transcription factor (TF) cascade is a fundamental question for understanding cell-cycle regulation. In budding yeast, we find that the transcription of nearly all genes, as assessed by RNA-seq or fluorescence microscopy in single cells, is dictated by CDK-APC/C. Three exceptional genes are transcribed in a pulsatile pattern in a variety of CDK-APC/C arrests. Pursuing one of these transcripts, the SIC1 inhibitor of B-type cyclins, we use a combination of mathematical modeling and experimentation to provide evidence that, counter-intuitively, Sic1 provides a failsafe mechanism promoting nuclear division when levels of mitotic cyclins are low. PMID:27058667

  10. Structure of an APC3–APC16 Complex: Insights into Assembly of the Anaphase-Promoting Complex/Cyclosome

    SciTech Connect

    Yamaguchi, Masaya; Yu, Shanshan; Qiao, Renping; Weissmann, Florian; Miller, Darcie J.; VanderLinden, Ryan; Brown, Nicholas G.; Frye, Jeremiah J.; Peters, Jan-Michael; Schulman, Brenda A.

    2015-08-21

    The anaphase-promoting complex/cyclosome (APC/C) is a massive E3 ligase that controls mitosis by catalyzing ubiquitination of key cell cycle regulatory proteins. The APC/C assembly contains two subcomplexes: the “Platform” centers around a cullin-RING-like E3 ligase catalytic core; the “Arc Lamp” is a hub that mediates transient association with regulators and ubiquitination substrates. The Arc Lamp contains the small subunits APC16, CDC26, and APC13, and tetratricopeptide repeat (TPR) proteins (APC7, APC3, APC6, and APC8) that homodimerize and stack with quasi-2-fold symmetry. Within the APC/C complex, APC3 serves as center for regulation. APC3's TPR motifs recruit substrate-binding coactivators, CDC20 and CDH1, via their C-terminal conserved Ile-Arg (IR) tail sequences. Human APC3 also binds APC16 and APC7 and contains a > 200-residue loop that is heavily phosphorylated during mitosis, although the basis for APC3 interactions and whether loop phosphorylation is required for ubiquitination are unclear. Here, we map the basis for human APC3 assembly with APC16 and APC7, report crystal structures of APC3Δloop alone and in complex with the C-terminal domain of APC16, and test roles of APC3's loop and IR tail binding surfaces in APC/C-catalyzed ubiquitination. The structures show how one APC16 binds asymmetrically to the symmetric APC3 dimer and, together with biochemistry and prior data, explain how APC16 recruits APC7 to APC3, show how APC3's C-terminal domain is rearranged in the full APC/C assembly, and visualize residues in the IR tail binding cleft important for coactivator-dependent ubiquitination. Overall, the results provide insights into assembly, regulation, and interactions of TPR proteins and the APC/C.

  11. APC functions at the centrosome to stimulate microtubule growth.

    PubMed

    Lui, Christina; Ashton, Cahora; Sharma, Manisha; Brocardo, Mariana G; Henderson, Beric R

    2016-01-01

    The adenomatous polyposis coli (APC) tumor suppressor is multi-functional. APC is known to localize at the centrosome, and in mitotic cells contributes to formation of the mitotic spindle. To test whether APC contributes to nascent microtubule (MT) growth at interphase centrosomes, we employed MT regrowth assays in U2OS cells to measure MT assembly before and after nocodazole treatment and release. We showed that siRNA knockdown of full-length APC delayed both initial MT aster formation and MT elongation/regrowth. In contrast, APC-mutant SW480 cancer cells displayed a defect in MT regrowth that was unaffected by APC knockdown, but which was rescued by reconstitution of full-length APC. Our findings identify APC as a positive regulator of centrosome MT initial assembly and suggest that this process is disrupted by cancer mutations. We confirmed that full-length APC associates with the MT-nucleation factor γ-tubulin, and found that the APC cancer-truncated form (1-1309) also bound to γ-tubulin through APC amino acids 1-453. While binding to γ-tubulin may help target APC to the site of MT nucleation complexes, additional C-terminal sequences of APC are required to stimulate and stabilize MT growth. PMID:26556314

  12. Attenuated familial adenomatous polyposis with desmoids caused by an APC mutation.

    PubMed

    Ikenoue, Tsuneo; Yamaguchi, Kiyoshi; Komura, Mitsuhiro; Imoto, Seiya; Yamaguchi, Rui; Shimizu, Eigo; Kasuya, Shinichi; Shibuya, Tetsuo; Hatakeyama, Seira; Miyano, Satoru; Furukawa, Yoichi

    2015-01-01

    We present here a case of attenuated familial adenomatous polyposis (AFAP) with a family history of desmoids and thyroid tumors. This patient had no colonic polyps but did have multiple desmoids. Genetic analysis identified a 4-bp deletion in codon 2644 (c.7932_7935delTTAT: p.Tyr2645LysfsX14) of the adenomatous polyposis coli (APC) gene. In cases with limited numbers of colonic polyps and desmoids, AFAP may be caused by a mutation in the 3' region of APC. PMID:27081525

  13. Attenuated familial adenomatous polyposis with desmoids caused by an APC mutation

    PubMed Central

    Ikenoue, Tsuneo; Yamaguchi, Kiyoshi; Komura, Mitsuhiro; Imoto, Seiya; Yamaguchi, Rui; Shimizu, Eigo; Kasuya, Shinichi; Shibuya, Tetsuo; Hatakeyama, Seira; Miyano, Satoru; Furukawa, Yoichi

    2015-01-01

    We present here a case of attenuated familial adenomatous polyposis (AFAP) with a family history of desmoids and thyroid tumors. This patient had no colonic polyps but did have multiple desmoids. Genetic analysis identified a 4-bp deletion in codon 2644 (c.7932_7935delTTAT: p.Tyr2645LysfsX14) of the adenomatous polyposis coli (APC) gene. In cases with limited numbers of colonic polyps and desmoids, AFAP may be caused by a mutation in the 3′ region of APC. PMID:27081525

  14. Regulation of APC and AXIN2 expression by intestinal tumor suppressor CDX2 in colon cancer cells.

    PubMed

    Olsen, Anders Krüger; Coskun, Mehmet; Bzorek, Michael; Kristensen, Michael Holmsgaard; Danielsen, Erik Thomas; Jørgensen, Steffen; Olsen, Jørgen; Engel, Ulla; Holck, Susanne; Troelsen, Jesper Thorvald

    2013-06-01

    Wnt signaling is often constitutively active in colorectal cancer cells. The expression of the intestinal specific transcription factor CDX2 is found to be transiently decreased in invasive cells at the tumor/stroma interface. A recent ChIP-Seq study has indicated that several Wnt signaling-related genes are regulated by CDX2. The aim was to investigate the role of decreased CDX2 level on the expression of APC, AXIN2 and GSK3β in migrating colon cancer cells at the invasive front. CDX2-bound promoter and enhancer regions from APC, AXIN2 and GSK3β were analyzed for gene regulatory activity and the expression pattern of APC and GSK3β at the invasive front was evaluated by immunohistochemical procedures. Transfection of intestinal and non-intestinal cell lines demonstrated that CDX2 activated APC and AXIN2 promoter activities via intestinal cell-specific enhancer elements. Suppressed CDX2 expression was associated with endogenous downregulation of APC and AXIN2 expression in Caco-2 cells but did not affect GSK3β expression. Furthermore, elevated levels of nuclear β-catenin and reduced levels of cytoplasmic APC were correlated to a low CDX2 expression in migrating colon cancer cells in vivo. These results suggest that a low CDX2 level has influence on the Wnt signaling in invasive colon cancer cells possibly promoting cellular migration. PMID:23393221

  15. Integrated metrology: an enabler for advanced process control (APC)

    NASA Astrophysics Data System (ADS)

    Schneider, Claus; Pfitzner, Lothar; Ryssel, Heiner

    2001-04-01

    Advanced process control (APC) techniques become more and more important as short innovation cycles in microelectronics and a highly competitive market requires cost-effective solutions in semiconductor manufacturing. APC marks a paradigm shift from statistically based techniques (SPC) using monitor wafers for sampling measurement data towards product wafer control. The APC functionalities including run-to-run control, fault detection, and fault analysis allow to detect process drifts and excursions at an early stage and to minimize the number of misprocessed wafers. APC is being established as part of factory control systems through the definition of an APC framework. A precondition for APC is the availability of sensors and measurement methods providing the necessary wafer data. This paper discusses integrated metrology as an enabler for APC and demonstrates practical implementations in semiconductor manufacturing.

  16. Adenomatous polyposis coli (APC) membrane recruitment 3, a member of the APC membrane recruitment family of APC-binding proteins, is a positive regulator of Wnt-β-catenin signalling.

    PubMed

    Brauburger, Katharina; Akyildiz, Senem; Ruppert, Jan G; Graeb, Michael; Bernkopf, Dominic B; Hadjihannas, Michel V; Behrens, Jürgen

    2014-02-01

    The adenomatous polyposis coli (APC) membrane recruitment (Amer) family proteins Amer1/Wilms tumour gene on the X chromosome and Amer2 are binding partners of the APC tumour suppressor protein, and act as negative regulators in the Wnt signalling cascade. So far, nothing has been known about the third member of the family, Amer3. Here we show that Amer3 binds to the armadillo repeat domain of APC, similarly to Amer1 and Amer2. Amer3 also binds to the Wnt pathway regulator conductin/axin2. Furthermore, we identified Amer1 as binding partner of Amer3. Whereas Amer1 and Amer2 are linked to the plasma membrane by an N-terminal membrane localization domain, Amer3 lacks this domain. Amer3 localizes to the cytoplasm and nucleus of epithelial cells, and this is dependent on specific nuclear import and export sequences. Functionally, exogenous Amer3 enhances the expression of a β-catenin/T-cell factor-dependent reporter gene, and knockdown of endogenous Amer3 reduces Wnt target gene expression in colorectal cancer cells. Thus, Amer3 acts as an activator of Wnt signalling, in contrast to Amer1 and Amer2, which are inhibitors, suggesting a nonredundant role of Amer proteins in the regulation of this pathway. Our data, together with those of previous studies, provide a comprehensive picture of similarities and differences within the Amer protein family. PMID:24251807

  17. Molecular mechanism of APC/C activation by mitotic phosphorylation.

    PubMed

    Zhang, Suyang; Chang, Leifu; Alfieri, Claudio; Zhang, Ziguo; Yang, Jing; Maslen, Sarah; Skehel, Mark; Barford, David

    2016-05-12

    In eukaryotes, the anaphase-promoting complex (APC/C, also known as the cyclosome) regulates the ubiquitin-dependent proteolysis of specific cell-cycle proteins to coordinate chromosome segregation in mitosis and entry into the G1 phase. The catalytic activity of the APC/C and its ability to specify the destruction of particular proteins at different phases of the cell cycle are controlled by its interaction with two structurally related coactivator subunits, Cdc20 and Cdh1. Coactivators recognize substrate degrons, and enhance the affinity of the APC/C for its cognate E2 (refs 4-6). During mitosis, cyclin-dependent kinase (Cdk) and polo-like kinase (Plk) control Cdc20- and Cdh1-mediated activation of the APC/C. Hyperphosphorylation of APC/C subunits, notably Apc1 and Apc3, is required for Cdc20 to activate the APC/C, whereas phosphorylation of Cdh1 prevents its association with the APC/C. Since both coactivators associate with the APC/C through their common C-box and Ile-Arg tail motifs, the mechanism underlying this differential regulation is unclear, as is the role of specific APC/C phosphorylation sites. Here, using cryo-electron microscopy and biochemical analysis, we define the molecular basis of how phosphorylation of human APC/C allows for its control by Cdc20. An auto-inhibitory segment of Apc1 acts as a molecular switch that in apo unphosphorylated APC/C interacts with the C-box binding site and obstructs engagement of Cdc20. Phosphorylation of the auto-inhibitory segment displaces it from the C-box-binding site. Efficient phosphorylation of the auto-inhibitory segment, and thus relief of auto-inhibition, requires the recruitment of Cdk-cyclin in complex with a Cdk regulatory subunit (Cks) to a hyperphosphorylated loop of Apc3. We also find that the small-molecule inhibitor, tosyl-l-arginine methyl ester, preferentially suppresses APC/C(Cdc20) rather than APC/C(Cdh1), and interacts with the binding sites of both the C-box and Ile-Arg tail motifs. Our

  18. Detection of APC mosaicism by next-generation sequencing in an FAP patient.

    PubMed

    Yamaguchi, Kiyoshi; Komura, Mitsuhiro; Yamaguchi, Rui; Imoto, Seiya; Shimizu, Eigo; Kasuya, Shinichi; Shibuya, Tetsuo; Hatakeyama, Seira; Takahashi, Norihiko; Ikenoue, Tsuneo; Hata, Keisuke; Tsurita, Giichiro; Shinozaki, Masaru; Suzuki, Yutaka; Sugano, Sumio; Miyano, Satoru; Furukawa, Yoichi

    2015-05-01

    Familial adenomatous polyposis (FAP) of the colon is characterized by multiple polyps in the intestine and extra-colonic manifestations. Most FAP cases are caused by a germline mutation in the tumor-suppressor gene APC, but some cases of adenomatous polyposis result from germline mutations in MUTYH, POLD1 or POLE. Although sequence analysis of APC by the Sanger method is routinely performed for genetic testing, there remain cases whose mutations are not detected by the analysis. Next-generation sequencing has enabled us to analyze the comprehensive human genome, improving the chance of identifying disease causative variants. In this study, we conducted whole-genome sequencing of a sporadic FAP patient in which we did not find any pathogenic APC mutations by the conventional Sanger sequencing. Whole-genome sequencing and subsequent deep sequencing identified a mosaic mutation of c.3175G>T, p.E1059X in ~12% of his peripheral leukocytes. Additional deep sequencing of his buccal mucosa, hair follicles, non-cancerous mucosa of the stomach and colon disclosed that these tissues harbored the APC mutation at different frequencies. Our data implied that genetic analysis by next-generation sequencing is an effective strategy to identify genetic mosaicism in hereditary diseases. PMID:25716913

  19. PIK3CA and APC Mutations are Synergistic in the Development of Intestinal Cancers

    PubMed Central

    Deming, Dustin A.; Leystra, Alyssa A.; Nettekoven, Laura; Sievers, Chelsea; Miller, Devon; Middlebrooks, Malisa; Clipson, Linda; Albrecht, Dawn; Bacher, Jeff; Washington, Mary Kay; Weichert, Jamey; Halberg, Richard B.

    2013-01-01

    Human colorectal cancers are known to possess multiple mutations, though how these mutations interact in tumor development and progression has not been fully investigated. We have previously described the FCPIK3ca* murine colon cancer model which expresses a constitutively activated phosphoinositide-3 kinase (PI3K) in the intestinal epithelium. The expression of this dominantly active form of PI3K results in hyperplasia and invasive mucinous adenocarcinomas. These cancers form via a non-canonical mechanism of tumor initiation that is mediated through activation of PI3K and not through aberrations in WNT signaling. Since the Adenomatous Polyposis Coli (APC) gene is mutated in the vast majority of human colon cancers and often occurs simultaneously with PIK3CA mutations, we sought to better understand the interaction between APC and PIK3CA mutations in the mammalian intestine. In this study, we have generated mice in which the expression of a constitutively active PI3K and the loss of APC occur simultaneously in the distal small intestine and colon. Here we demonstrate that expression of a dominant active PI3K synergizes with loss of APC activity resulting in a dramatic changes in tumor multiplicity, size, morphology, and invasiveness. Activation of the PI3K pathway is not able to directly activate WNT signaling through the nuclear localization of CTNNB1 (β-catenin) in the absence of aberrant WNT signaling. Alterations at the transcriptional level, including increased CCND1, may be the etiology of synergy between these activated pathways. PMID:23708654

  20. Alterations in K-ras, APC and p53-multiple genetic pathway in colorectal cancer among Indians.

    PubMed

    Malhotra, Pooja; Anwar, Mumtaz; Nanda, Neha; Kochhar, Rakesh; Wig, Jai Dev; Vaiphei, Kim; Mahmood, Safrun

    2013-06-01

    The incidence of colorectal cancer (CRC) is increasing rapidly in Asian countries during the past few decades, but no comprehensive analysis has been done to find out the exact cause of this disease. In this study, we investigated the frequencies of mutations and expression pattern of K-ras, APC (adenomatosis polyposis coli) and p53 in tumor, adjoining and distant normal mucosa and to correlate these alterations with patients clinicopathological parameters as well as with the survival. Polymerase chain reaction (PCR)-restriction digestion was used to detect mutations in K-ras and PCR-SSCP (Single Strand Conformation Polymorphism) followed by DNA sequencing was used to detect mutations in APC and p53 genes. Immunohistochemistry was used to detect the expression pattern of K-ras, APC and p53 proteins. The frequencies of mutations of K-ras, APC and p53 in 30 tumor tissues samples were 26.7 %, 46.7 % and 20 %, respectively. Only 3.3 % of tumors contained mutations in all the three genes. The most common combination of mutation was APC and p53 whereas mutation in both p53 and K-ras were extremely rare. There was no association between the mutations and expression pattern of K-ras, APC and p53 (p>0.05). In Indians, the frequency of alterations of K-ras and APC is similar as in Westerns, whereas the frequency of p53 mutation is slightly lower. The lack of multiple mutations in tumor specimens suggests that these genetic alterations might have independent influences on CRC development and there could be multiple alternative genetic pathways to CRC in our present study cohort. PMID:23526092

  1. Identification of five novel modifier loci of ApcMin harbored in the BXH14 recombinant inbred strain

    PubMed Central

    Siracusa, Linda D.

    2012-01-01

    Every year thousands of people in the USA are diagnosed with small intestine and colorectal cancers (CRC). Although environmental factors affect disease etiology, uncovering underlying genetic factors is imperative for risk assessment and developing preventative therapies. Familial adenomatous polyposis is a heritable genetic disorder in which individuals carry germ-line mutations in the adenomatous polyposis coli (APC) gene that predisposes them to CRC. The Apc Min mouse model carries a point mutation in the Apc gene and develops polyps along the intestinal tract. Inbred strain background influences polyp phenotypes in Apc Min mice. Several Modifier of Min (Mom) loci that alter tumor phenotypes associated with the Apc Min mutation have been identified to date. We screened BXH recombinant inbred (RI) strains by crossing BXH RI females with C57BL/6J (B6) Apc Min males and quantitating tumor phenotypes in backcross progeny. We found that the BXH14 RI strain harbors five modifier loci that decrease polyp multiplicity. Furthermore, we show that resistance is determined by varying combinations of these modifier loci. Gene interaction network analysis shows that there are multiple networks with proven gene–gene interactions, which contain genes from all five modifier loci. We discuss the implications of this result for studies that define susceptibility loci, namely that multiple networks may be acting concurrently to alter tumor phenotypes. Thus, the significance of this work resides not only with the modifier loci we identified but also with the combinations of loci needed to get maximal protection against polyposis and the impact of this finding on human disease studies. Abbreviations:APCadenomatous polyposis coliGWASgenome-wide association studiesQTLquantitative trait lociSNPsingle-nucleotide polymorphism. PMID:22637734

  2. The postsynaptic adenomatous polyposis coli (APC) multiprotein complex is required for localizing neuroligin and neurexin to neuronal nicotinic synapses in vivo.

    PubMed

    Rosenberg, Madelaine M; Yang, Fang; Mohn, Jesse L; Storer, Elizabeth K; Jacob, Michele H

    2010-08-18

    Synaptic efficacy requires that presynaptic and postsynaptic specializations align precisely and mature coordinately. The underlying mechanisms are poorly understood, however. We propose that adenomatous polyposis coli protein (APC) is a key coordinator of presynaptic and postsynaptic maturation. APC organizes a multiprotein complex that directs nicotinic acetylcholine receptor (nAChR) localization at postsynaptic sites in avian ciliary ganglion neurons in vivo. We hypothesize that the APC complex also provides retrograde signals that direct presynaptic active zones to develop in register with postsynaptic nAChR clusters. In our model, the APC complex provides retrograde signals via postsynaptic neuroligin that interacts extracellularly with presynaptic neurexin. S-SCAM (synaptic cell adhesion molecule) and PSD-93 (postsynaptic density-93) are scaffold proteins that bind to neuroligin. We identify S-SCAM as a novel component of neuronal nicotinic synapses. We show that S-SCAM, PSD-93, neuroligin and neurexin are enriched at alpha3*-nAChR synapses. PSD-93 and S-SCAM bind to APC and its binding partner beta-catenin, respectively. Blockade of selected APC and beta-catenin interactions, in vivo, leads to decreased postsynaptic accumulation of S-SCAM, but not PSD-93. Importantly, neuroligin synaptic clusters are also decreased. On the presynaptic side, there are decreases in neurexin and active zone proteins. Further, presynaptic terminals are less mature structurally and functionally. We define a novel neural role for APC by showing that the postsynaptic APC multiprotein complex is required for anchoring neuroligin and neurexin at neuronal synapses in vivo. APC human gene mutations correlate with autism spectrum disorders, providing strong support for the importance of the association, demonstrated here, between APC, neuroligin and neurexin. PMID:20720115

  3. Methylation status of the APC and RASSF1A promoter in cell-free circulating DNA and its prognostic role in patients with colorectal cancer

    PubMed Central

    MATTHAIOS, DIMITRIOS; BALGKOURANIDOU, IOANNA; KARAYIANNAKIS, ANASTASIOS; BOLANAKI, HELEN; XENIDIS, NIKOLAOS; AMARANTIDIS, KYRIAKOS; CHELIS, LEONIDAS; ROMANIDIS, KONSTANTINOS; CHATZAKI, AIKATERINI; LIANIDOU, EVI; TRYPSIANIS, GRIGORIOS; KAKOLYRIS, STYLIANOS

    2016-01-01

    DNA methylation is the most frequent epigenetic alteration. Using methylation-specific polymerase chain reaction (MSP), the methylation status of the adenomatous polyposis coli (APC) and Ras association domain family 1 isoform A (RASSF1A) genes was examined in cell-free circulating DNA from 155 plasma samples obtained from patients with early and advanced colorectal cancer (CRC). APC and RASSF1A hypermethylation was frequently observed in both early and advanced disease, and was significantly associated with a poorer disease outcome. The methylation status of the APC and RASSF1A promoters was investigated in cell-free DNA of patients with CRC. Using MSP, the promoter methylation status of APC and RASSF1A was examined in 155 blood samples obtained from patients with CRC, 88 of whom had operable CRC (oCRC) and 67 had metastatic CRC (mCRC). The frequency of APC methylation in patients with oCRC was 33%. Methylated APC promoter was significantly associated with older age (P=0.012), higher stage (P=0.014) and methylated RASSF1A status (P=0.050). The frequency of APC methylation in patients with mCRC was 53.7%. In these patients, APC methylation was significantly associated with methylated RASSF1A status (P=0.016). The frequency of RASSF1A methylation in patients with oCRC was 25%. Methylated RASSF1A in oCRC was significantly associated with higher stage (P=0.021). The frequency of RASSF1A methylation in mCRC was 44.8%. Methylated RASSF1A in mCRC was associated with moderate differentiation (P=0.012), high levels of carcinoembryonic antigen (P=0.023) and methylated APC status (P=0.016). Patients with an unmethylated APC gene had better survival in both early (81±5 vs. 27±4 months, P<0.001) and advanced disease (37±7 vs. 15±3 months, P<0.001), compared with patients with methylated APC. Patients with an unmethylated RASSF1A gene had better survival in both early (71±6 vs. 46±8 months, P<0.001) and advanced disease (28±4 vs. 16±3 months, P<0.001) than patients with

  4. Tumor promotion and inhibition by phenobarbital in livers of conditional Apc-deficient mice.

    PubMed

    Braeuning, Albert; Gavrilov, Alina; Geissler, Miriam; Wenz, Christine; Colnot, Sabine; Templin, Markus F; Metzger, Ute; Römer, Michael; Zell, Andreas; Schwarz, Michael

    2016-06-01

    Activation of Wnt/β-catenin signaling is important for human and rodent hepatocarcinogenesis. In mice, the tumor promoter phenobarbital (PB) selects for hepatocellular tumors with activating β-catenin mutations via constitutive androstane receptor activation. PB-dependent tumor promotion was studied in mice with genetic inactivation of Apc, a negative regulator of β-catenin, to circumvent the problem of randomly induced mutations by chemical initiators and to allow monitoring of PB- and Wnt/β-catenin-dependent tumorigenesis in the absence of unknown genomic alterations. Moreover, the study was designed to investigate PB-induced proliferation of liver cells with activated β-catenin. PB treatment provided Apc-deficient hepatocytes with only a minor proliferative advantage, and additional connexin 32 deficiency did not affect the proliferative response. PB significantly promoted the outgrowth of Apc-deficient hepatocellular adenoma (HCA), but simultaneously inhibited the formation of Apc-deficient hepatocellular carcinoma (HCC). The probability of tumor promotion by PB was calculated to be much lower for hepatocytes with loss of Apc, as compared to mutational β-catenin activation. Comprehensive transcriptomic and phosphoproteomic characterization of HCA and HCC revealed molecular details of the two tumor types. HCC were characterized by a loss of differentiated hepatocellular gene expression, enhanced proliferative signaling, and massive over-activation of Wnt/β-catenin signaling. In conclusion, PB exerts a dual role in liver tumor formation by promoting the growth of HCA but inhibiting the growth of HCC. Data demonstrate that one and the same compound can produce opposite effects on hepatocarcinogenesis, depending on context, highlighting the necessity to develop a more differentiated view on the tumorigenicity of this model compound. PMID:26838046

  5. HectD1 E3 ligase modifies adenomatous polyposis coli (APC) with polyubiquitin to promote the APC-axin interaction.

    PubMed

    Tran, Hoanh; Bustos, Daisy; Yeh, Ronald; Rubinfeld, Bonnee; Lam, Cynthia; Shriver, Stephanie; Zilberleyb, Inna; Lee, Michelle W; Phu, Lilian; Sarkar, Anjali A; Zohn, Irene E; Wertz, Ingrid E; Kirkpatrick, Donald S; Polakis, Paul

    2013-02-01

    The adenomatous polyposis coli (APC) protein functions as a negative regulator of the Wnt signaling pathway. In this capacity, APC forms a "destruction complex" with Axin, CK1α, and GSK3β to foster phosphorylation of the Wnt effector β-catenin earmarking it for Lys-48-linked polyubiquitylation and proteasomal degradation. APC is conjugated with Lys-63-linked ubiquitin chains when it is bound to Axin, but it is unclear whether this modification promotes the APC-Axin interaction or confers upon APC an alternative function in the destruction complex. Here we identify HectD1 as a candidate E3 ubiquitin ligase that modifies APC with Lys-63 polyubiquitin. Knockdown of HectD1 diminished APC ubiquitylation, disrupted the APC-Axin interaction, and augmented Wnt3a-induced β-catenin stabilization and signaling. These results indicate that HectD1 promotes the APC-Axin interaction to negatively regulate Wnt signaling. PMID:23277359

  6. Specific immunotherapy of experimental myasthenia by genetically engineered APCs: the "guided missile" strategy.

    PubMed

    Drachman, D B; Wu, J-M; Miagkov, A; Williams, M A; Adams, R N; Wu, B

    2003-09-01

    Although treatment of MG with general immunosuppressive agents is often effective, it has important drawbacks, including suppression of the immune system as a whole, with the risks of infection and neoplasia, and numerous other adverse side effects. Ideally, treatment of MG should eliminate the specific pathogenic autoimmune response to AChR, without otherwise suppressing the immune system or producing other adverse side effects. Although antibodies to AChR are directly responsible for the loss of AChRs at neuromuscular junctions in MG, the AChR antibody response is T cell-dependent, and immunotherapy directed at T cells can abrogate the autoantibody response, with resulting benefit. As in other autoimmune diseases, the T cell response in MG is highly heterogeneous. The design of specific immunotherapy must take this heterogeneity into account and target the entire repertoire of AChR-specific T cells. We describe our investigation of a novel strategy for specific immunotherapy of MG, involving gene transfer to convert antigen-presenting cells (APCs) to "guided missiles" that target AChR-specific T cells, and that induce apoptosis and elimination of those T cells. This strategy uses the ability of APCs from a given individual to present the entire spectrum of AChR epitopes unique for that individual, and thereby to target the entire repertoire of antigen-specific T cells of the same individual. Using viral vectors, we have genetically engineered the APCs to process and present the most important domain of the AChR molecule, and to express a "warhead" of Fas ligand (FasL) to eliminate the activated AChR-specific T cells with which they interact. Our results show that the APCs express the appropriate gene products, and effectively and specifically eliminate AChR-specific T cells by the Fas/FasL pathway, while sparing T cells of other specificities. PMID:14592923

  7. Proteomic profiling of a mouse model of acute intestinal Apc deletion leads to identification of potential novel biomarkers of human colorectal cancer (CRC).

    PubMed

    Hammoudi, Abeer; Song, Fei; Reed, Karen R; Jenkins, Rosalind E; Meniel, Valerie S; Watson, Alastair J M; Pritchard, D Mark; Clarke, Alan R; Jenkins, John R

    2013-10-25

    Colorectal cancer (CRC) is the fourth most common cause of cancer-related death worldwide. Accurate non-invasive screening for CRC would greatly enhance a population's health. Adenomatous polyposis coli (Apc) gene mutations commonly occur in human colorectal adenomas and carcinomas, leading to Wnt signalling pathway activation. Acute conditional transgenic deletion of Apc in murine intestinal epithelium (AhCre(+)Apc(fl)(/)(fl)) causes phenotypic changes similar to those found during colorectal tumourigenesis. This study comprised a proteomic analysis of murine small intestinal epithelial cells following acute Apc deletion to identify proteins that show altered expression during human colorectal carcinogenesis, thus identifying proteins that may prove clinically useful as blood/serum biomarkers of colorectal neoplasia. Eighty-one proteins showed significantly increased expression following iTRAQ analysis, and validation of nine of these by Ingenuity Pathaway Analysis showed they could be detected in blood or serum. Expression was assessed in AhCre(+)Apc(fl)(/)(fl) small intestinal epithelium by immunohistochemistry, western blot and quantitative real-time PCR; increased nucelolin concentrations were also detected in the serum of AhCre(+)Apc(fl)(/)(fl) and Apc(Min)(/)(+) mice by ELISA. Six proteins; heat shock 60kDa protein 1, Nucleolin, Prohibitin, Cytokeratin 18, Ribosomal protein L6 and DEAD (Asp-Glu-Ala-Asp) box polypeptide 5,were selected for further investigation. Increased expression of 4 of these was confirmed in human CRC by qPCR. In conclusion, several novel candidate biomarkers have been identified from analysis of transgenic mice in which the Apc gene was deleted in the intestinal epithelium that also showed increased expression in human CRC. Some of these warrant further investigation as potential serum-based biomarkers of human CRC. PMID:23998936

  8. Antigen targeting to APC: from mice to veterinary species.

    PubMed

    Alvarez, B; Poderoso, T; Alonso, F; Ezquerra, A; Domínguez, J; Revilla, C

    2013-10-01

    Antigen delivery to receptors expressed on antigen presenting cells (APC) has shown to improve immunogenicity of vaccines in mice. An enhancement of cytotoxic T lymphocyte (CTL), helper T cell or humoral responses was obtained depending on the type of APC and the surface molecule targeted. Although this strategy is being also evaluated in livestock animals with promising results, some discrepancies have been found between species and pathogens. The genetic diversity of livestock animals, the different pattern of expression of some receptors among species, the use of different markers to characterize APC in large animals and sometimes the lack of reagents make difficult to compare results obtained in different species. In this review, we summarize the data available regarding antigen targeting to APC receptors in cattle, sheep and pig and discuss the results found in these animals in the context of what has been obtained in mice. PMID:23648645

  9. Los1p, Involved in Yeast Pre-Trna Splicing, Positively Regulates Members of the Sol Gene Family

    PubMed Central

    Shen, W. C.; Stanford, D. R.; Hopper, A. K.

    1996-01-01

    To understand the role of Los1p in pre-tRNA splicing, we sought los1 multicopy suppressors. We found SOL1 that suppresses both point and null LOS1 mutations. Since, when fused to the Gal4p DNA-binding domain, Los1p activates transcription, we tested whether Los1p regulates SOL1. We found that los1 mutants have depleted levels of SOL1 mRNA and Sollp. Thus, LOS1 appears to positively regulate SOL1. SOL1 belongs to a multigene family with at least two additional members, SOL2 and SOL3. Sol proteins have extensive similarity to an unusual group of glucose-6-phosphate dehydrogenases. As the similarities are restricted to areas separate from the catalytic domain, these G6PDs may have more than one function. The SOL family appears to be unessential since cells with a triple disruption of all three SOL genes are viable. SOL gene disruptions negatively affect tRNA-mediated nonsense suppression and the severity increases with the number of mutant SOL genes. However, tRNA levels do not vary with either multicopy SOL genes or with SOL disruptions. Therefore, the Sol proteins affect tRNA expression/function at steps other than transcription or splicing. We propose that LOS1 regulates gene products involved in tRNA expression/function as well as pre-tRNA splicing. PMID:8725220

  10. Tumour Suppressor Adenomatous Polyposis Coli (APC) localisation is regulated by both Kinesin-1 and Kinesin-2

    PubMed Central

    Ruane, Peter T.; Gumy, Laura F.; Bola, Becky; Anderson, Beverley; Wozniak, Marcin J.; Hoogenraad, Casper C.; Allan, Victoria J.

    2016-01-01

    Microtubules and their associated proteins (MAPs) underpin the polarity of specialised cells. Adenomatous polyposis coli (APC) is one such MAP with a multifunctional agenda that requires precise intracellular localisations. Although APC has been found to associate with kinesin-2 subfamily members, the exact mechanism for the peripheral localization of APC remains unclear. Here we show that the heavy chain of kinesin-1 directly interacts with the APC C-terminus, contributing to the peripheral localisation of APC in fibroblasts. In rat hippocampal neurons the kinesin-1 binding domain of APC is required for its axon tip enrichment. Moreover, we demonstrate that APC requires interactions with both kinesin-2 and kinesin-1 for this localisation. Underlining the importance of the kinesin-1 association, neurons expressing APC lacking kinesin-1-binding domain have shorter axons. The identification of this novel kinesin-1-APC interaction highlights the complexity and significance of APC localisation in neurons. PMID:27272132

  11. Tumour Suppressor Adenomatous Polyposis Coli (APC) localisation is regulated by both Kinesin-1 and Kinesin-2.

    PubMed

    Ruane, Peter T; Gumy, Laura F; Bola, Becky; Anderson, Beverley; Wozniak, Marcin J; Hoogenraad, Casper C; Allan, Victoria J

    2016-01-01

    Microtubules and their associated proteins (MAPs) underpin the polarity of specialised cells. Adenomatous polyposis coli (APC) is one such MAP with a multifunctional agenda that requires precise intracellular localisations. Although APC has been found to associate with kinesin-2 subfamily members, the exact mechanism for the peripheral localization of APC remains unclear. Here we show that the heavy chain of kinesin-1 directly interacts with the APC C-terminus, contributing to the peripheral localisation of APC in fibroblasts. In rat hippocampal neurons the kinesin-1 binding domain of APC is required for its axon tip enrichment. Moreover, we demonstrate that APC requires interactions with both kinesin-2 and kinesin-1 for this localisation. Underlining the importance of the kinesin-1 association, neurons expressing APC lacking kinesin-1-binding domain have shorter axons. The identification of this novel kinesin-1-APC interaction highlights the complexity and significance of APC localisation in neurons. PMID:27272132

  12. Identification of Mom12 and Mom13, two novel modifier loci of Apc (Min) -mediated intestinal tumorigenesis.

    PubMed

    Crist, Richard C; Roth, Jacquelyn J; Lisanti, Michael P; Siracusa, Linda D; Buchberg, Arthur M

    2011-04-01

    Colorectal cancer is a heterogeneous disease resulting from a combination of genetic and environmental factors. The C57BL/6J (B6) Apc (Min/+) mouse develops polyps throughout the gastrointestinal tract and has been a valuable model for understanding the genetic basis of intestinal tumorigenesis. Apc (Min/+) mice have been used to study known oncogenes and tumor suppressor genes on a controlled genetic background. These studies often utilize congenic knockout alleles, which can carry an unknown amount of residual donor DNA. The Apc (Min) model has also been used to identify modifer loci, known as Modifier of Min (Mom) loci, which alter Apc (Min) -mediated intestinal tumorigenesis. B6 mice carrying a knockout allele generated in WW6 embryonic stem cells were crossed to B6 Apc (Min/+) mice to determine the effect on polyp multiplicity. The newly generated colony developed significantly more intestinal polyps than Apc (Min/+) controls. Polyp multiplicity did not correlate with inheritance of the knockout allele, suggesting the presence of one or more modifier loci segregating in the colony. Genotyping of simple sequence length polymorphism (SSLP) markers revealed residual 129X1/SvJ genomic DNA within the congenic region of the parental knockout line. An analysis of polyp multiplicity data and SSLP genotyping indicated the presence of two Mom loci in the colony: 1) Mom12, a dominant modifier linked to the congenic region on chromosome 6, and 2) Mom13, which is unlinked to the congenic region and whose effect is masked by Mom12. The identification of Mom12 and Mom13 demonstrates the potential problems resulting from residual heterozygosity present in congenic lines. PMID:21386660

  13. APC germline mutations in families with familial adenomatous polyposis.

    PubMed

    De Queiroz Rossanese, Lillian Barbosa; De Lima Marson, Fernando Augusto; Ribeiro, José Dirceu; Coy, Claudio Saddy Rodrigues; Bertuzzo, Carmen Silvia

    2013-11-01

    Adenomatous polyposis coli (APC) germline mutations are responsible for the occurrence of familial adenomatous polyposis (FAP). Somatic mutations lead to malignant transformation of adenomas. In this context, considering the significance of APC germline mutations in FAP, we aimed to identify APC germline mutations. In the present study, 20 FAP patients were enrolled. The determination of APC germline mutations was performed using sequencing, and the mutations were compared with clinical markers (gender, age at diagnosis, smoking habits, TNM stage, Astler‑Coller stage, degree of differentiation of adenocarcinoma). The data were compared using the SPSS program, with the Fisher's exact test and χ2 test, considering α=0.05. According to the main results in our sample, 16 alleles with deleterious mutations (80% of the patients) were identified while 7 (35%) patients had no deleterious mutations. There was a predominance of nonsense (45% of the patients) and frameshift (20% of the patients) mutations. There was no statistical significance between the APC germline mutations identified and the clinical variables considered in our study. Only TNM stage was associated with the presence of deleterious mutations. Patients with deleterious mutations had an OR, 0.086 (IC=0.001-0.984); TNM stage I+II in comparison with III+IV, when compared with the patients with no deleterious mutations identified. In this context, as a conclusion, we demonstrated the molecular heterogeneity of APC germline mutations in FAP and the difficulty to perform molecular diagnostics in a Brazilian population, considering the admixed population analyzed. PMID:23970361

  14. The utility of Apc-mutant rats in modeling human colon cancer

    PubMed Central

    Irving, Amy A.; Yoshimi, Kazuto; Hart, Marcia L.; Parker, Taybor; Clipson, Linda; Ford, Madeline R.; Kuramoto, Takashi; Dove, William F.; Amos-Landgraf, James M.

    2014-01-01

    Prior to the advent of genetic engineering in the mouse, the rat was the model of choice for investigating the etiology of cancer. Now, recent advances in the manipulation of the rat genome, combined with a growing recognition of the physiological differences between mice and rats, have reignited interest in the rat as a model of human cancer. Two recently developed rat models, the polyposis in the rat colon (Pirc) and Kyoto Apc Delta (KAD) strains, each carry mutations in the intestinal-cancer-associated adenomatous polyposis coli (Apc) gene. In contrast to mouse models carrying Apc mutations, in which cancers develop mainly in the small intestine rather than in the colon and there is no gender bias, these rat models exhibit colonic predisposition and gender-specific susceptibility, as seen in human colon cancer. The rat also provides other experimental resources as a model organism that are not provided by the mouse: the structure of its chromosomes facilitates the analysis of genomic events, the size of its colon permits longitudinal analysis of tumor growth, and the size of biological samples from the animal facilitates multiplexed molecular analyses of the tumor and its host. Thus, the underlying biology and experimental resources of these rat models provide important avenues for investigation. We anticipate that advances in disease modeling in the rat will synergize with resources that are being developed in the mouse to provide a deeper understanding of human colon cancer. PMID:25288683

  15. The ABBA motif binds APC/C activators and is shared by APC/C substrates and regulators

    PubMed Central

    Hagting, Anja; Izawa, Daisuke; Mansfeld, Jörg; Gibson, Toby J.; Pines, Jonathon

    2016-01-01

    The APC/C is the ubiquitin ligase that regulates mitosis by targeting specific proteins for degradation at specific times under the control of the Spindle Assembly Checkpoint (SAC). How the APC/C recognises its different substrates is a key problem in the control of cell division. Here, we have identified the ABBA motif in Cyclin A, BUBR1, BUB1 and Acm1, and show that it binds to the APC/C co-activator CDC20. The ABBA motif in Cyclin A is required for its proper degradation in prometaphase through competing with BUBR1 for the same site on CDC20. Moreover, the ABBA motifs in BUBR1 and BUB1 are necessary for the SAC to work at full strength and to recruit CDC20 to kinetochores. Thus, we have identified a conserved motif integral to the proper control of mitosis that connects APC/C substrate recognition with the SAC. PMID:25669885

  16. SNW1 enables sister chromatid cohesion by mediating the splicing of sororin and APC2 pre-mRNAs

    PubMed Central

    van der Lelij, Petra; Stocsits, Roman R; Ladurner, Rene; Petzold, Georg; Kreidl, Emanuel; Koch, Birgit; Schmitz, Julia; Neumann, Beate; Ellenberg, Jan; Peters, Jan-Michael

    2014-01-01

    Although splicing is essential for the expression of most eukaryotic genes, inactivation of splicing factors causes specific defects in mitosis. The molecular cause of this defect is unknown. Here, we show that the spliceosome subunits SNW1 and PRPF8 are essential for sister chromatid cohesion in human cells. A transcriptome-wide analysis revealed that SNW1 or PRPF8 depletion affects the splicing of specific introns in a subset of pre-mRNAs, including pre-mRNAs encoding the cohesion protein sororin and the APC/C subunit APC2. SNW1 depletion causes cohesion defects predominantly by reducing sororin levels, which causes destabilisation of cohesin on DNA. SNW1 depletion also reduces APC/C activity and contributes to cohesion defects indirectly by delaying mitosis and causing “cohesion fatigue”. Simultaneous expression of sororin and APC2 from intron-less cDNAs restores cohesion in SNW1-depleted cells. These results indicate that the spliceosome is required for mitosis because it enables expression of genes essential for cohesion. Our transcriptome-wide identification of retained introns in SNW1- and PRPF8-depleted cells may help to understand the aetiology of diseases associated with splicing defects, such as retinosa pigmentosum and cancer. PMID:25257309

  17. Liver-targeted disruption of Apc in mice activates β-catenin signaling and leads to hepatocellular carcinomas

    PubMed Central

    Colnot, S.; Decaens, T.; Niwa-Kawakita, M.; Godard, C.; Hamard, G.; Kahn, A.; Giovannini, M.; Perret, C.

    2004-01-01

    Although inappropriate activation of the Wnt/β-catenin pathway has been implicated in the development of hepatocellular carcinoma (HCC), the role of this signaling in liver carcinogenesis remains unclear. To investigate this issue, we constructed a mutant mouse strain, Apclox/lox, in which exon 14 of the tumor-suppressor gene adenomatous polyposis coli (Apc) is flanked by loxP sequences. i.v. injection of adenovirus encoding Cre recombinase (AdCre) at high multiplicity [109 plaque-forming units (pfu) per mouse] inactivated the Apc gene in the liver and resulted in marked hepatomegaly, hepatocyte hyperplasia, and rapid mortality. β-Catenin signaling activation was demonstrated by nuclear and cytoplasmic accumulation of β-catenin in the hepatocytes and by the induction of β-catenin target genes (glutamine synthetase, glutamate transporter 1, ornithine aminotransferase, and leukocyte cell-derived chemotaxin 2) in the liver. To test a long-term oncogenic effect, we inoculated mice with lower doses of AdCre (0.5 × 109 pfu per mouse), compatible with both survival and persistence of β-catenin-activated cells. In these conditions, 67% of mice developed HCC. β-Catenin signaling was strongly activated in these Apc-inactivated HCCs. The HCCs were well, moderately, or poorly differentiated. Indeed, their histological and molecular features mimicked human HCC. Thus, deletion of Apc in the liver provides a valuable model of human HCC, and, in this model, activation of the Wnt/β-catenin pathway by invalidation of Apc is required for liver tumorigenesis. PMID:15563600

  18. NUP98 fusion oncoproteins interact with the APC/C(Cdc20) as a pseudosubstrate and prevent mitotic checkpoint complex binding.

    PubMed

    Salsi, Valentina; Fantini, Sebastian; Zappavigna, Vincenzo

    2016-09-01

    NUP98 is a recurrent partner gene in translocations causing acute myeloid leukemias and myelodisplastic syndrome. The expression of NUP98 fusion oncoproteins has been shown to induce mitotic spindle defects and chromosome missegregation, which correlate with the capability of NUP98 fusions to cause mitotic checkpoint attenuation. We show that NUP98 oncoproteins physically interact with the APC/C(Cdc20) in the absence of the NUP98 partner protein RAE1, and prevent the binding of the mitotic checkpoint complex to the APC/C(Cdc20). NUP98 oncoproteins require the GLEBS-like domain present in their NUP98 moiety to bind the APC/C(Cdc20). We found that NUP98 wild-type is a substrate of APC/C(Cdc20) prior to mitotic entry, and that its binding to APC/C(Cdc20) is controlled via phosphorylation of a PEST sequence located within its C-terminal portion. We identify S606, within the PEST sequence, as a key target site, whose phosphorylation modulates the capability of NUP98 to interact with APC/C(Cdc20). We finally provide evidence for an involvement of the peptidyl-prolyl isomerase PIN1 in modulating the possible conformational changes within NUP98 that lead to its dissociation from the APC/C(Cdc20) during mitosis. Our results provide novel insight into the mechanisms underlying the aberrant capability of NUP98 oncoproteins to interact with APC/C(Cdc20) and to interfere with its function. PMID:27097363

  19. Los1p, involved in yeast pre-tRNA splicing, positively regulates members of the SOL gene family

    SciTech Connect

    Shen, W.C.; Stanford, D.R.; Hopper, A.K.

    1996-06-01

    To understand the role of Los1p in pre-tRNA splicing, we sought los1 multicopy suppressors. We found SOL1 that suppresses both point and null LOS1 mutations. Since, when fused to the Gal4p DNA-binding domain, Los1p activates transcription, we tested whether Los1p regulates SOL1. We found that los1 mutants have depleted levels of SOL1 mRNA and Sol1p. Thus, LOS1 appears to positively regulate SOL1. SOL1 belongs to a multigene family with at least two additional members, SOL2 and SOL3. Sol proteins have extensive similarity to an unusual group of glucose-6-phosphate dehydrogenases (G6PDs). As the similarities are restricted to areas separate from the catalytic domain, these G6PDs may have more than one function. The SOL gene disruptions negatively affect tRNA-mediated nonsense suppression and the severity increases with the number of mutant SOL genes. However, tRNA levels do not vary with either multicopy SOL genes or with SOL disruptions. Therefore, the Sol proteins affect tRNA expression/function at steps other than transcription or splicing. We propose that LOS1 regulates gene products involved in tRNA expression/function as well as pre-tRNA splicing. 64 refs., 6 figs., 6 tabs.

  20. Association of APC, GSTP1 and SOCS1 promoter methylation with the risk of hepatocellular carcinoma: a meta-analysis.

    PubMed

    Liu, Meng; Cui, Lian-Hua; Li, Cheng-Cheng; Zhang, Li

    2015-11-01

    Studies of the relationships of adenomatous polyposis coli (APC), glutathione-S-transferase P1 (GSTP1) and suppressor of the cytokine signalling 1 (SOCS1) promoter region methylation with the risk of hepatocellular carcinoma (HCC) have yielded inconsistent results. We carried out the current meta-analysis to comprehensively assess the associations between APC, GSTP1 and SOCS1 promoter methylation frequency and the risk of HCC. All relevant reports were identified by searching the PubMed, Embase, Web of Science, CNKI and the Chinese BioMedical Literature databases before 1 March 2014, with restriction to articles published in the Chinese and English languages. Pooled odds ratios (ORs) and corresponding 95% confidence intervals (CIs) were calculated to investigate the rates of APC, GSTP1 and SOCS1 promoter methylation and the risk of HCC. Our meta-analysis identified relationships of APC (12 studies with 592 HCC tumour tissues), GSTP1 (14 studies including 646 HCC tumour tissues) and SOCS1 (11 studies with 512 HCC tumour tissues) promoter methylation with the risk of HCC. Compared with paracancerous tissues, the pooled ORs of APC, GSTP1 and SOCS1 promoter region methylation in HCC cancer tissues were 5.32 (95% CI=2.96-9.56), 5.65, (95% CI=3.41-9.35) and 2.73 (95% CI=1.37-5.44), respectively. Compared with normal liver tissues as controls, the pooled ORs of APC, GSTP1 and SOCS1 promoter region methylation in HCC cancer tissues were 20.43 (95% CI=5.56-75.08), 18.78 (95% CI=5.76-61.19) and 13.00 (95% CI=5.20-32.47), respectively. Subgroup analysis by ethnicity showed that APC, GSTP1 and SOCS1 promoter methylation was associated significantly with the risk of HCC in both Asian and White populations (all P<0.05). Our meta-analysis suggested strong associations between APC, GSTP1 and SOCS1 gene promoter methylation and the risk of HCC, suggesting these to be promising biomarkers for HCC. PMID:25853848

  1. Cell-Type Specific Expression of Apc in Lung Development, Injury and Repair

    PubMed Central

    Li, Aimin; Xing, Yiming; Chan, Belinda; Heisterkamp, Nora; Groffen, John; Borok, Zea; Minoo, Parviz; Li, Changgong

    2010-01-01

    Adenomatous polyposis coli (Apc) is critical for Wnt signaling and cell migration. The current study examined Apc expression during lung development, injury and repair. Apc was first detectable in smooth muscle layers in early lung morphogenesis, and was highly expressed in ciliated and neuroendocrine cells in the advanced stages. No Apc immunoreactivity was detected in Clara or basal cells, which function as stem/progenitor cell in adult lung. In ciliated cells, Apc is associated mainly with apical cytoplasmic domain. In response to naphthalene induced injury, Apcpositive cells underwent squamous metaplasia, accompanied by changes in Apc subcellular distribution. In conclusion, both spatial and temporal expression of Apc is dynamically regulated during lung development and injury repair. Differential expression of Apc in progenitor vs. non-progenitor cells suggests a functional role in cell type specification. Subcellular localization changes of Apc in response to naphthalene injury suggest a role in cell shape and cell migration. PMID:20658693

  2. Targeted apc;twist double-mutant mice: a new model of spontaneous osteosarcoma that mimics the human disease.

    PubMed

    Entz-Werlé, Natacha; Choquet, Philippe; Neuville, Agnès; Kuchler-Bopp, Sabine; Clauss, François; Danse, Jean-Marc; Simo-Noumbissie, Pauline; Guérin, Eric; Gaub, Marie-Pierre; Freund, Jean-Noel; Boehm, Nelly; Constantinesco, André; Lutz, Patrick; Guenot, Dominique; Perrin-Schmitt, Fabienne

    2010-01-01

    TWIST and adenomatosis polyposis coli (APC) are critical signaling factors in normal bone development. In previous studies examining a homogeneously treated cohort of pediatric osteosarcoma patients, we reported the frequent and concurrent loss of both TWIST and APC genes. On these bases, we created a related animal model to further explore the oncogenic cooperation between these two genes. We performed intercrosses between twist-null/+ and Apc1638N/+ mice and studied their progeny. The Apc1638N/+;twistnull/+ mice developed bone abnormalities observed by macroscopic skeletal analyses and in vivo imaging. Complementary histologic, cellular, and molecular analyses were used to characterize the identified bone tumors, including cell culture and immunofluorescence of bone differentiation markers. Spontaneous localized malignant bone tumors were frequently identified in Apc1638N/+;twist-null/+ mice by in vivo imaging evaluation and histologic analyses. These tumors possessed several features similar to those observed in human localized osteosarcomas. In particular, the murine tumors presented with fibroblastic, chondroblastic, and osteoblastic osteosarcoma histologies, as well as mixtures of these subtypes. In addition, cellular analyses and bone differentiation markers detected by immunofluorescence on tumor sections reproduced most murine and human osteosarcoma characteristics. For example, the early bone differentiation marker Runx2, interacting physically with hypophosphorylated pRb, was undetectable in these murine osteosarcomas, whereas phosphorylated retinoblastoma was abundant in the osteoblastic and chondroblastic tumor subtypes. These characteristics, similar to those observed in human osteosarcomas, indicated that our animal model may be a powerful tool to further understand the development of localized osteosarcoma. PMID:21151473

  3. Targeted Apc;Twist Double-Mutant Mice: A New Model of Spontaneous Osteosarcoma That Mimics the Human Disease123

    PubMed Central

    Entz-Werlé, Natacha; Choquet, Philippe; Neuville, Agnès; Kuchler-Bopp, Sabine; Clauss, François; Danse, Jean-Marc; Simo-Noumbissie, Pauline; Guérin, Eric; Gaub, Marie-Pierre; Freund, Jean-Noel; Boehm, Nelly; Constantinesco, André; Lutz, Patrick; Guenot, Dominique; Perrin-Schmitt, Fabienne

    2010-01-01

    TWIST and adenomatosis polyposis coli (APC) are critical signaling factors in normal bone development. In previous studies examining a homogeneously treated cohort of pediatric osteosarcoma patients, we reported the frequent and concurrent loss of both TWIST and APC genes. On these bases, we created a related animal model to further explore the oncogenic cooperation between these two genes. We performed intercrosses between twist-null/+ and Apc1638N/+ mice and studied their progeny. The Apc1638N/+;twistnull/+ mice developed bone abnormalities observed by macroscopic skeletal analyses and in vivo imaging. Complementary histologic, cellular, and molecular analyses were used to characterize the identified bone tumors, including cell culture and immunofluorescence of bone differentiation markers. Spontaneous localized malignant bone tumors were frequently identified in Apc1638N/+;twist-null/+ mice by in vivo imaging evaluation and histologic analyses. These tumors possessed several features similar to those observed in human localized osteosarcomas. In particular, the murine tumors presented with fibroblastic, chondroblastic, and osteoblastic osteosarcoma histologies, as well as mixtures of these subtypes. In addition, cellular analyses and bone differentiation markers detected by immunofluorescence on tumor sections reproduced most murine and human osteosarcoma characteristics. For example, the early bone differentiation marker Runx2, interacting physically with hypophosphorylated pRb, was undetectable in these murine osteosarcomas, whereas phosphorylated retinoblastoma was abundant in the osteoblastic and chondroblastic tumor subtypes. These characteristics, similar to those observed in human osteosarcomas, indicated that our animal model may be a powerful tool to further understand the development of localized osteosarcoma. PMID:21151473

  4. Human Immunodeficiencies Related to Defective APC/T Cell Interaction

    PubMed Central

    Kallikourdis, Marinos; Viola, Antonella; Benvenuti, Federica

    2015-01-01

    The primary event for initiating adaptive immune responses is the encounter between T lymphocytes and antigen presenting cells (APCs) in the T cell area of secondary lymphoid organs and the formation of highly organized intercellular junctions referred to as immune synapses (IS). In vivo live-cell imaging of APC–T cell interactions combined to functional studies unveiled that T cell fate is dictated, in large part, by the stability of the initial contact. Immune cell interaction is equally important during delivery of T cell help to B cells and for the killing of target cells by cytotoxic T cells and NK cells. The critical role of contact dynamics and synapse stability on the immune response is well illustrated by human immune deficiencies in which disease pathogenesis is linked to altered adhesion or defective cross-talk between the synaptic partners. The Wiskott–Aldrich syndrome (WAS) is a severe primary immunodeficiency caused by mutations in the Wiskott–Aldrich syndrome protein (WASp), a scaffold that promotes actin polymerization and links TCR stimulation to T cell activation. Absence or mutations in WASp affects intercellular APC–T cell communications by interfering with multiple mechanisms on both sides of the IS. The warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome is caused by mutations in CXCR4, a chemokine receptor that in mutant form leads to impairment of APC–T cell interactions. Present evidences suggest that other recently characterized primary immune deficiencies caused by mutation in genes linked to actin cytoskeletal reorganization, such as WIP and DOCK8, may also depend on altered synapse stability. Here, we will discuss in details the mechanisms of disturbed APC–T cell interactions in WAS and WHIM. Moreover, we will summarize the evidence pointing to a compromised conjugate formation in WIP, DOCK8, and X-linked lymphoproliferative syndrome. PMID:26379669

  5. Dietary Methyl Donor Depletion Protects Against Intestinal Tumorigenesis in ApcMin/+ Mice

    PubMed Central

    Kadaveru, Krishna; Protiva, Petr; Greenspan, Emily J; Kim, Young-In; Rosenberg, Daniel W

    2012-01-01

    Despite recent population data, the influence of dietary folate supplementation on colon cancer risk remains controversial. This study examines the effects of folate deficiency, in combination with choline, methionine and vitamin B12 depletion, on intestinal tumorigenesis in ApcMin/+ mice. Methyl donor sufficient (MDS) and deficient (MDD) diets were started at 5 or 10 weeks of age and tumors evaluated at 16 weeks. MDD suppressed intestinal tumor formation in ApcMin/+ mice (~80%) when started at 5 weeks of age. The protective effect was lost when MDD was initiated at 10 weeks of age, indicating an important time-dependency on cancer suppression. Concomitant with cancer protection, MDD restricted body weight gain. Therefore, a second study was conducted in which MDS was given ad libitum or pair-fed with MDD. While small intestinal tumors were reduced 54% in pair-fed MDS mice, MDD caused a further reduction (96%). In colon, although MDD did not affect tumor numbers, tumor size was reduced. Gene expression profiling of normal-appearing colonic mucosa after 11 weeks on MDD identified a total of 493 significantly down-regulated genes relative to the MDS group. Pathway analysis placed many of these genes within general categories of inflammatory signaling and cell cycle regulation, consistent with recently published human data obtained during folate depletion (1). Further studies are warranted to investigate the complex interplay of methyl donor status and cancer protection in high-risk populations. PMID:22677908

  6. Cdh1 regulates craniofacial development via APC-dependent ubiquitination and activation of Goosecoid.

    PubMed

    Shao, Rui; Liu, Jia; Yan, Guang; Zhang, Jinfang; Han, Yujiao; Guo, Jianfeng; Xu, Zhan; Yuan, Zhu; Liu, Jiankang; Malumbres, Marcos; Wan, Lixin; Wei, Wenyi; Zou, Weiguo

    2016-06-01

    Craniofacial anomalies (CFAs) characterized by birth defects of skull and facial bones are the most frequent congenital disease. Genomic analysis has identified multiple genes responsible for CFAs; however, the underlying genetic mechanisms for the majority of CFAs remain largely unclear. Our previous study revealed that the Wwp2 E3 ubiquitin ligase facilitates craniofacial development in part through inducing monoubiquitination and activation of the paired-like homeobox transcription factor, Goosecoid (Gsc). Here we report that Gsc is also ubiquitinated and activated by the APC(Cdh1) E3 ubiquitin ligase, leading to transcriptional activation of various Gsc target genes crucial for craniofacial development. Consistenly, neural crest-specific Cdh1-knockout mice display similar bone malformation as Wwp2-deficient mice in the craniofacial region, characterized by a domed skull, a short snout and a twisted nasal bone. Mechanistically, like Wwp2-deficient mice, mice with Cdh1 deficiency in neural crest cells exhibit reduced Gsc/Sox6 transcriptional activities. Simultaneous deletion of Cdh1 and Wwp2 results in a more severe craniofacial defect compared with single gene deletion, suggesting a synergistic augmentation of Gsc activity by these two E3 ubiquitin ligases. Hence, our study reveals a novel role for Cdh1 in craniofacial development through promoting APC-dependent non-proteolytic ubiquitination and activation of Gsc. PMID:27126000

  7. The tumor suppressor APC differentially regulates multiple β-catenins through the function of axin and CKIα during C. elegans asymmetric stem cell divisions.

    PubMed

    Baldwin, Austin T; Phillips, Bryan T

    2014-06-15

    The APC tumor suppressor regulates diverse stem cell processes including gene regulation through Wnt-β-catenin signaling and chromosome stability through microtubule interactions, but how the disparate functions of APC are controlled is not well understood. Acting as part of a Wnt-β-catenin pathway that controls asymmetric cell division, Caenorhabditis elegans APC, APR-1, promotes asymmetric nuclear export of the β-catenin WRM-1 by asymmetrically stabilizing microtubules. Wnt function also depends on a second β-catenin, SYS-1, which binds to the C. elegans TCF POP-1 to activate gene expression. Here, we show that APR-1 regulates SYS-1 levels in asymmetric stem cell division, in addition to its known role in lowering nuclear levels of WRM-1. We demonstrate that SYS-1 is also negatively regulated by the C. elegans homolog of casein kinase 1α (CKIα), KIN-19. We show that KIN-19 restricts APR-1 localization, thereby regulating nuclear WRM-1. Finally, the polarity of APR-1 cortical localization is controlled by PRY-1 (C. elegans Axin), such that PRY-1 controls the polarity of both SYS-1 and WRM-1 asymmetries. We propose a model whereby Wnt signaling, through CKIα, regulates the function of two distinct pools of APC - one APC pool negatively regulates SYS-1, whereas the second pool stabilizes microtubules and promotes WRM-1 nuclear export. PMID:24762815

  8. High performance APCS conceptual design and evaluation scoping study

    SciTech Connect

    Soelberg, N.; Liekhus, K.; Chambers, A.; Anderson, G.

    1998-02-01

    This Air Pollution Control System (APCS) Conceptual Design and Evaluation study was conducted to evaluate a high-performance (APC) system for minimizing air emissions from mixed waste thermal treatment systems. Seven variations of high-performance APCS designs were conceptualized using several design objectives. One of the system designs was selected for detailed process simulation using ASPEN PLUS to determine material and energy balances and evaluate performance. Installed system capital costs were also estimated. Sensitivity studies were conducted to evaluate the incremental cost and benefit of added carbon adsorber beds for mercury control, specific catalytic reduction for NO{sub x} control, and offgas retention tanks for holding the offgas until sample analysis is conducted to verify that the offgas meets emission limits. Results show that the high-performance dry-wet APCS can easily meet all expected emission limits except for possibly mercury. The capability to achieve high levels of mercury control (potentially necessary for thermally treating some DOE mixed streams) could not be validated using current performance data for mercury control technologies. The engineering approach and ASPEN PLUS modeling tool developed and used in this study identified APC equipment and system performance, size, cost, and other issues that are not yet resolved. These issues need to be addressed in feasibility studies and conceptual designs for new facilities or for determining how to modify existing facilities to meet expected emission limits. The ASPEN PLUS process simulation with current and refined input assumptions and calculations can be used to provide system performance information for decision-making, identifying best options, estimating costs, reducing the potential for emission violations, providing information needed for waste flow analysis, incorporating new APCS technologies in existing designs, or performing facility design and permitting activities.

  9. Tumor suppressor gene adenomatous polyposis coli downregulates intestinal transport.

    PubMed

    Rexhepaj, Rexhep; Rotte, Anand; Gu, Shuchen; Michael, Diana; Pasham, Venkanna; Wang, Kan; Kempe, Daniela S; Ackermann, Teresa F; Brücher, Björn; Fend, Falko; Föller, Michael; Lang, Florian

    2011-05-01

    Loss of function mutations of the tumor suppressor gene adenomatous polyposis coli (APC) underly the familial adenomatous polyposis. Mice carrying an inactivating mutation in the apc gene (apc (Min/+)) similarly develop intestinal polyposis. APC is effective at least in part by degrading β-catenin and lack of APC leads to markedly enhanced cellular β-catenin levels. β-Catenin has most recently been shown to upregulate the Na+/K+ ATPase. The present study, thus, explored the possibility that APC could influence intestinal transport. The abundance and localization of β-catenin were determined utilizing Western blotting and confocal microscopy, the activity of the electrogenic glucose carrier (SGLT1) was estimated from the glucose-induced current in jejunal segments utilizing Ussing chamber experiments and the Na+/H+ exchanger (NHE3) activity from Na+ -dependent re-alkalinization of cytosolic pH (ΔpH(i)) following an ammonium pulse employing BCECF fluorescence. As a result, β-catenin abundance in intestinal tissue was significantly higher in apc (Min/+) mice than in wild-type mice (apc (+/+)). The β-catenin protein was localized in the basolateral membrane. Both, the glucose-induced current and ΔpH(i) were significantly higher in apc (Min/+) mice than in apc (+/+) mice. In conclusion, intestinal electrogenic transport of glucose and intestinal Na+/H+ exchanger activity are both significantly enhanced in apc (Min/+) mice, pointing to a role of APC in the regulation of epithelial transport. PMID:21476133

  10. Chlorinated Water Modulates the Development of Colorectal Tumors with Chromosomal Instability and Gut Microbiota in Apc-Deficient Mice

    PubMed Central

    Sasada, Tatsunari; Hinoi, Takao; Saito, Yasufumi; Adachi, Tomohiro; Takakura, Yuji; Kawaguchi, Yasuo; Sotomaru, Yusuke; Sentani, Kazuhiro; Oue, Naohide; Yasui, Wataru; Ohdan, Hideki

    2015-01-01

    The gastrointestinal tract is continuously exposed to a variety of chemicals and commensal bacteria. Recent studies have shown that changes in gut microbial populations caused by chlorine or other chemicals in the drinking water influence the development of human colorectal cancer, although the mechanism of tumorigenesis in the gut epithelium is obfuscated by the diversity of microflora and complexity of the tumor microenvironment. In this regard, mouse models that recapitulate human colorectal cancer are an invaluable tool. In this study, we used two conditional adenomatous polyposis coli (Apc) knockout mouse models to investigate the effect of chlorinated water on tumorigenesis in the digestive tract. Mice with colon-specific carcinoma—caused by either chromosomal (CDX2P 9.5-NLS Cre;Apc+/flox, abbreviated to CPC;Apc) or microsatellite (CDX2P9.5-G19Cre;Apcflox/flox and CDX2P9.5-G22Cre;Apcflox/flox) instability, respectively—were administered chlorinated (10.0 mg/L chlorine) or tap (0.7 mg/L chlorine) water and evaluated for colon polyp formation. In CPC;Apc mice given chlorinated drinking water, tumors tended to develop in the colon, whereas in those that drank tap water, tumors were mostly observed in the small intestine. There was no difference in the rate of tumor formation of CDX2P9.5-G19Cre;Apcflox/flox and CDX2P9.5-G22Cre;Apcflox/flox mice consuming chlorinated as compared to tap water, suggesting that microsatellite instability in the Apc gene does not significantly affect tumorigenesis. Chlorinated water altered the enteric environment by reducing the fecal populations of the obligatory anaerobes Clostridium perfringens and C. difficile, as well as species belonging to the Atopobium cluster, including Enterobacteriaceae and Staphylococcus sp., which was associated with colon tumorigenesis in CPC;Apc mice. These results suggest that differences in tumorigenesis among CPC;Apc mice consuming chlorinated versus tap water may be due to differences in

  11. The E3 ligase APC/C(Cdh1) promotes ubiquitylation-mediated proteolysis of PAX3 to suppress melanocyte proliferation and melanoma growth.

    PubMed

    Cao, Juxiang; Dai, Xiangpeng; Wan, Lixin; Wang, Hongshen; Zhang, Jinfang; Goff, Philip S; Sviderskaya, Elena V; Xuan, Zhenyu; Xu, Zhixiang; Xu, Xiaowei; Hinds, Philip; Flaherty, Keith T; Faller, Douglas V; Goding, Colin R; Wang, Yongjun; Wei, Wenyi; Cui, Rutao

    2015-09-01

    The anaphase-promoting complex or cyclosome with the subunit Cdh1 (APC/C(Cdh1)) is an E3 ubiquitin ligase involved in the control of the cell cycle. Here, we identified sporadic mutations occurring in the genes encoding APC components, including Cdh1, in human melanoma samples and found that loss of APC/C(Cdh1) may promote melanoma development and progression, but not by affecting cell cycle regulatory targets of APC/C. Most of the mutations we found in CDH1 were those associated with ultraviolet light (UV)-induced melanomagenesis. Compared with normal human skin tissue and human or mouse melanocytes, the abundance of Cdh1 was decreased and that of the transcription factor PAX3 was increased in human melanoma tissue and human or mouse melanoma cell lines, respectively; Cdh1 abundance was further decreased with advanced stages of human melanoma. PAX3 was a substrate of APC/C(Cdh1) in melanocytes, and APC/C(Cdh1)-mediated ubiquitylation marked PAX3 for proteolytic degradation in a manner dependent on the D-box motif in PAX3. Either mutating the D-box in PAX3 or knocking down Cdh1 prevented the ubiquitylation and degradation of PAX3 and increased proliferation and melanin production in melanocytes. Knocking down Cdh1 in melanoma cells in culture or before implantation in mice promoted doxorubicin resistance, whereas reexpressing wild-type Cdh1, but not E3 ligase-deficient Cdh1 or a mutant that could not interact with PAX3, restored doxorubicin sensitivity in melanoma cells both in culture and in xenografts. Thus, our findings suggest a tumor suppressor role for APC/C(Cdh1) in melanocytes and that targeting PAX3 may be a strategy for treating melanoma. PMID:26329581

  12. Belgian MSWI fly ashes and APC residues: a characterisation study.

    PubMed

    De Boom, Aurore; Degrez, Marc

    2012-06-01

    Municipal Solid Waste Incineration (MSWI) produces different sorts of residues, bottom ash, fly ashes and Air Pollution Control (APC) residues. Generally, fly ashes and APC residues are mixed at the MSWI plant and manage as a sole residue. In this study, fly ashes and APC residues have been sampled separately at different Belgian MSWI plant and analysed by X-ray fluorescence in order to highlight the composition differences that may appear between the solids. Ca and Cl are found to be the major elements in most of the samples. Lithophilic elements, such as Al and Si, are richer in furnace and boiler ashes, as can be expected. Leaching tests also show differences between the residues; leachates from furnace and boiler ashes are alkaline while those from bag filter residues present a pH value of 6, which impacts the leaching of heavy metals (Pb and Zn). The results suggest that it could be advantageous to manage fly ashes and APC residues separately by adjusting the treatment to their specificities. PMID:22244614

  13. Timing of APC/C substrate degradation is determined by fzy/fzr specificity of destruction boxes

    PubMed Central

    Zur, Amit; Brandeis, Michael

    2002-01-01

    The anaphase promoting complex/cyclosome (APC/C), activated by fzy and fzr, degrades cell cycle proteins that carry RXXL or KEN destruction boxes (d-boxes). APC/C substrates regulate sequential events and must be degraded in the correct order during mitosis and G1. We studied how d-boxes determine APC/Cfzy/APC/Cfzr specificity and degradation timing. Cyclin B1 has an RXXL box and is degraded by both APC/Cfzy and APC/Cfzr; fzy has a KEN box and is degraded by APC/Cfzr only. We characterized the degradation of substrates with swapped d-boxes. Cyclin B1 with KEN was degraded by APC/Cfzr only. Fzy with RXXL could be degraded by APC/Cfzy and APC/Cfzr. Interestingly, APC/Cfzy- but not APC/Cfzr-specific degradation is highly dependent on the location of RXXL. We studied degradation of tagged substrates in real time and observed that APC/Cfzr is activated in early G1. These observations demonstrate how d-box specificities of APC/Cfzy and APC/Cfzr, and the successive activation of APC/C by fzy and fzr, establish the temporal degradation pattern. Our observations can explain further why some endogenous RXXL substrates are degraded by APC/Cfzy, while others are restricted to APC/Cfzr. PMID:12198152

  14. Redefining the subcellular location and transport of APC: new insights using a panel of antibodies

    PubMed Central

    Brocardo, Mariana; Näthke, Inke S; Henderson, Beric R

    2005-01-01

    Adenomatous polyposis coli (APC) is a tumour suppressor involved in colon cancer progression. We and others previously described nuclear–cytoplasmic shuttling of APC. However, there are conflicting reports concerning the localization of endogenous wild-type and tumour-associated, truncated APC. To resolve this issue, we compared APC localization using immunofluorescence (IF) microscopy and cell fractionation with nine different APC antibodies. We found that three commonly used APC antibodies showed nonspecific nuclear staining by IF and validated this conclusion in cells where APC was inactivated using small interfering RNA or Cre/Flox. Fractionation showed that wild-type and truncated APC from colon cancer cells were primarily cytoplasmic, but increased in the nucleus after leptomycin B treatment, consistent with CRM1-dependent nuclear export. In contrast to recent reports, our biochemical data indicate that APC nuclear localization is not regulated by changes in cell density, and that APC nuclear export is not prevented by truncating mutations in cancer. These results verify that the bulk of APC resides in the cytoplasm and indicate the need for caution when evaluating the nuclear accumulation of APC. PMID:15678162

  15. 42 CFR 419.31 - Ambulatory payment classification (APC) system and payment weights.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 42 Public Health 3 2010-10-01 2010-10-01 false Ambulatory payment classification (APC) system and... Outpatient Services § 419.31 Ambulatory payment classification (APC) system and payment weights. (a) APC... OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICARE PROGRAM PROSPECTIVE PAYMENT SYSTEM FOR...

  16. Apc-driven colon carcinogenesis in Pirc rat is strongly reduced by polyethylene glycol.

    PubMed

    Femia, Angelo Pietro; Becherucci, Caterina; Crucitta, Stefania; Caderni, Giovanna

    2015-11-01

    Polyethylene glycol (PEG) is one of the most powerful agents in reducing chemically induced carcinogenesis in rat colon. However, contrasting results in Min mice dampened the enthusiasm on this potentially strong and virtually safe, cancer chemopreventing agent. Pirc (F344/NTac-Apc (am1137) ) rats carrying a germline heterozygous mutation in the Apc gene, spontaneously develop multiple tumours in the colon thus modelling both familial adenomatous polyposis (FAP) and sporadic colorectal cancer (CRC). Given this similarity, we thought that these rats could be appropriate to test the efficacy of PEG 8000 in reducing carcinogenesis. Pirc male rats aged one month were treated with 5% PEG in drinking water for 2 or 6 months. Precancerous lesions were dramatically reduced after 2 months of PEG treatment (Mucin depleted foci (MDF)/colon were 99 ± 17 and 12 ± 8 in Controls and PEG-treated rats, respectively; p < 0.001; mean ± SD). Similarly, colon tumors were significantly reduced after 6 months of treatment (tumors/rat were 8.1 ± 2.3 and 3.6 ± 2.2 in Controls and PEG-treated rats, respectively; p < 0.05; mean ± SD). Colon proliferation, a parameter correlated to cancer risk, was also significantly lower in PEG-treated rats than in Controls, while apoptosis was not significantly affected. In conclusion, PEG markedly reduces colon carcinogenesis in Pirc rats mutated in Apc; we thus suggest that PEG may be used as chemopreventive agent to reduce cancer risk in FAP and CRC patients. PMID:25912754

  17. Dietary acrylamide intake and the risk of colorectal cancer with specific mutations in KRAS and APC.

    PubMed

    Hogervorst, Janneke G F; de Bruijn-Geraets, Daisy; Schouten, Leo J; van Engeland, Manon; de Kok, Theo M C M; Goldbohm, R Alexandra; van den Brandt, Piet A; Weijenberg, Matty P

    2014-05-01

    Acrylamide, a probable human carcinogen, is present in heat-treated carbohydrate-rich foods. Epidemiological studies have not shown a clear association between acrylamide intake and colorectal cancer (CRC) risk. This may be due to the molecular heterogeneity in colorectal tumors, which was not taken into consideration before. Since the acrylamide metabolite glycidamide induces specific DNA mutations in rodents, we investigated whether acrylamide is associated with CRC risk characterized by mutations in Kirsten-ras (KRAS) and adenomatous polyposis coli (APC); key genes in colorectal carcinogenesis. This case-cohort analysis, within the Netherlands Cohort Study on diet and cancer, was based on 7.3 years of follow-up. Acrylamide intake was assessed with a food frequency questionnaire. Mutation analysis of codons 1286-1520 in exon 15 in APC and codons 12 and 13 in exon 1 in KRAS was performed on tumor tissue of 733 cases. Hazard ratios (HR) were calculated using Cox proportional hazards analysis. Among men, acrylamide intake was statistically significantly associated with an increased risk of particularly tumors with an activating KRAS mutation {HR fourth versus first quartile: 2.12 [95% confidence interval (CI): 1.16-3.87], P trend: 0.01}. Among women, acrylamide intake was statistically significantly associated with a decreased risk of particularly tumors with a truncating APC mutation (fourth versus first quartile: 0.47 (95% CI: 0.23-0.94), P trend: 0.02), but only in the highest quartile of intake. This is the first study to show that acrylamide might be associated with CRC with specific somatic mutations, differentially in men and women. More research is needed to corroborate or refute these findings. PMID:24398672

  18. The ABBA motif binds APC/C activators and is shared by APC/C substrates and regulators.

    PubMed

    Di Fiore, Barbara; Davey, Norman E; Hagting, Anja; Izawa, Daisuke; Mansfeld, Jörg; Gibson, Toby J; Pines, Jonathon

    2015-02-01

    The anaphase-promoting complex or cyclosome (APC/C) is the ubiquitin ligase that regulates mitosis by targeting specific proteins for degradation at specific times under the control of the spindle assembly checkpoint (SAC). How the APC/C recognizes its different substrates is a key problem in the control of cell division. Here, we have identified the ABBA motif in cyclin A, BUBR1, BUB1, and Acm1, and we show that it binds to the APC/C coactivator CDC20. The ABBA motif in cyclin A is required for its proper degradation in prometaphase through competing with BUBR1 for the same site on CDC20. Moreover, the ABBA motifs in BUBR1 and BUB1 are necessary for the SAC to work at full strength and to recruit CDC20 to kinetochores. Thus, we have identified a conserved motif integral to the proper control of mitosis that connects APC/C substrate recognition with the SAC. PMID:25669885

  19. Single-molecule spectroscopy and femtosecond transient absorption studies on the excitation energy transfer process in ApcE(1-240) dimers.

    PubMed

    Long, Saran; Zhou, Meng; Tang, Kun; Zeng, Xiao-Li; Niu, Yingli; Guo, Qianjin; Zhao, Kai-Hong; Xia, Andong

    2015-05-28

    ApcE(1-240) dimers with one intrinsic phycocyanobilin (PCB) chromophore in each monomer that is truncated from the core-membrane linker (ApcE) of phycobilisomes (PBS) in Nostoc sp. PCC 7120 show a sharp and significantly red-shifted absorption. Two explanations either conformation-dependent Förster resonance energy transfer (FRET) or the strong exciton coupling limit have been proposed for red-shifted absorption. This is a classic example of the special pair in the photosynthetic light harvesting proteins, but the mechanism of this interaction is still a matter of intense debate. We report the studies using single-molecule and transient absorption spectra on the interaction in the special pair of ApcE dimers. Our results demonstrate the presence of conformation-dependent FRET between the two PCB chromophores in ApcE dimers. The broad distributions of fluorescence intensities, lifetimes and polarization difference from single-molecule measurements reveal the heterogeneity of local protein-pigment environments in ApcE dimers, where the same molecular structures but different protein environments are the main reason for the two PCB chromophores with different spectral properties. The excitation energy transfer rate between the donor and the acceptor about (110 ps)(-1) is determined from transient absorption measurements. The red-shifted absorption in ApcE dimers could result from more extending conformation, which shows another type of absorption redshift that does not depend on strong exciton coupling. The results here stress the importance of conformation-controlled spectral properties of the chemically identical chromophores, which could be a general feature to control energy/electron transfer, widely existing in the light harvesting complexes. PMID:25925197

  20. Comprehensive proteome analysis of an Apc mouse model uncovers proteins associated with intestinal tumorigenesis.

    PubMed

    Hung, Kenneth E; Faca, Vitor; Song, Kenneth; Sarracino, David A; Richard, Larissa Georgeon; Krastins, Bryan; Forrester, Sara; Porter, Andrew; Kunin, Alexandra; Mahmood, Umar; Haab, Brian B; Hanash, Samir M; Kucherlapati, Raju

    2009-03-01

    Tumor-derived proteins may occur in the circulation as a result of secretion, shedding from the cell surface, or cell turnover. We have applied an in-depth comprehensive proteomic strategy to plasma from intestinal tumor-bearing Apc mutant mice to identify proteins associated with tumor development. We used quantitative tandem mass spectrometry of fractionated mouse plasma to identify differentially expressed proteins in plasma from intestinal tumor-bearing Apc mutant mice relative to matched controls. Up-regulated proteins were assessed for the expression of corresponding genes in tumor tissue. A subset of proteins implicated in colorectal cancer were selected for further analysis at the tissue level using antibody microarrays, Western blotting, tumor immunohistochemistry, and novel fluorescent imaging. We identified 51 proteins that were elevated in plasma with concordant up-regulation at the RNA level in tumor tissue. The list included multiple proteins involved in colon cancer pathogenesis: cathepsin B and cathepsin D, cullin 1, Parkinson disease 7, muscle pyruvate kinase, and Ran. Of these, Parkinson disease 7, muscle pyruvate kinase, and Ran were also found to be up-regulated in human colon adenoma samples. We have identified proteins with direct relevance to colorectal carcinogenesis that are present both in plasma and in tumor tissue in intestinal tumor-bearing mice. Our results show that integrated analysis of the plasma proteome and tumor transcriptome of genetically engineered mouse models is a powerful approach for the identification of tumor-related plasma proteins. PMID:19240248

  1. Lovastatin, but not orlistat, reduces intestinal polyp volume in an ApcMin/+ mouse model.

    PubMed

    Notarnicola, Maria; Barone, Michele; Francavilla, Antonio; Tutino, Valeria; Bianco, Giusy; Tafaro, Angela; Minoia, Mario; Polimeno, Lorenzo; Napoli, Anna; Scavo, Maria Principia; Caruso, Maria Gabriella

    2016-08-01

    The statins, inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCoAR) and orlistat, an inhibitor of fatty acid synthase (FAS), inhibit tumor cell growth by restricting cholesterol and fatty acid synthesis, respectively. We previously demonstrated that an omega (ω)-3 polyunsaturated fatty acid (PUFA)- or olive oil-enriched diet reduced the polyp number and volume in ApcMin/+ mice. This phenomenon was associated with a significant inhibition of FAS and HMGCoAR, as well as an increase in the estrogen receptor (ER)β/α ratio. Herein, we evaluated the effect of lovastatin and orlistat on polyp development and ER expression in ApcMin/+ mice, in order to confirm previous data obtained with ω‑3-PUFAs and olive oil. As expected, the use of lovastatin and orlistat significantly reduced HMGCoAR and FAS enzymatic activities and gene expression in colonic tissues, but did not affect the number of intestinal polyps, while there was a statistically significant reduction in polyp volume only in the mouse group treated with lovastatin. In the mice receiving orlistat, we observed a significant increase in cell proliferation in the polyp tissue, as well as enhanced expression of ERα. Moreover, the overexpression of ERα was associated with a statistically significant increase in PES1, Shh and Gli1 protein levels, considered ERα-related molecular targets. PMID:27277576

  2. Cellular Factors Targeting APCs to Modulate Adaptive T Cell Immunity

    PubMed Central

    Do, Jeongsu; Min, Booki

    2014-01-01

    The fate of adaptive T cell immunity is determined by multiple cellular and molecular factors, among which the cytokine milieu plays the most important role in this process. Depending on the cytokines present during the initial T cell activation, T cells become effector cells that produce different effector molecules and execute adaptive immune functions. Studies thus far have primarily focused on defining how these factors control T cell differentiation by targeting T cells themselves. However, other non-T cells, particularly APCs, also express receptors for the factors and are capable of responding to them. In this review, we will discuss how APCs, by responding to those cytokines, influence T cell differentiation and adaptive immunity. PMID:25126585

  3. 75 FR 78246 - Medicare Program; Re-Chartering of the Advisory Panel on Ambulatory Payment Classification (APC...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-12-15

    ... Charter effective through November 21, 2012. FOR FURTHER INFORMATION CONTACT: Shirl Ackerman-Ross, (410... new technology APCs to clinical APCs). Evaluating APC group weights. Reviewing packaging the cost of... methodology for packaging and the impact of packaging on APC group structure and payment. Removing...

  4. APC/C is an essential regulator of centrosome clustering.

    PubMed

    Drosopoulos, Konstantinos; Tang, Chan; Chao, William C H; Linardopoulos, Spiros

    2014-01-01

    Centrosome amplification has been extensively associated with cancer. Cancer cells with extra centrosomes have the ability to cluster the extra centrosomes and divide in a bipolar fashion. Although a number of proteins have been shown to be involved in centrosome clustering, a mechanistic understanding of how this process is coordinated is not yet well defined. Here, to reveal regulators of centrosome clustering, we perform small interfering RNA (siRNA) screens with multiple assay readouts in a human isogenic cellular model. We find that APC/C activity is essential for centrosome clustering. We show that the motor kinesin Eg5 is a substrate of APC/C-CDH1, and that inhibition of APC/C results in stabilization of Eg5. Increased Eg5 protein levels disturb the balance of forces on the spindle and prevent centrosome clustering. This process is completely reversed after a short treatment with the Eg5 inhibitor, monastrol. These data advance our understanding of the regulation of centrosome clustering. PMID:24751481

  5. Multiloop photolithography control using hierarchical context information for APC models

    NASA Astrophysics Data System (ADS)

    Stuber, John D.

    2003-06-01

    Automated process control loops running in semiconductor manufacturing facilities must be able to compensate for machine variations as well as identify differences between products. With a number of exposure tools, pattern levels, and active devices manufactured in a typical ASIC fab, for photo APC system must maintain thousands of control loops. Control loop context information in TI's Semiconductor Manufacturing System (SMS) is defined in a hierarchal fashion which allows default values in a manufacturing specification to be overridden for particular products or lots. ProcessWORKS APC software automatically adds new control loops for new devices and pattern levels to a defined model structure. This system scales well and supports hundreds of thousands of control loops from a single database server in TI fabs. Application of product specific control systems for alignment and exposure control has provided increased exposure capacity due to decreased reworks and setup time, a substantial reduction in engineering maintenance and improved process capability. The APC system has evolved into a requirement for leading edge photolithography processes in Texas Instruments.

  6. Intestinal Peyer's patches prevent tumorigenesis in Apc (Min/+) mice.

    PubMed

    Fujimoto, Kyoko; Fujii, Gen; Sakurai, Hitomi; Yoshitome, Hiroko; Mutoh, Michihiro; Wada, Morimasa

    2015-01-01

    Peyer's patches are nodules that play a central role in intestinal immunity. Few studies demonstrate the relationship between the number of Peyer's patches and intestinal polyps. Here we identify a statistically significant inverse correlation between the quantity of Peyer's patches and of the development of intestinal polyps in Apc (Min/+) mice, which are a useful model to clarify the role of Peyer's patches in intestinal tumorigenesis. Using this model, we increased the number of Peyer's patches using 0.1% and 1% corn husk arabinoxylan through feed. Intestinal polyp formation significantly decreased, concomitant with an increase in Peyer's patches development (n = 12/group). In Aly (-/-) Apc (Min/+) mice (negative control; no Peyer's patches) there was no change in the amount of intestinal polyps (n = 10/group). Immune reaction following corn husk arabinoxylan treatment was measured by cytokine array. Increasing the number of Peyer's patches decreased interleukin-17 production, which showed a dose dependent correlation with transcription factor/lymphoid enhancer-binding factor. This study identified a relationship between levels of Peyer's patches and intestinal polyp formation, partly explained by the involvement of interleukin-17 production and β-catenin signaling in Apc (Min/+) mice. PMID:25678750

  7. Heterozygous deletion of ATG5 in Apc(Min/+) mice promotes intestinal adenoma growth and enhances the antitumor efficacy of interferon-gamma.

    PubMed

    Wang, Lu; Wang, Yan; Lu, Yuyin; Zhang, Qianyun; Qu, Xianjun

    2015-01-01

    Autophagy related gene 5 (ATG5) was lost in 23% of the patients with colorectal cancer (CRC) and the role of loss of ATG5 in the pathogenesis of CRC remains unclear. Knockdown of ATG5 in cancer cells enhances the antitumor efficacy of lots of chemotherapeutic agents. However, there is still no animal model to validate these in vitro observations in vivo. In this study, we found that heterozygous deletion of ATG5 in Apc(Min/+) mice increased the number and size of adenomas as compared with those in Apc(Min/+)ATG5(+/+) mice. To investigate whether ATG5 deficiency could sensitize tumors to chemotherapies, we compared the antitumor effects of Interferon-gamma (IFN-γ) between Apc(Min/+)ATG5(+/+) and Apc(Min/+)ATG5(+/-) mice, as IFN-γ is a potential tumor suppressor for CRC and has been used clinically as an efficient adjuvant to chemotherapy of cancer. We revealed that heterozygous deletion of ATG5 significantly enhanced the antitumor efficacy of IFN-γ. Early treatment of Apc(Min/+)ATG5(+/-) mice with IFN-γ decreased tumor incidence rate to 16.7% and reduced the number of adenomas by 95.5% and late treatment led to regression of tumor. Moreover, IFN-γ treatment did not cause any evident toxic reaction. Mechanistic analysis revealed that heterozygous deletion of ATG5 activated EGFR/ERK1/2 and Wnt/β-catenin pathways in adenomas of Apc(Min/+) mice and enhanced the effects of IFN-γ-dependent inhibition of these 2 pathways. Our results demonstrate that ATG5 plays important roles in intestinal tumor growth and combination of IFN-γ and ATG5 deficiency or ATG5-targeted inhibition is a promising strategy for prevention and treatment of CRC. PMID:25695667

  8. The tissue effect of argon-plasma coagulation with prior submucosal injection (Hybrid-APC) versus standard APC: A randomized ex-vivo study

    PubMed Central

    Neugebauer, Alexander; Scharpf, Marcus; Braun, Kirsten; May, Andrea; Ell, Christian; Fend, Falko; Enderle, Markus D

    2014-01-01

    Background Thermal ablation for Barrett’s oesophagus has widely been established in gastrointestinal endoscopy during the last decade. The mainly used methods of radiofrequency ablation (RFA) and argon-plasma coagulation (APC) carry a relevant risk of stricture formation of up to 5–15%. Newer ablation techniques that are able to overcome this disadvantage would therefore be desirable. The aim of the present study was to compare the depth of tissue injury of the new method of Hybrid-APC versus standard APC within a randomized study in a porcine oesophagus model. Methods Using a total of eight explanted pig oesophagi, 48 oesophageal areas were ablated either by standard or Hybrid-APC (APC with prior submucosal fluid injection) using power settings of 50 and 70 W. The depth of tissue injury to the oesophageal wall was analysed macroscopically and histopathologically. Results Using 50 W, mean coagulation depth was 937 ± 469 µm during standard APC, and 477 ± 271 µm during Hybrid-APC (p = 0.064). Using 70 W, coagulation depth was 1096 ± 320 µm (standard APC) and 468 ± 136 µm (Hybrid-APC; p = 0.003). During all settings, damage to the muscularis mucosae was observed. Using standard APC, damage to the submucosal layer was observed in 4/6 (50 W) and 6/6 cases (70 W). During Hybrid-APC, coagulation of the submucosal layer occurred in 2/6 (50 W) and 1/6 cases (70 W). The proper muscle layer was only damaged during conventional APC (50 W: 1/6; 70 W: 3/6). Limitations Ex-vivo animal study with limited number of cases. Conclusions Hybrid-APC reduces coagulation depth by half in comparison with standard APC, with no thermal injury to the proper muscle layer. It may therefore lead to a lower rate of stricture formation during clinical application. PMID:25360316

  9. The ins and outs of APC and β-catenin nuclear transport

    PubMed Central

    Henderson, Beric R.; Fagotto, Francois

    2002-01-01

    Adenomatous polyposis coli (APC) and β-catenin, two key interacting proteins implicated in development and cancer, were recently found to traffic into and out of the nucleus in response to internal and external signals. The two proteins can enter and exit the nucleus independently, a discovery that has prompted debate about the previously proposed role of APC as a β-catenin chaperone. Here, we review the regulation of APC and β-catenin subcellular localization, in particular in cancer cells. We speculate that, in non-stimulated cells, APC actively exports β-catenin from the nucleus to the cytoplasm where its levels are regulated by degradation; and, conversely, that, in cancer cells or those stimulated by Wnt signaling, β-catenin degradation is inhibited and the accruing protein is capable of moving between the nucleus and cytoplasm independently of APC. Models that link APC and β-catenin transport to function are discussed. PMID:12223464

  10. The ins and outs of APC and beta-catenin nuclear transport.

    PubMed

    Henderson, Beric R; Fagotto, Francois

    2002-09-01

    Adenomatous polyposis coli (APC) and beta-catenin, two key interacting proteins implicated in development and cancer, were recently found to traffic into and out of the nucleus in response to internal and external signals. The two proteins can enter and exit the nucleus independently, a discovery that has prompted debate about the previously proposed role of APC as a beta-catenin chaperone. Here, we review the regulation of APC and beta-catenin subcellular localization, in particular in cancer cells. We speculate that, in non-stimulated cells, APC actively exports beta-catenin from the nucleus to the cytoplasm where its levels are regulated by degradation; and, conversely, that, in cancer cells or those stimulated by Wnt signaling, beta-catenin degradation is inhibited and the accruing protein is capable of moving between the nucleus and cytoplasm independently of APC. Models that link APC and beta-catenin transport to function are discussed. PMID:12223464

  11. Hematopoietic prostaglandin D synthase suppresses intestinal adenomas in ApcMin/+ mice.

    PubMed

    Park, Jae Man; Kanaoka, Yoshihide; Eguchi, Naomi; Aritake, Kosuke; Grujic, Sava; Materi, Alicia M; Buslon, Virgilio S; Tippin, Brigette L; Kwong, Alan M; Salido, Eduardo; French, Samuel W; Urade, Yoshihiro; Lin, Henry J

    2007-02-01

    Aspirin and other nonsteroidal anti-inflammatory drugs prevent some cases of colon cancer by inhibiting prostaglandin (PG) synthesis. PGE(2) promotes colon neoplasia, as shown by knockout mouse studies on enzymes and receptors in the PG cascade. A few experiments 20 to 30 years ago suggested that PGD(2) may suppress tumors, but a role for biosynthetic enzymes for PGD(2) in tumor development has not been studied. We report here that disruption of the gene for hematopoietic PGD synthase in Apc(Min/+) mice led to approximately 50% more intestinal adenomas compared with controls. Tumor size was not affected. By immunohistochemistry, we detected hematopoietic PGD synthase mainly in macrophages and monocytes of the gut mucosa. The mean number of tumors did not increase with knockout of the gene for the lipocalin type of the enzyme, which is not produced in the intestine. On the other hand, Apc(Min/+) mice with transgenic human hematopoietic PGD synthase tended to have 80% fewer intestinal adenomas. The transgene produced high mRNA levels (375-fold over endogenous). There was a suggestion of higher urinary excretion of 11beta-PGF(2alpha) and a lower excretion of a PGE(2) metabolite in transgenic mice, but differences (30-40%) were not statistically significant. The results support an interpretation that hematopoietic PGD synthase controls an inhibitory effect on intestinal tumors. Further studies will be needed to prove possible mechanisms, such as routing of PG production away from protumorigenic PGE(2) or inhibition of the nuclear factor-kappaB cascade by PGD(2) metabolites. PMID:17283118

  12. Analytic Patch Configuration (APC) gateway version 1.0 user's guide

    NASA Technical Reports Server (NTRS)

    Bingel, Bradford D.

    1990-01-01

    The Analytic Patch Configuration (APC) is an interactive software tool which translates aircraft configuration geometry files from one format into another. This initial release of the APC Gateway accommodates six formats: the four accepted APC formats (89f, 89fd, 89u, and 89ud), the PATRAN 2.x phase 1 neutral file format, and the Integrated Aerodynamic Analysis System (IAAS) General Geometry (GG) format. Written in ANSI FORTRAN 77 and completely self-contained, the APC Gateway is very portable and was already installed on CDC/NOS, VAX/VMS, SUN, SGI/IRIS, CONVEX, and GRAY hosts.

  13. Apc bridges Wnt/{beta}-catenin and BMP signaling during osteoblast differentiation of KS483 cells

    SciTech Connect

    Miclea, Razvan L.; Horst, Geertje van der; Robanus-Maandag, Els C.; Loewik, Clemens W.G.M.; Oostdijk, Wilma; Wit, Jan M.; Karperien, Marcel

    2011-06-10

    The canonical Wnt signaling pathway influences the differentiation of mesenchymal cell lineages in a quantitative and qualitative fashion depending on the dose of {beta}-catenin signaling. Adenomatous polyposis coli (Apc) is the critical intracellular regulator of {beta}-catenin turnover. To better understand the molecular mechanisms underlying the role of Apc in regulating the differentiation capacity of skeletal progenitor cells, we have knocked down Apc in the murine mesenchymal stem cell-like KS483 cells by stable expression of Apc-specific small interfering RNA. In routine culture, KSFrt-Apc{sub si} cells displayed a mesenchymal-like spindle shape morphology, exhibited markedly decreased proliferation and increased apoptosis. Apc knockdown resulted in upregulation of the Wnt/{beta}-catenin and the BMP/Smad signaling pathways, but osteogenic differentiation was completely inhibited. This effect could be rescued by adding high concentrations of BMP-7 to the differentiation medium. Furthermore, KSFrt-Apc{sub si} cells showed no potential to differentiate into chondrocytes or adipocytes. These results demonstrate that Apc is essential for the proliferation, survival and differentiation of KS483 cells. Apc knockdown blocks the osteogenic differentiation of skeletal progenitor cells, a process that can be overruled by high BMP signaling.

  14. APC mutation and the crypt cycle in murine and human intestine.

    PubMed Central

    Bjerknes, M.; Cheng, H.; Hay, K.; Gallinger, S.

    1997-01-01

    Dysplastic colon adenomas are thought to arise from growth of clones of APC -/- colonic epithelial cells. Isolated clusters of dysplastic crypts are often observed in patients with familial adenomatous polyposis. These patients have genotype APC +/-, and the clusters of dysplastic crypts (called microadenoma or aberrant crypt foci) are thought to represent an early stage in the expansion of a mutant clone of APC -/- cells. It is thought that the growth of these clusters of mutant crypts results from crypt replication through a process similar to what occurs in the normal crypt cycle. We measured the relative replication rate of mutant crypts by analyzing the size of clusters of mutant crypts in APC +/- individuals and found that mutant APC -/- crypts replicate more rapidly than do normal APC +/- (i.e., nonneoplastic) crypts. In contrast, the replication rate of mutant crypts in Apc +/- mice is not significantly different from that of normal crypts, thus supporting previous findings that aberrant crypt foci do not contribute significantly to the colon adenoma population in adult Apc +/- mice. Intriguingly, we found an effect of Apc heterozygosity on the frequency of branching crypts in young mice. PMID:9060821

  15. APC: A New Code for Atmospheric Polarization Computations

    NASA Technical Reports Server (NTRS)

    Korkin, Sergey V.; Lyapustin, Alexei I.; Rozanov, Vladimir V.

    2014-01-01

    A new polarized radiative transfer code Atmospheric Polarization Computations (APC) is described. The code is based on separation of the diffuse light field into anisotropic and smooth (regular) parts. The anisotropic part is computed analytically. The smooth regular part is computed numerically using the discrete ordinates method. Vertical stratification of the atmosphere, common types of bidirectional surface reflection and scattering by spherical particles or spheroids are included. A particular consideration is given to computation of the bidirectional polarization distribution function (BPDF) of the waved ocean surface.

  16. A trellis-searched APC (adaptive predictive coding) speech coder

    SciTech Connect

    Malone, K.T. ); Fischer, T.R. . Dept. of Electrical and Computer Engineering)

    1990-01-01

    In this paper we formulate a speech coding system that incorporates trellis coded vector quantization (TCVQ) and adaptive predictive coding (APC). A method for optimizing'' the TCVQ codebooks is presented and experimental results concerning survivor path mergings are reported. Simulation results are given for encoding rates of 16 and 9.6 kbps for a variety of coder parameters. The quality of the encoded speech is deemed excellent at an encoding rate of 16 kbps and very good at 9.6 kbps. 13 refs., 2 figs., 4 tabs.

  17. Rh-I-UEA-1 polymerized liposomes target and image adenomatous polyps in the APC(Min/+) mouse using optical colonography.

    PubMed

    Roney, Celeste A; Xu, Biying; Xie, Jianwu; Yuan, Shuai; Wierwille, Jeremiah; Chen, Chao-Wei; Chen, Yu; Griffiths, Gary L; Summers, Ronald M

    2011-08-01

    Mutated adenomatous polyposis coli (APC) genes predispose transformations to neoplasia, progressing to colorectal carcinoma. Early detection facilitates clinical management and therapy. Novel lectin-mediated polymerized targeted liposomes (Rh-I-UEA-1), with polyp specificity and incorporated imaging agents were fabricated to locate and image adenomatous polyps in APC(Min/+) mice. The biomarker α-L-fucose covalently joins the liposomal conjugated lectin Ulexeuropaeus agglutinin (UEA-1), via glycosidic linkage to the polyp mucin layer. Multispectral optical imaging (MSI) corroborated a global perspective of specific binding (rhodamine B 532 nm emission, 590-620 nm excitation) of targeted Rh-I-UEA-1 polymerized liposomes to polyps with 1.4-fold labeling efficiency. High-resolution coregistered optical coherence tomography (OCT) and fluorescence molecular imaging (FMI) reveal the spatial correlation of contrast distribution and tissue morphology. Freshly excised APC(Min) bowels were incubated with targeted liposomes (UEA-1 lectin), control liposomes (no lectin), or iohexol (Omnipaque) and imaged by the three techniques. Computed tomographic quantitative analyses did not confirm that targeted liposomes more strongly bound polyps than nontargeted liposomes or iohexol (Omnipaque) alone. OCT, with anatomic depth capabilities, along with the coregistered FMI, substantiated Rh-I-UEA-1 liposome binding along the mucinous polyp surface. UEA-1 lectin denotes α-l-fucose biomarker carbohydrate expression at the mucin glycoprotein layer; Rh-I-UEA-1 polymerized liposomes target and image adenomatous polyps in APC(Min) mice. PMID:21521550

  18. Sipuleucel-T: APC 8015, APC-8015, prostate cancer vaccine--Dendreon.

    PubMed

    2006-01-01

    Sipuleucel-T [APC 8015, Provenge] is an autologous, dendritic cell-based vaccine under development with Dendreon Corporation for the treatment of androgen-independent and androgen-dependent prostate cancer. It was generated using the company's active immunotherapy platform to stimulate a patient's own immune system to specifically target and destroy cancer cells, while leaving healthy cells unharmed. This approach could provide patients with a meaningful survival benefit and an improved tolerability profile over existing anticancer therapies. Sipuleucel-T selectively targets the prostate-specific antigen (PSA) known as prostatic acid phosphatase (PAP) that is expressed in approximately 95% of prostate cancers. It is produced by ex vivo exposure of dendritic cell precursors to PA 2024, a recombinant fusion protein composed of the PAP target fused to granulocyte-macrophage colony-stimulating factor (GM-CSF) and incorporated into Dendreon's proprietary Antigen Delivery Cassette. Patients are typically administered three intravenous (IV)-infusions of the vaccine over a 1-month period as a complete course of therapy. It is undergoing late-stage clinical evaluation among patients with early and advanced prostate cancer. In November 2003, Kirin Brewery returned to Dendreon the full rights to Sipuleucel-T for Asia. In exchange, Dendreon licensed patent rights relating to the use of certain HLA-DR antibodies to Kirin for $US20 million. This amended agreement enables Dendreon to complete ongoing discussions for a worldwide marketing and sales partnership for Sipuleucel-T. Similarly, Kirin is able to develop its HLA-DR monoclonal antibodies free of potential infringement claims arising from Dendreon's patent rights to HLA-DR. The licensing agreement relates to patent rights owned by Dendreon relating to monoclonal antibodies against the HLA-DR antigen. In addition, Dendreon retains rights to develop and commercialise its two existing HLA-DR monoclonal antibodies, DN 1921 and

  19. Tumorigenic fragments of APC cause dominant defects in directional cell migration in multiple model systems.

    PubMed

    Nelson, Scott A; Li, Zhouyu; Newton, Ian P; Fraser, David; Milne, Rachel E; Martin, David M A; Schiffmann, David; Yang, Xuesong; Dormann, Dirk; Weijer, Cornelis J; Appleton, Paul L; Näthke, Inke S

    2012-11-01

    Nonsense mutations that result in the expression of truncated, N-terminal, fragments of the adenomatous polyposis coli (APC) tumour suppressor protein are found in most sporadic and some hereditary colorectal cancers. These mutations can cause tumorigenesis by eliminating β-catenin-binding sites from APC, which leads to upregulation of β-catenin and thereby results in the induction of oncogenes such as MYC. Here we show that, in three distinct experimental model systems, expression of an N-terminal fragment of APC (N-APC) results in loss of directionality, but not speed, of cell motility independently of changes in β-catenin regulation. We developed a system to culture and fluorescently label live pieces of gut tissue to record high-resolution three-dimensional time-lapse movies of cells in situ. This revealed an unexpected complexity of normal gut cell migration, a key process in gut epithelial maintenance, with cells moving with spatial and temporal discontinuity. Quantitative comparison of gut tissue from wild-type mice and APC heterozygotes (APC(Min/+); multiple intestinal neoplasia model) demonstrated that cells in precancerous epithelia lack directional preference when moving along the crypt-villus axis. This effect was reproduced in diverse experimental systems: in developing chicken embryos, mesoderm cells expressing N-APC failed to migrate normally; in amoeboid Dictyostelium, which lack endogenous APC, expressing an N-APC fragment maintained cell motility, but the cells failed to perform directional chemotaxis; and multicellular Dictyostelium slug aggregates similarly failed to perform phototaxis. We propose that N-terminal fragments of APC represent a gain-of-function mutation that causes cells within tissue to fail to migrate directionally in response to relevant guidance cues. Consistent with this idea, crypts in histologically normal tissues of APC(Min/+) intestines are overpopulated with cells, suggesting that a lack of migration might cause cell

  20. Characterization of an APC Promoter 1B deletion in a Patient Diagnosed with Familial Adenomatous Polyposis via Whole Genome Shotgun Sequencing

    PubMed Central

    Kalbfleisch, Ted; Brock, Pamela; Snow, Angela; Neklason, Deborah; Gowans, Gordon; Klein, Jon

    2015-01-01

    Recently, deletions have been identified and published as causal for Familial Adenomatous Polyposis in the 1B promoter region of the APC gene.  Those deletions were measured using multiplex ligation-dependent probe amplification.  Here, we present and characterize an ~11kb deletion identified by whole genome shotgun sequencing.  The deletion occurred in a patient diagnosed with Familial Adenomatous Polyposis, and was located on chr5, between bases 112,034,824 and 112,045,845, fully encompassing the 1B promoter region of the APC gene.   Results are presented here that include the sequence evidence supporting the presence of the deletion as well as base level characterization of the deletion site.  These results demonstrate the capacity of whole genome sequencing for the detection of large structural variants in single individuals. PMID:26213617

  1. Characterization of an APC Promoter 1B deletion in a Patient Diagnosed with Familial Adenomatous Polyposis via Whole Genome Shotgun Sequencing.

    PubMed

    Kalbfleisch, Ted; Brock, Pamela; Snow, Angela; Neklason, Deborah; Gowans, Gordon; Klein, Jon

    2015-01-01

    Recently, deletions have been identified and published as causal for Familial Adenomatous Polyposis in the 1B promoter region of the APC gene.  Those deletions were measured using multiplex ligation-dependent probe amplification.  Here, we present and characterize an ~11kb deletion identified by whole genome shotgun sequencing.  The deletion occurred in a patient diagnosed with Familial Adenomatous Polyposis, and was located on chr5, between bases 112,034,824 and 112,045,845, fully encompassing the 1B promoter region of the APC gene.   Results are presented here that include the sequence evidence supporting the presence of the deletion as well as base level characterization of the deletion site.  These results demonstrate the capacity of whole genome sequencing for the detection of large structural variants in single individuals. PMID:26213617

  2. 42 CFR 419.31 - Ambulatory payment classification (APC) system and payment weights.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 42 Public Health 3 2011-10-01 2011-10-01 false Ambulatory payment classification (APC) system and payment weights. 419.31 Section 419.31 Public Health CENTERS FOR MEDICARE & MEDICAID SERVICES, DEPARTMENT... Outpatient Services § 419.31 Ambulatory payment classification (APC) system and payment weights. (a)...

  3. REMOVAL OF ARSENIC IN DRINKING WATER: ARS CFU-50 APC ELECTROFLOCCULATION AND FILTRATION WATER TREATMENT SYSTEM

    EPA Science Inventory

    ETV testing of the ARS CFU-50 APC Electroflocculation and Filtration Water Treatment System (ARS CFU-50 APC) for arsenic removal was conducted at the Town of Bernalillo Well #3 site from April 18 through May 2, 2006. The source water was chlorinated groundwater from two supply w...

  4. Modeling the Effect of APC Truncation on Destruction Complex Function in Colorectal Cancer Cells

    SciTech Connect

    Barua, Dipak; Hlavacek, William S.

    2013-09-26

    In colorectal cancer cells, APC, a tumor suppressor protein, is commonly expressed in truncated form. Truncation of APC is believed to disrupt degradation of β—catenin, which is regulated by a multiprotein complex called the destruction complex. The destruction complex comprises APC, Axin, β—catenin, serine/threonine kinases, and other proteins. The kinases CK1α and GSK–3β, which are recruited by Axin, mediate phosphorylation of β—catenin, which initiates its ubiquitination and proteosomal degradation. The mechanism of regulation of β—catenin degradation by the destruction complex and the role of truncation of APC in colorectal cancer are not entirely understood. Through formulation and analysis of a rule-based computational model, we investigated the regulation of β—catenin phosphorylation and degradation by APC and the effect of APC truncation on function of the destruction complex. The model integrates available mechanistic knowledge about site-specific interactions and phosphorylation of destruction complex components and is consistent with an array of published data. In this paper, we find that the phosphorylated truncated form of APC can outcompete Axin for binding to β—catenin, provided that Axin is limiting, and thereby sequester β—catenin away from Axin and the Axin-recruited kinases CK1α and GSK–3β. Full-length APC also competes with Axin for binding to β—catenin; however, full-length APC is able, through its SAMP repeats, which bind Axin and which are missing in truncated oncogenic forms of APC, to bring β—catenin into indirect association with Axin and Axin-recruited kinases. Because our model indicates that the positive effects of truncated APC on β—catenin levels depend on phosphorylation of APC, at the first 20-amino acid repeat, and because phosphorylation of this site is mediated by CK1ϵ, we suggest that CK1ϵ is a potential target for therapeutic intervention in colorectal cancer. Finally, specific inhibition

  5. Modeling the Effect of APC Truncation on Destruction Complex Function in Colorectal Cancer Cells

    DOE PAGESBeta

    Barua, Dipak; Hlavacek, William S.

    2013-09-26

    In colorectal cancer cells, APC, a tumor suppressor protein, is commonly expressed in truncated form. Truncation of APC is believed to disrupt degradation of β—catenin, which is regulated by a multiprotein complex called the destruction complex. The destruction complex comprises APC, Axin, β—catenin, serine/threonine kinases, and other proteins. The kinases CK1α and GSK–3β, which are recruited by Axin, mediate phosphorylation of β—catenin, which initiates its ubiquitination and proteosomal degradation. The mechanism of regulation of β—catenin degradation by the destruction complex and the role of truncation of APC in colorectal cancer are not entirely understood. Through formulation and analysis of amore » rule-based computational model, we investigated the regulation of β—catenin phosphorylation and degradation by APC and the effect of APC truncation on function of the destruction complex. The model integrates available mechanistic knowledge about site-specific interactions and phosphorylation of destruction complex components and is consistent with an array of published data. In this paper, we find that the phosphorylated truncated form of APC can outcompete Axin for binding to β—catenin, provided that Axin is limiting, and thereby sequester β—catenin away from Axin and the Axin-recruited kinases CK1α and GSK–3β. Full-length APC also competes with Axin for binding to β—catenin; however, full-length APC is able, through its SAMP repeats, which bind Axin and which are missing in truncated oncogenic forms of APC, to bring β—catenin into indirect association with Axin and Axin-recruited kinases. Because our model indicates that the positive effects of truncated APC on β—catenin levels depend on phosphorylation of APC, at the first 20-amino acid repeat, and because phosphorylation of this site is mediated by CK1ϵ, we suggest that CK1ϵ is a potential target for therapeutic intervention in colorectal cancer. Finally, specific

  6. Modeling the effect of APC truncation on destruction complex function in colorectal cancer cells.

    PubMed

    Barua, Dipak; Hlavacek, William S

    2013-01-01

    In colorectal cancer cells, APC, a tumor suppressor protein, is commonly expressed in truncated form. Truncation of APC is believed to disrupt degradation of β-catenin, which is regulated by a multiprotein complex called the destruction complex. The destruction complex comprises APC, Axin, β-catenin, serine/threonine kinases, and other proteins. The kinases CK1α and GSK -3β, which are recruited by Axin, mediate phosphorylation of β-catenin, which initiates its ubiquitination and proteosomal degradation. The mechanism of regulation of β-catenin degradation by the destruction complex and the role of truncation of APC in colorectal cancer are not entirely understood. Through formulation and analysis of a rule-based computational model, we investigated the regulation of β-catenin phosphorylation and degradation by APC and the effect of APC truncation on function of the destruction complex. The model integrates available mechanistic knowledge about site-specific interactions and phosphorylation of destruction complex components and is consistent with an array of published data. We find that the phosphorylated truncated form of APC can outcompete Axin for binding to β-catenin, provided that Axin is limiting, and thereby sequester β-catenin away from Axin and the Axin-recruited kinases CK1α and GSK -3β. Full-length APC also competes with Axin for binding to β-catenin; however, full-length APC is able, through its SAMP repeats, which bind Axin and which are missing in truncated oncogenic forms of APC, to bring β-catenin into indirect association with Axin and Axin-recruited kinases. Because our model indicates that the positive effects of truncated APC on β-catenin levels depend on phosphorylation of APC, at the first 20-amino acid repeat, and because phosphorylation of this site is mediated by CK1ε, we suggest that CK1ε is a potential target for therapeutic intervention in colorectal cancer. Specific inhibition of CK1ε is predicted to limit binding of

  7. Critical Current Properties in Longitudinal Magnetic Field of YBCO Superconductor with APC

    NASA Astrophysics Data System (ADS)

    Kido, R.; Kiuchi, M.; Otabe, E. S.; Matsushita, T.; Jha, A. K.; Matsumoto, K.

    The critical current density (Jc) properties of the Artificial Pinning Center (APC) introduced YBa2Cu3O7 (YBCO) films in the longitudinal magnetic field were measured. Y2O3 or Y2BaCuO5 (Y211) was introduced as APCs to YBCO, and YBCO films with APC were fabricated on SrTiO3 single crystal substrate. The sizes of Y2O3 and Y211 were 5-10 nm and 10-20 nm, respectively. As a result, Jc enhancement in the longitudinal magnetic field was observed in Y2O3 introduced YBCO films. However, it was not observed in Y211 introduced YBCO films. Therefore, it was considered that Jc properties in the longitudinal magnetic field were affected by introducing of small size APC, and it was necessary that APC does not disturb the current pathway in the superconductor.

  8. Berberine potently attenuates intestinal polyps growth in ApcMin mice and familial adenomatous polyposis patients through inhibition of Wnt signalling

    PubMed Central

    Zhang, Junfang; Cao, Hailong; Zhang, Bing; Cao, Hanwei; Xu, Xiuqin; Ruan, Hang; Yi, Tingting; Tan, Li; Qu, Rui; Song, Gang; Wang, Bangmao; Hu, Tianhui

    2013-01-01

    As a traditional anti-inflammatory Chinese herbal medicine, Alkaloid berberine has been recently reported to exhibit anti-tumour effects against a wide spectrum of cancer. However, the mechanism was largely unknown. Gene chip array reveals that with berberine treatment, c-Myc, the target gene of Wnt pathway, was down-regulated 5.3-folds, indicating that berberine might inhibit Wnt signalling. TOPflash analysis revealed that Wnt activity was significantly reduced after berberine treatment, and the mechanism of which might be that berberine disrupted β-catenin transfer to nucleus through up-regulating the expression of adenomatous polyposis coli (APC) gene and stabilized APC-β-catenin complex. Berberine administration in ApcMin/+ mice exhibited fewer and smaller polyps in intestine, along with reduction in cyclin D1 and c-Myc expression. In clinical practice, oral administration of berberine also significantly reduced the familial adenomatous polyposis patients' polyp size along with the inhibition of cyclin D1 expression in polyp samples. These observations indicate that berberine inhibits colon tumour formation through inhibition of Wnt/β-catenin signalling and berberine might be a promising drug for the prevention of colon cancer. PMID:24015932

  9. APC2 and Axin promote mitotic fidelity by facilitating centrosome separation and cytoskeletal regulation.

    PubMed

    Poulton, John S; Mu, Frank W; Roberts, David M; Peifer, Mark

    2013-10-01

    To ensure the accurate transmission of genetic material, chromosome segregation must occur with extremely high fidelity. Segregation errors lead to chromosomal instability (CIN), with deleterious consequences. Mutations in the tumor suppressor adenomatous polyposis coli (APC) initiate most colon cancers and have also been suggested to promote disease progression through increased CIN, but the mechanistic role of APC in preventing CIN remains controversial. Using fly embryos as a model, we investigated the role of APC proteins in CIN. Our findings suggest that APC2 loss leads to increased rates of chromosome segregation error. This occurs through a cascade of events beginning with incomplete centrosome separation leading to failure to inhibit formation of ectopic cleavage furrows, which result in mitotic defects and DNA damage. We test several hypotheses related to the mechanism of action of APC2, revealing that APC2 functions at the embryonic cortex with several protein partners, including Axin, to promote mitotic fidelity. Our in vivo data demonstrate that APC2 protects genome stability by modulating mitotic fidelity through regulation of the cytoskeleton. PMID:24026117

  10. Oscillation of APC/C activity during cell cycle arrest promotes centrosome amplification

    PubMed Central

    Prosser, Suzanna L.; Samant, Mugdha D.; Baxter, Joanne E.; Morrison, Ciaran G.; Fry, Andrew M.

    2014-01-01

    Centrosome duplication is licensed by the disengagement, or ‘uncoupling’, of centrioles during late mitosis. However, arrest of cells in G2 can trigger premature centriole disengagement. Here, we show that premature disengagement results from untimely activation of the APC/C leading to securin degradation and release of active separase. APC/C activation during G2 arrest is dependent on Plk1-mediated degradation of the APC/C inhibitor, Emi1, but Plk1 also has a second APC/C-independent role in promoting disengagement. Importantly, APC/C and Plk1 activity also stimulate centriole disengagement in response to hydroxyurea or DNA damage-induced cell cycle arrest and this leads to centrosome amplification. However, the re-duplication of disengaged centrioles is dependent on Cdk2 activity and Cdk2 activation coincides with a subsequent inactivation of the APC/C and re-accumulation of cyclin A. Release from these arrests leads to mitotic entry but, due to the presence of disengaged and/or amplified centrosomes, formation of abnormal mitotic spindles that lead to chromosome missegregation. Thus, oscillation of APC/C activity during cell cycle arrest promotes both centrosome amplification and genome instability. PMID:22956538

  11. Copy number variants associated with 18p11.32, DCC and the promoter 1B region of APC in colorectal polyposis patients

    PubMed Central

    Masson, Amy L.; Talseth-Palmer, Bente A.; Evans, Tiffany-Jane; McElduff, Patrick; Spigelman, Allan D.; Hannan, Garry N.; Scott, Rodney J.

    2015-01-01

    Familial Adenomatous Polyposis (FAP) is the second most common inherited predisposition to colorectal cancer (CRC) associated with the development of hundreds to thousands of adenomas in the colon and rectum. Mutations in APC are found in ~ 80% polyposis patients with FAP. In the remaining 20% no genetic diagnosis can be provided suggesting other genes or mechanisms that render APC inactive may be responsible. Copy number variants (CNVs) remain to be investigated in FAP and may account for disease in a proportion of polyposis patients. A cohort of 56 polyposis patients and 40 controls were screened for CNVs using the 2.7M microarray (Affymetrix) with data analysed using ChAS (Affymetrix). A total of 142 CNVs were identified unique to the polyposis cohort suggesting their involvement in CRC risk. We specifically identified CNVs in four unrelated polyposis patients among CRC susceptibility genes APC, DCC, MLH1 and CTNNB1 which are likely to have contributed to disease development in these patients. A recurrent deletion was observed at position 18p11.32 in 9% of the patients screened that was of particular interest. Further investigation is necessary to fully understand the role of these variants in CRC risk given the high prevalence among the patients screened. PMID:26909336

  12. Copy number variants associated with 18p11.32, DCC and the promoter 1B region of APC in colorectal polyposis patients.

    PubMed

    Masson, Amy L; Talseth-Palmer, Bente A; Evans, Tiffany-Jane; McElduff, Patrick; Spigelman, Allan D; Hannan, Garry N; Scott, Rodney J

    2016-02-01

    Familial Adenomatous Polyposis (FAP) is the second most common inherited predisposition to colorectal cancer (CRC) associated with the development of hundreds to thousands of adenomas in the colon and rectum. Mutations in APC are found in ~ 80% polyposis patients with FAP. In the remaining 20% no genetic diagnosis can be provided suggesting other genes or mechanisms that render APC inactive may be responsible. Copy number variants (CNVs) remain to be investigated in FAP and may account for disease in a proportion of polyposis patients. A cohort of 56 polyposis patients and 40 controls were screened for CNVs using the 2.7M microarray (Affymetrix) with data analysed using ChAS (Affymetrix). A total of 142 CNVs were identified unique to the polyposis cohort suggesting their involvement in CRC risk. We specifically identified CNVs in four unrelated polyposis patients among CRC susceptibility genes APC, DCC, MLH1 and CTNNB1 which are likely to have contributed to disease development in these patients. A recurrent deletion was observed at position 18p11.32 in 9% of the patients screened that was of particular interest. Further investigation is necessary to fully understand the role of these variants in CRC risk given the high prevalence among the patients screened. PMID:26909336

  13. Tumorigenic fragments of APC cause dominant defects in directional cell migration in multiple model systems

    PubMed Central

    Nelson, Scott A.; Li, Zhouyu; Newton, Ian P.; Fraser, David; Milne, Rachel E.; Martin, David M. A.; Schiffmann, David; Yang, Xuesong; Dormann, Dirk; Weijer, Cornelis J.; Appleton, Paul L.; Näthke, Inke S.

    2012-01-01

    SUMMARY Nonsense mutations that result in the expression of truncated, N-terminal, fragments of the adenomatous polyposis coli (APC) tumour suppressor protein are found in most sporadic and some hereditary colorectal cancers. These mutations can cause tumorigenesis by eliminating β-catenin-binding sites from APC, which leads to upregulation of β-catenin and thereby results in the induction of oncogenes such as MYC. Here we show that, in three distinct experimental model systems, expression of an N-terminal fragment of APC (N-APC) results in loss of directionality, but not speed, of cell motility independently of changes in β-catenin regulation. We developed a system to culture and fluorescently label live pieces of gut tissue to record high-resolution three-dimensional time-lapse movies of cells in situ. This revealed an unexpected complexity of normal gut cell migration, a key process in gut epithelial maintenance, with cells moving with spatial and temporal discontinuity. Quantitative comparison of gut tissue from wild-type mice and APC heterozygotes (APCMin/+; multiple intestinal neoplasia model) demonstrated that cells in precancerous epithelia lack directional preference when moving along the crypt-villus axis. This effect was reproduced in diverse experimental systems: in developing chicken embryos, mesoderm cells expressing N-APC failed to migrate normally; in amoeboid Dictyostelium, which lack endogenous APC, expressing an N-APC fragment maintained cell motility, but the cells failed to perform directional chemotaxis; and multicellular Dictyostelium slug aggregates similarly failed to perform phototaxis. We propose that N-terminal fragments of APC represent a gain-of-function mutation that causes cells within tissue to fail to migrate directionally in response to relevant guidance cues. Consistent with this idea, crypts in histologically normal tissues of APCMin/+ intestines are overpopulated with cells, suggesting that a lack of migration might cause

  14. Adenomatous polyposis coli (APC) regulates multiple signaling pathways by enhancing glycogen synthase kinase-3 (GSK-3) activity.

    PubMed

    Valvezan, Alexander J; Zhang, Fang; Diehl, J Alan; Klein, Peter S

    2012-02-01

    Glycogen synthase kinase-3 (GSK-3) is essential for many signaling pathways and cellular processes. As Adenomatous Polyposis Coli (APC) functions in many of the same processes, we investigated a role for APC in the regulation of GSK-3-dependent signaling. We find that APC directly enhances GSK-3 activity. Furthermore, knockdown of APC mimics inhibition of GSK-3 by reducing phosphorylation of glycogen synthase and by activating mTOR, revealing novel roles for APC in the regulation of these enzymes. Wnt signaling inhibits GSK-3 through an unknown mechanism, and this results in both stabilization of β-catenin and activation of mTOR. We therefore hypothesized that Wnts may regulate GSK-3 by disrupting the interaction between APC and the Axin-GSK-3 complex. We find that Wnts rapidly induce APC dissociation from Axin, correlating with β-catenin stabilization. Furthermore, Axin interaction with the Wnt co-receptor LRP6 causes APC dissociation from Axin. We propose that APC regulates multiple signaling pathways by enhancing GSK-3 activity, and that Wnts induce APC dissociation from Axin to reduce GSK-3 activity and activate downstream signaling. APC regulation of GSK-3 also provides a novel mechanism for Wnt regulation of multiple downstream effectors, including β-catenin and mTOR. PMID:22184111

  15. β-Catenin destruction complex-independent regulation of Hippo–YAP signaling by APC in intestinal tumorigenesis

    PubMed Central

    Cai, Jing; Maitra, Anirban; Anders, Robert A.; Taketo, Makoto M.; Pan, Duojia

    2015-01-01

    Mutations in Adenomatous polyposis coli (APC) underlie familial adenomatous polyposis (FAP), an inherited cancer syndrome characterized by the widespread development of colorectal polyps. APC is best known as a scaffold protein in the β-catenin destruction complex, whose activity is antagonized by canonical Wnt signaling. Whether other effector pathways mediate APC's tumor suppressor function is less clear. Here we report that activation of YAP, the downstream effector of the Hippo signaling pathway, is a general hallmark of tubular adenomas from FAP patients. We show that APC functions as a scaffold protein that facilitates the Hippo kinase cascade by interacting with Sav1 and Lats1. Consistent with the molecular link between APC and the Hippo signaling pathway, genetic analysis reveals that YAP is absolutely required for the development of APC-deficient adenomas. These findings establish Hippo–YAP signaling as a critical effector pathway downstream from APC, independent from its involvement in the β-catenin destruction complex. PMID:26193883

  16. Impact of Atomic Gap Size on Sensitivity and Backaction of APC Displacement Detectors

    NASA Astrophysics Data System (ADS)

    Flowers-Jacobs, N. E.; Lehnert, K. W.

    2008-03-01

    Recently our group created a mesoscopic displacement detector formed by coupling an atomic point contact (APC) to a nanomechanical beam and demonstrated a displacement imprecision limited by the fundamental shot-noise in the number of electrons that tunnel across the APC [1]. We continue this work by using a cryogenic apparatus that flexes the device substrate to mechanically adjust the size of the APC atomic gap in situ. The resulting changes in the APC displacement detector's intrinsic noise properties are measured by observing the 1 K random thermal motion of the nanomechanical beam at resonance frequencies up to 200 MHz. The goal of this work is to explore the effect of atomic gap size and shape on displacement sensitivity, understand the origin of the observed measurement backaction, and measure the recoil force of tunneling electrons. [1] N. E. Flowers-Jacobs, D. R. Schmidt, and K. W. Lehnert, Phys. Rev. Lett. 98, 096804 (2007)

  17. Quercetin Supplementation Attenuates the Progression of Cancer Cachexia in ApcMin/+ Mice123

    PubMed Central

    Velázquez, Kandy T.; Enos, Reilly T.; Narsale, Aditi A.; Puppa, Melissa J.; Davis, J. Mark; Murphy, E. Angela; Carson, James A.

    2014-01-01

    Although there are currently no approved treatments for cancer cachexia, there is an intensified interest in developing therapies because of the high mortality index associated with muscle wasting diseases. Successful treatment of the cachectic patient focuses on improving or maintaining body weight and musculoskeletal function. Nutraceutical compounds, including the natural phytochemical quercetin, are being examined as potential treatments because of their anti-inflammatory, antioxidant, and anticarcinogenic properties. The purpose of this study was to determine the effect of quercetin supplementation on the progression of cachexia in the adenomatous polyposis coli (Apc)Min/+ mouse model of colorectal cancer. At 15 wk of age, C57BL/6 and male ApcMin/+ mice were supplemented with 25 mg/kg of quercetin or vehicle solution mix of Tang juice and water (V) daily for 3 wk. Body weight, strength, neuromuscular performance, and fatigue were assessed before and after quercetin or V interventions. Indicators of metabolic dysfunction and inflammatory signaling were also assessed. During the treatment period, the relative decrease in body weight in the ApcMin/+ mice gavaged with V (ApcMin/+V; −14% ± 2.3) was higher than in control mice gavaged with V (+0.6% ± 1.0), control mice gavaged with quercetin (−2% ± 1.0), and ApcMin/+ mice gavaged with quercetin (ApcMin/+Q; −9% ± 1.3). At 18 wk of age, the loss of grip strength and muscle mass shown in ApcMin/+V mice was significantly attenuated (P < 0.05) in ApcMin/+Q mice. Furthermore, ApcMin/+V mice had an induction of plasma interleukin-6 and muscle signal transducer and activator of transcription 3 phosphorylation, which were significantly (P < 0.05) mitigated in ApcMin/+Q mice, despite having a similar tumor burden. Quercetin treatment did not improve treadmill run-time-to-fatigue, hyperglycemia, or hyperlipidemia in cachectic ApcMin/+ mice. Overall, quercetin supplementation positively affected several aspects of

  18. Evaluation and Analysis of Seasat a Scanning Multichannel Microwave Radiometer (SMMR) Antenna Pattern Correction (APC) Algorithm

    NASA Technical Reports Server (NTRS)

    Kitzis, S. N.; Kitzis, J. L.

    1979-01-01

    The accuracy of the SEASAT-A SMMR antenna pattern correction (APC) algorithm was assessed. Interim APC brightness temperature measurements for the SMMR 6.6 GHz channels are compared with surface truth derived sea surface temperatures. Plots and associated statistics are presented for SEASAT-A SMMR data acquired for the Gulf of Alaska experiment. The cross-track gradients observed in the 6.6 GHz brightness temperature data are discussed.

  19. Synergistic Blockade of Mitotic Exit by Two Chemical Inhibitors of the APC/C

    PubMed Central

    Sackton, Katharine L.; Dimova, Nevena; Zeng, Xing; Tian, Wei; Zhang, Mengmeng; Sackton, Timothy B.; Meaders, Johnathan; Pfaff, Kathleen L.; Sigoillot, Frederic; Yu, Hongtao; Luo, Xuelian; King, Randall W.

    2014-01-01

    Summary Protein machines are multi-subunit protein complexes that orchestrate highly regulated biochemical tasks. An example is the Anaphase-Promoting Complex/Cyclosome (APC/C), a thirteen-subunit ubiquitin ligase that initiates the metaphase-anaphase transition and mitotic exit by targeting proteins such as securin and cyclin B1 for ubiquitin-dependent destruction by the proteasome1,2. Because blocking mitotic exit is an effective approach for inducing tumor cell death3,4, the APC/C represents a potential novel target for cancer therapy. APC/C activation in mitosis requires binding of Cdc205, which forms a co-receptor with the APC/C to recognize substrates containing a Destruction box (D-box)6-14. Here we demonstrate that we can synergistically inhibit APC/C-dependent proteolysis and mitotic exit by simultaneously disrupting two protein-protein interactions within the APC/C-Cdc20-substrate ternary complex. We identified a small molecule, called apcin (APC inhibitor), which binds to Cdc20 and competitively inhibits the ubiquitylation of D-box-containing substrates. Analysis of the crystal structure of the apcin-Cdc20 complex suggests that apcin occupies the D-box-binding pocket on the side face of the WD40-domain. The ability of apcin to block mitotic exit is synergistically amplified by co-addition of tosyl-L-arginine methyl ester (TAME), a small molecule that blocks the APC/C-Cdc20 interaction15,16. This work suggests that simultaneous disruption of multiple, weak protein-protein interactions is an effective approach for inactivating a protein machine. PMID:25156254

  20. Selection for Phase Variation of LOS Biosynthetic Genes Frequently Occurs in Progression of Non-Typeable Haemophilus influenzae Infection from the Nasopharynx to the Middle Ear of Human Patients

    PubMed Central

    Srikhanta, Yogitha N.; Eckert, Anja; Novotny, Laura A.; Bakaletz, Lauren O.; Jennings, Michael P.

    2014-01-01

    Surface structures in Haemophilus influenzae are subject to rapid ON/OFF switching of expression, a process termed phase variation. We analyse tetranucleotide repeats controlling phase variation in lipo-oligosaccharide (LOS) genes of H. influenzae in paired isolates from both the nasopharynx and middle ears of paediatric patients with chronic or recurrent otitis media. A change in expression of at least one of the seven phase variable LOS biosynthesis genes was seen in 12 of the 21 strain pairs. Several strains showed switching of expression in multiple LOS genes, consistent with a key role for phase variable LOS biosynthetic genes in human infection. PMID:24587383

  1. Atomic structure of the APC/C and its mechanism of protein ubiquitination

    PubMed Central

    Yang, Jing; McLaughlin, Stephen H.; Barford, David

    2015-01-01

    The anaphase-promoting complex (APC/C) is a multimeric RING E3 ubiquitin ligase that controls chromosome segregation and mitotic exit. Its regulation by coactivator subunits, phosphorylation, the mitotic checkpoint complex, and interphase inhibitor Emi1 ensures the correct order and timing of distinct cell cycle transitions. Here, we used cryo-electron microscopy to determine atomic structures of APC/C-coactivator complexes with either Emi1 or a UbcH10-ubiquitin conjugate. These structures define the architecture of all APC/C subunits, the position of the catalytic module, and explain how Emi1 mediates inhibition of the two E2s UbcH10 and Ube2S. Definition of Cdh1 interactions with the APC/C indicates how they are antagonized by Cdh1 phosphorylation. The structure of the APC/C with UbcH10-ubiquitin reveals insights into the initiating ubiquitination reaction. Our results provide a quantitative framework for the design of experiments to further investigate APC/C functions in vivo. PMID:26083744

  2. The APC/C Ubiquitin Ligase: From Cell Biology to Tumorigenesis

    PubMed Central

    Penas, Clara; Ramachandran, Vimal; Ayad, Nagi George

    2011-01-01

    The ubiquitin proteasome system (UPS) is required for normal cell proliferation, vertebrate development, and cancer cell transformation. The UPS consists of multiple proteins that work in concert to target a protein for degradation via the 26S proteasome. Chains of an 8.5-kDa protein called ubiquitin are attached to substrates, thus allowing recognition by the 26S proteasome. Enzymes called ubiquitin ligases or E3s mediate specific attachment to substrates. Although there are over 600 different ubiquitin ligases, the Skp1–Cullin–F-box (SCF) complexes and the anaphase promoting complex/cyclosome (APC/C) are the most studied. SCF involvement in cancer has been known for some time while APC/C’s cancer role has recently emerged. In this review we will discuss the importance of APC/C to normal cell proliferation and development, underscoring its possible contribution to transformation. We will also examine the hypothesis that modulating a specific interaction of the APC/C may be therapeutically attractive in specific cancer subtypes. Finally, given that the APC/C pathway is relatively new as a cancer target, therapeutic interventions affecting APC/C activity may be beneficial in cancers that are resistant to classical chemotherapy. PMID:22655255

  3. Atomic structure of the APC/C and its mechanism of protein ubiquitination.

    PubMed

    Chang, Leifu; Zhang, Ziguo; Yang, Jing; McLaughlin, Stephen H; Barford, David

    2015-06-25

    The anaphase-promoting complex (APC/C) is a multimeric RING E3 ubiquitin ligase that controls chromosome segregation and mitotic exit. Its regulation by coactivator subunits, phosphorylation, the mitotic checkpoint complex and interphase early mitotic inhibitor 1 (Emi1) ensures the correct order and timing of distinct cell-cycle transitions. Here we use cryo-electron microscopy to determine atomic structures of APC/C-coactivator complexes with either Emi1 or a UbcH10-ubiquitin conjugate. These structures define the architecture of all APC/C subunits, the position of the catalytic module and explain how Emi1 mediates inhibition of the two E2s UbcH10 and Ube2S. Definition of Cdh1 interactions with the APC/C indicates how they are antagonized by Cdh1 phosphorylation. The structure of the APC/C with UbcH10-ubiquitin reveals insights into the initiating ubiquitination reaction. Our results provide a quantitative framework for the design of future experiments to investigate APC/C functions in vivo. PMID:26083744

  4. Fizzy is required for activation of the APC/cyclosome in Xenopus egg extracts.

    PubMed

    Lorca, T; Castro, A; Martinez, A M; Vigneron, S; Morin, N; Sigrist, S; Lehner, C; Dorée, M; Labbé, J C

    1998-07-01

    The Xenopus homologue of Drosophila Fizzy and budding yeast CDC20 has been characterized. The encoded protein (X-FZY) is a component of a high molecular weight complex distinct from the APC/cyclosome. Antibodies directed against FZY were produced and shown to prevent calmodulin-dependent protein kinase II (CaMKII) from inducing the metaphase to anaphase transition of spindles assembled in vitro in Xenopus egg extracts, and this was associated with suppression of the degradation of mitotic cyclins. The same antibodies suppressed M phase-promoting factor (MPF)-dependent activation of the APC/cyclosome in interphase egg extracts, although they did not appear to alter the pattern or extent of MPF-dependent phosphorylation of APC/cyclosome subunits. As these phosphorylations are thought to be essential for APC/cyclosome activation in eggs and early embryos, we conclude that at least two events are required for MPF to activate the APC/cyclosome, allowing both chromatid segregation and full degradation of mitotic cyclins. The first one, which does not require FZY function, is the phosphorylation of APC/cyclosome subunits. The second one, that requires FZY function (even in the absence of MAD2 protein and when the spindle assembly checkpoint is not activated) is not yet understood at its molecular level. PMID:9649427

  5. Point Mutations in Exon 1B of APC Reveal Gastric Adenocarcinoma and Proximal Polyposis of the Stomach as a Familial Adenomatous Polyposis Variant.

    PubMed

    Li, Jun; Woods, Susan L; Healey, Sue; Beesley, Jonathan; Chen, Xiaoqing; Lee, Jason S; Sivakumaran, Haran; Wayte, Nicci; Nones, Katia; Waterfall, Joshua J; Pearson, John; Patch, Anne-Marie; Senz, Janine; Ferreira, Manuel A; Kaurah, Pardeep; Mackenzie, Robertson; Heravi-Moussavi, Alireza; Hansford, Samantha; Lannagan, Tamsin R M; Spurdle, Amanda B; Simpson, Peter T; da Silva, Leonard; Lakhani, Sunil R; Clouston, Andrew D; Bettington, Mark; Grimpen, Florian; Busuttil, Rita A; Di Costanzo, Natasha; Boussioutas, Alex; Jeanjean, Marie; Chong, George; Fabre, Aurélie; Olschwang, Sylviane; Faulkner, Geoffrey J; Bellos, Evangelos; Coin, Lachlan; Rioux, Kevin; Bathe, Oliver F; Wen, Xiaogang; Martin, Hilary C; Neklason, Deborah W; Davis, Sean R; Walker, Robert L; Calzone, Kathleen A; Avital, Itzhak; Heller, Theo; Koh, Christopher; Pineda, Marbin; Rudloff, Udo; Quezado, Martha; Pichurin, Pavel N; Hulick, Peter J; Weissman, Scott M; Newlin, Anna; Rubinstein, Wendy S; Sampson, Jone E; Hamman, Kelly; Goldgar, David; Poplawski, Nicola; Phillips, Kerry; Schofield, Lyn; Armstrong, Jacqueline; Kiraly-Borri, Cathy; Suthers, Graeme K; Huntsman, David G; Foulkes, William D; Carneiro, Fatima; Lindor, Noralane M; Edwards, Stacey L; French, Juliet D; Waddell, Nicola; Meltzer, Paul S; Worthley, Daniel L; Schrader, Kasmintan A; Chenevix-Trench, Georgia

    2016-05-01

    Gastric adenocarcinoma and proximal polyposis of the stomach (GAPPS) is an autosomal-dominant cancer-predisposition syndrome with a significant risk of gastric, but not colorectal, adenocarcinoma. We mapped the gene to 5q22 and found loss of the wild-type allele on 5q in fundic gland polyps from affected individuals. Whole-exome and -genome sequencing failed to find causal mutations but, through Sanger sequencing, we identified point mutations in APC promoter 1B that co-segregated with disease in all six families. The mutations reduced binding of the YY1 transcription factor and impaired activity of the APC promoter 1B in luciferase assays. Analysis of blood and saliva from carriers showed allelic imbalance of APC, suggesting that these mutations lead to decreased allele-specific expression in vivo. Similar mutations in APC promoter 1B occur in rare families with familial adenomatous polyposis (FAP). Promoter 1A is methylated in GAPPS and sporadic FGPs and in normal stomach, which suggests that 1B transcripts are more important than 1A in gastric mucosa. This might explain why all known GAPPS-affected families carry promoter 1B point mutations but only rare FAP-affected families carry similar mutations, the colonic cells usually being protected by the expression of the 1A isoform. Gastric polyposis and cancer have been previously described in some FAP-affected individuals with large deletions around promoter 1B. Our finding that GAPPS is caused by point mutations in the same promoter suggests that families with mutations affecting the promoter 1B are at risk of gastric adenocarcinoma, regardless of whether or not colorectal polyps are present. PMID:27087319

  6. APC binds the Miro/Milton motor complex to stimulate transport of mitochondria to the plasma membrane.

    PubMed

    Mills, Kate M; Brocardo, Mariana G; Henderson, Beric R

    2016-02-01

    Mutations in adenomatous polyposis coli (APC) disrupt regulation of Wnt signaling, mitosis, and the cytoskeleton. We describe a new role for APC in the transport of mitochondria. Silencing of wild-type APC by small interfering RNA caused mitochondria to redistribute from the cell periphery to the perinuclear region. We identified novel APC interactions with the mitochondrial kinesin-motor complex Miro/Milton that were mediated by the APC C-terminus. Truncating mutations in APC abolished its ability to bind Miro/Milton and reduced formation of the Miro/Milton complex, correlating with disrupted mitochondrial distribution in colorectal cancer cells that could be recovered by reconstitution of wild-type APC. Using proximity ligation assays, we identified endogenous APC-Miro/Milton complexes at mitochondria, and live-cell imaging showed that loss of APC slowed the frequency of anterograde mitochondrial transport to the membrane. We propose that APC helps drive mitochondria to the membrane to supply energy for cellular processes such as directed cell migration, a process disrupted by cancer mutations. PMID:26658612

  7. APC binds the Miro/Milton motor complex to stimulate transport of mitochondria to the plasma membrane

    PubMed Central

    Mills, Kate M.; Brocardo, Mariana G.; Henderson, Beric R.

    2016-01-01

    Mutations in adenomatous polyposis coli (APC) disrupt regulation of Wnt signaling, mitosis, and the cytoskeleton. We describe a new role for APC in the transport of mitochondria. Silencing of wild-type APC by small interfering RNA caused mitochondria to redistribute from the cell periphery to the perinuclear region. We identified novel APC interactions with the mitochondrial kinesin-motor complex Miro/Milton that were mediated by the APC C-terminus. Truncating mutations in APC abolished its ability to bind Miro/Milton and reduced formation of the Miro/Milton complex, correlating with disrupted mitochondrial distribution in colorectal cancer cells that could be recovered by reconstitution of wild-type APC. Using proximity ligation assays, we identified endogenous APC-Miro/Milton complexes at mitochondria, and live-cell imaging showed that loss of APC slowed the frequency of anterograde mitochondrial transport to the membrane. We propose that APC helps drive mitochondria to the membrane to supply energy for cellular processes such as directed cell migration, a process disrupted by cancer mutations. PMID:26658612

  8. APC promoter is frequently methylated in pancreatic juice of patients with pancreatic carcinomas or periampullary tumors

    PubMed Central

    Ginesta, Mireia M.; Diaz-Riascos, Zamira Vanessa; Busquets, Juli; Pelaez, Núria; Serrano, Teresa; Peinado, Miquel Àngel; Jorba, Rosa; García-Borobia, Francisco Javier; Capella, Gabriel; Fabregat, Joan

    2016-01-01

    Early detection of pancreatic and periampullary neoplasms is critical to improve their clinical outcome. The present authors previously demonstrated that DNA hypermethylation of adenomatous polyposis coli (APC), histamine receptor H2 (HRH2), cadherin 13 (CDH13), secreted protein acidic and cysteine rich (SPARC) and engrailed-1 (EN-1) promoters is frequently detected in pancreatic tumor cells. The aim of the present study was to assess their prevalence in pancreatic juice of carcinomas of the pancreas and periampullary area. A total of 135 pancreatic juices obtained from 85 pancreatic cancer (PC), 26 ampullary carcinoma (AC), 10 intraductal papillary mucinous neoplasm (IPMN) and 14 chronic pancreatitis (CP) patients were analyzed. The methylation status of the APC, HRH2, CDH13, SPARC and EN-1 promoters was analyzed using methylation specific-melting curve analysis (MS-MCA). Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations were also tested with allele-specific quantitative polymerase chain reaction amplification. Out of the 5 promoters analyzed, APC (71%) and HRH2 (65%) were the most frequently methylated in PC juice. APC methylation was also detected at a high frequency in AC (76%) and IPMN (80%), but only occasionally observed in CP (7%). APC methylation had a high sensitivity (71–80%) for all types of cancer analyzed. The panel (where a sample scored as positive when ≥2 markers were methylated) did not outperform APC as a single marker. Finally, KRAS detection in pancreatic juice offered a lower sensitivity (50%) and specificity (71%) for detection of any cancer. APC hypermethylation in pancreatic juice, as assessed by MS-MCA, is a frequent event of potential clinical usefulness in the diagnosis of pancreatic and periampullary neoplasms. PMID:27602165

  9. Detection and Analysis of Cell Cycle-Associated APC/C-Mediated Cellular Ubiquitylation In Vitro and In Vivo.

    PubMed

    Cedeño, Cesyen; La Monaca, Esther; Esposito, Mara; Gutierrez, Gustavo J

    2016-01-01

    The anaphase-promoting complex or cyclosome (APC/C) is one of the major orchestrators of the cell division cycle in mammalian cells. The APC/C acts as a ubiquitin ligase that triggers sequential ubiquitylation of a significant number of substrates which will be eventually degraded by proteasomes during major transitions of the cell cycle. In this chapter, we present accessible methodologies to assess both in in vitro conditions and in cellular systems ubiquitylation reactions mediated by the APC/C. In addition, we also describe techniques to evidence the changes in protein stability provoked by modulation of the activity of the APC/C. Finally, specific methods to analyze interactors or posttranslational modifications of particular APC/C subunits are also discussed. Given the crucial role played by the APC/C in the regulation of the cell cycle, this review only focuses on its action and effects in actively proliferating cells. PMID:27613041

  10. Paternal B Vitamin Intake Is a Determinant of Growth, Hepatic Lipid Metabolism and Intestinal Tumor Volume in Female Apc1638N Mouse Offspring

    PubMed Central

    Sabet, Julia A.; Park, Lara K.; Iyer, Lakshmanan K.; Tai, Albert K.; Koh, Gar Yee; Pfalzer, Anna C.; Parnell, Laurence D.; Mason, Joel B.; Liu, Zhenhua; Byun, Alexander J.; Crott, Jimmy W.

    2016-01-01

    Background The importance of maternal nutrition to offspring health and risk of disease is well established. Emerging evidence suggests paternal diet may affect offspring health as well. Objective In the current study we sought to determine whether modulating pre-conception paternal B vitamin intake alters intestinal tumor formation in offspring. Additionally, we sought to identify potential mechanisms for the observed weight differential among offspring by profiling hepatic gene expression and lipid content. Methods Male Apc1638N mice (prone to intestinal tumor formation) were fed diets containing replete (control, CTRL), mildly deficient (DEF), or supplemental (SUPP) quantities of vitamins B2, B6, B12, and folate for 8 weeks before mating with control-fed wild type females. Wild type offspring were euthanized at weaning and hepatic gene expression profiled. Apc1638N offspring were fed a replete diet and euthanized at 28 weeks of age to assess tumor burden. Results No differences in intestinal tumor incidence or burden were found between male Apc1638N offspring of different paternal diet groups. Although in female Apc1638N offspring there were no differences in tumor incidence or multiplicity, a stepwise increase in tumor volume with increasing paternal B vitamin intake was observed. Interestingly, female offspring of SUPP and DEF fathers had a significantly lower body weight than those of CTRL fed fathers. Moreover, hepatic trigylcerides and cholesterol were elevated 3-fold in adult female offspring of SUPP fathers. Weanling offspring of the same fathers displayed altered expression of several key lipid-metabolism genes. Hundreds of differentially methylated regions were identified in the paternal sperm in response to DEF and SUPP diets. Aside from a few genes including Igf2, there was a striking lack of overlap between these genes differentially methylated in sperm and differentially expressed in offspring. Conclusions In this animal model, modulation of

  11. LATS1 and LATS2 phosphorylate CDC26 to modulate assembly of the tetratricopeptide repeat subcomplex of APC/C.

    PubMed

    Masuda, Kenta; Chiyoda, Tatsuyuki; Sugiyama, Naoyuki; Segura-Cabrera, Aldo; Kabe, Yasuaki; Ueki, Arisa; Banno, Kouji; Banno, Koji; Suematsu, Makoto; Aoki, Daisuke; Ishihama, Yasushi; Saya, Hideyuki; Kuninaka, Shinji

    2015-01-01

    In budding yeast, the Mitotic Exit Network (MEN) regulates anaphase promoting complex/cyclosome (APC/C) via the Dbf2-Cdc14 signaling cascade. Dbf2 kinase phosphorylates and activates Cdc14 phosphatase, which removes the inhibitory phosphorylation of the APC/C cofactor Cdh1. Although each component of the MEN was highly conserved during evolution, there is presently no evidence supporting direct phosphorylation of CDC14 by large tumor suppressor kinase 1 (LATS1), the human counterpart of Dbf2; hence, it is unclear how LATS1 regulates APC/C. Here, we demonstrate that LATS1 phosphorylates the Thr7 (T7) residue of the APC/C component CDC26 directly. Nocodazole-induced phosphorylation of T7 was reduced by knockdown of LATS1 and LATS2 in HeLa cells, indicating that both of these kinases contribute to the phosphorylation of CDC26 in vivo. The T7 residue of CDC26 is critical for its interaction with APC6, a tetratricopeptide repeat-containing subunit of APC/C, and mutation of this residue to Asp (T7D) reduced the interaction of CDC26 with APC6. Replacement of endogenous CDC26 in HeLa cells with exogenous phosphor-mimic T7D-mutated CDC26 increased the elution size of APC/C subunits in a gel filtration assay, implying a change in the APC/C assembly upon phosphorylation of CDC26. Furthermore, T7D-mutated CDC26 promoted the ubiquitination of polo-like kinase 1, a well-known substrate of APC/C. Overall, these results suggest that LATS1/2 are novel kinases involved in APC/C phosphorylation and indicate a direct regulatory link between LATS1/2 and APC/C. PMID:25723520

  12. LATS1 and LATS2 Phosphorylate CDC26 to Modulate Assembly of the Tetratricopeptide Repeat Subcomplex of APC/C

    PubMed Central

    Masuda, Kenta; Chiyoda, Tatsuyuki; Sugiyama, Naoyuki; Segura-Cabrera, Aldo; Kabe, Yasuaki; Ueki, Arisa; Banno, Koji; Suematsu, Makoto; Aoki, Daisuke; Ishihama, Yasushi; Saya, Hideyuki; Kuninaka, Shinji

    2015-01-01

    In budding yeast, the Mitotic Exit Network (MEN) regulates anaphase promoting complex/cyclosome (APC/C) via the Dbf2-Cdc14 signaling cascade. Dbf2 kinase phosphorylates and activates Cdc14 phosphatase, which removes the inhibitory phosphorylation of the APC/C cofactor Cdh1. Although each component of the MEN was highly conserved during evolution, there is presently no evidence supporting direct phosphorylation of CDC14 by large tumor suppressor kinase 1 (LATS1), the human counterpart of Dbf2; hence, it is unclear how LATS1 regulates APC/C. Here, we demonstrate that LATS1 phosphorylates the Thr7 (T7) residue of the APC/C component CDC26 directly. Nocodazole-induced phosphorylation of T7 was reduced by knockdown of LATS1 and LATS2 in HeLa cells, indicating that both of these kinases contribute to the phosphorylation of CDC26 in vivo. The T7 residue of CDC26 is critical for its interaction with APC6, a tetratricopeptide repeat-containing subunit of APC/C, and mutation of this residue to Asp (T7D) reduced the interaction of CDC26 with APC6. Replacement of endogenous CDC26 in HeLa cells with exogenous phosphor-mimic T7D-mutated CDC26 increased the elution size of APC/C subunits in a gel filtration assay, implying a change in the APC/C assembly upon phosphorylation of CDC26. Furthermore, T7D-mutated CDC26 promoted the ubiquitination of polo-like kinase 1, a well-known substrate of APC/C. Overall, these results suggest that LATS1/2 are novel kinases involved in APC/C phosphorylation and indicate a direct regulatory link between LATS1/2 and APC/C. PMID:25723520

  13. The Apc(min) mouse has altered hematopoietic stem cell function and provides a model for MPD/MDS.

    PubMed

    Lane, Steven W; Sykes, Stephen M; Al-Shahrour, Fatima; Shterental, Sebastian; Paktinat, Mahnaz; Lo Celso, Cristina; Jesneck, Jonathan L; Ebert, Benjamin L; Williams, David A; Gilliland, D Gary

    2010-04-29

    Apc, a negative regulator of the canonical Wnt signaling pathway, is a bona-fide tumor suppressor whose loss of function results in intestinal polyposis. APC is located in a commonly deleted region on human chromosome 5q, associated with myelodysplastic syndrome (MDS), suggesting that haploinsufficiency of APC contributes to the MDS phenotype. Analysis of the hematopoietic system of mice with the Apc(min) allele that results in a premature stop codon and loss of function showed no abnormality in steady state hematopoiesis. Bone marrow derived from Apc(min) mice showed enhanced repopulation potential, indicating a cell intrinsic gain of function in the long-term hematopoietic stem cell (HSC) population. However, Apc(min) bone marrow was unable to repopulate secondary recipients because of loss of the quiescent HSC population. Apc(min) mice developed a MDS/myeloproliferative phenotype. Our data indicate that Wnt activation through haploinsufficiency of Apc causes insidious loss of HSC function that is only evident in serial transplantation strategies. These data provide a cautionary note for HSC-expansion strategies through Wnt pathway activation, provide evidence that cell extrinsic factors can contribute to the development of myeloid disease, and indicate that loss of function of APC may contribute to the phenotype observed in patients with MDS and del(5q). PMID:20197553

  14. The Spindle Assembly Checkpoint Is Not Essential for Viability of Human Cells with Genetically Lowered APC/C Activity

    PubMed Central

    Wild, Thomas; Larsen, Marie Sofie Yoo; Narita, Takeo; Schou, Julie; Nilsson, Jakob; Choudhary, Chunaram

    2016-01-01

    Summary The anaphase-promoting complex/cyclosome (APC/C) and the spindle assembly checkpoint (SAC), which inhibits the APC/C, are essential determinants of mitotic timing and faithful division of genetic material. Activation of the APC/C is known to depend on two APC/C-interacting E2 ubiquitin-conjugating enzymes—UBE2C and UBE2S. We show that APC/C activity in human cells is tuned by the combinatorial use of three E2s, namely UBE2C, UBE2S, and UBE2D. Genetic deletion of UBE2C and UBE2S, individually or in combination, leads to discriminative reduction in APC/C function and sensitizes cells to UBE2D depletion. Reduction of APC/C activity results in loss of switch-like metaphase-to-anaphase transition and, strikingly, renders cells insensitive to chemical inhibition of MPS1 and genetic ablation of MAD2, both of which are essential for the SAC. These results provide insights into the regulation of APC/C activity and demonstrate that the essentiality of the SAC is imposed by the strength of the APC/C. PMID:26904940

  15. Deconstructing the ßcatenin destruction complex: mechanistic roles for the tumor suppressor APC in regulating Wnt signaling.

    PubMed

    Roberts, David M; Pronobis, Mira I; Poulton, John S; Waldmann, Jon D; Stephenson, Elise M; Hanna, Shahnaz; Peifer, Mark

    2011-06-01

    Negatively regulating signaling by targeting key effectors for ubiquitination/destruction is essential for development and oncogenesis. The tumor suppressor adenomatous polyposis coli (APC), an essential negative regulator of Wnt signaling, provides a paradigm. APC mutations occur in most colon cancers. Acting in the "destruction complex" with Axin, glycogen synthase kinase 3, and casein kinase, APC targets ßcatenin (ßcat) for phosphorylation and recognition by an E3 ubiquitin-ligase. Despite 20 years of work, the internal workings of the destruction complex and APC's role remain largely mysterious. We use both Drosophila and colon cancer cells to test hypotheses for APC's mechanism of action. Our data are inconsistent with current models suggesting that high-affinity ßcat-binding sites on APC play key roles. Instead, they suggest that multiple ßcat-binding sites act additively to fine-tune signaling via cytoplasmic retention. We identify essential roles for two putative binding sites for new partners--20-amino-acid repeat 2 and conserved sequence B--in destruction complex action. Finally, we demonstrate that APC interacts with Axin by two different modes and provide evidence that conserved sequence B helps ensure release of APC from Axin, with disassembly critical in regulating ßcat levels. Using these data, we suggest a new model for destruction complex action in development, which also provides new insights into functions of truncated APC proteins in cancer. PMID:21471006

  16. Xenopus polo-like kinase Plx1 regulates XErp1, a novel inhibitor of APC/C activity

    PubMed Central

    Schmidt, Andreas; Duncan, Peter I.; Rauh, Nadine R.; Sauer, Guido; Fry, Andrew M.; Nigg, Erich A.; Mayer, Thomas U.

    2005-01-01

    Metaphase-to-anaphase transition is a fundamental step in cell cycle progression where duplicated sister-chromatids segregate to the future daughter cells. The anaphase-promoting complex/cyclosome (APC/C) is a highly regulated ubiquitin-ligase that triggers anaphase onset and mitotic exit by targeting securin and mitotic cyclins for destruction. It was previously shown that the Xenopus polo-like kinase Plx1 is essential to activate APC/C upon release from cytostatic factor (CSF) arrest in Xenopus egg extract. Although the mechanism by which Plx1 regulates APC/C activation remained unclear, the existence of a putative APC/C inhibitor was postulated whose activity would be neutralized by Plx1 upon CSF release. Here we identify XErp1, a novel Plx1-regulated inhibitor of APC/C activity, and we demonstrate that XErp1 is required to prevent anaphase onset in CSF-arrested Xenopus egg extract. Inactivation of XErp1 leads to premature APC/C activation. Conversely, addition of excess XErp1 to Xenopus egg extract prevents APC/C activation. Plx1 phosphorylates XErp1 in vitro at a site that targets XErp1 for degradation upon CSF release. Thus, our data lead to a model of APC/C activation in Xenopus egg extract in which Plx1 targets the APC/C inhibitor XErp1 for degradation. PMID:15713843

  17. AGILE integration into APC for high mix logic fab

    NASA Astrophysics Data System (ADS)

    Gatefait, M.; Lam, A.; Le Gratiet, B.; Mikolajczak, M.; Morin, V.; Chojnowski, N.; Kocsis, Z.; Smith, I.; Decaunes, J.; Ostrovsky, A.; Monget, C.

    2015-09-01

    mix logic Fab) in term of product and technology portfolio AGILE corrects for up to 120nm of product topography error on process layer with less than 50nm depth of focus Based on tool functionalities delivered by ASML and on high volume manufacturing requirement, AGILE integration is a real challenge. Regarding ST requirements "Automatic AGILE" functionality developed by ASML was not a turnkey solution and a dedicated functionality was needed. A "ST homemade AGILE integration" has been fully developed and implemented within ASML and ST constraints. This paper describes this integration in our Advanced Process Control platform (APC).

  18. Polarizing T and B Cell Responses by APC-Targeted Subunit Vaccines

    PubMed Central

    Grødeland, Gunnveig; Fossum, Even; Bogen, Bjarne

    2015-01-01

    Current influenza vaccines mostly aim at the induction of specific neutralizing antibodies. While antibodies are important for protection against a particular virus strain, T cells can recognize epitopes that will offer broader protection against influenza. We have previously developed a DNA vaccine format by which protein antigens can be targeted specifically to receptors on antigen presenting cells (APCs). The DNA-encoded vaccine proteins are homodimers, each chain consisting of a targeting unit, a dimerization unit, and an antigen. The strategy of targeting antigen to APCs greatly enhances immune responses as compared to non-targeted controls. Furthermore, targeting of antigen to different receptors on APCs can polarize the immune response to different arms of immunity. Here, we discuss how targeting of hemagglutinin to MHC class II molecules increases Th2 and IgG1 antibody responses, whereas targeting to chemokine receptors XCR1 or CCR1/3/5 increases Th1 and IgG2a responses, in addition to CD8+ T cell responses. We also discuss these results in relation to work published by others on APC-targeting. Differential targeting of APC surface molecules may allow the induction of tailor-made phenotypes of adaptive immune responses that are optimal for protection against various infectious agents, including influenza virus. PMID:26257735

  19. Polyethylene glycol inhibits intestinal neoplasia and induces epithelial apoptosis in Apc(min) mice.

    PubMed

    Roy, Hemant K; Gulizia, James; DiBaise, John K; Karolski, William J; Ansari, Sajid; Madugula, Madhavi; Hart, John; Bissonnette, Marc; Wali, Ramesh K

    2004-11-01

    Efficacy of a safe and clinically utilized polyethylene glycol formulation (PEG-3350) to suppress intestinal tumors was investigated in the Apc(min) mouse-model of experimental carcinogenesis. Furthermore, based on our previous finding on the induction of apoptosis in HT-29 cells by PEG, we evaluated its ability to stimulate epithelial cell apoptosis in both Apc(min) mouse as well as AOM-treated rat as a potential molecular mechanism of chemoprevention. Twenty-two Apc(min) mice were randomized equally to PEG or vehicle (control) supplementation. Tumors were scored and uninvolved intestinal mucosal apoptosis was assayed using a modified terminal deoxynucleotidyl transferase-mediated nick end-labeling (TUNEL) assay and by immunohistochemical detection of cleaved caspase-3. Supplementation of Apc(min) mice with 10% PEG 3350 (in drinking water) resulted in a 48% (P<0.05) reduction in intestinal tumor burden and induced 2-3 fold increase in mucosal apoptosis. Dietary supplementation of polyethylene glycol (5%) also stimulated colonic mucosal apoptosis 4-5 fold in AOM-treated rats, the regimen that we previously reported to reduce tumor burden by 76% (P<0.05). In summary, we demonstrate, for the first time, that PEG does protect against Apc(min) mouse tumorigenesis. The correlation between pro-apoptotic actions and chemopreventive efficacy of PEG in these models strongly implicates induction of apoptosis as one of the impending mechanisms of chemoprevention. PMID:15374630

  20. Co-regulation proteomics reveals substrates and mechanisms of APC/C-dependent degradation

    PubMed Central

    Singh, Sasha A; Winter, Dominic; Kirchner, Marc; Chauhan, Ruchi; Ahmed, Saima; Ozlu, Nurhan; Tzur, Amit; Steen, Judith A; Steen, Hanno

    2014-01-01

    Using multiplexed quantitative proteomics, we analyzed cell cycle-dependent changes of the human proteome. We identified >4,400 proteins, each with a six-point abundance profile across the cell cycle. Hypothesizing that proteins with similar abundance profiles are co-regulated, we clustered the proteins with abundance profiles most similar to known Anaphase-Promoting Complex/Cyclosome (APC/C) substrates to identify additional putative APC/C substrates. This protein profile similarity screening (PPSS) analysis resulted in a shortlist enriched in kinases and kinesins. Biochemical studies on the kinesins confirmed KIFC1, KIF18A, KIF2C, and KIF4A as APC/C substrates. Furthermore, we showed that the APC/CCDH1-dependent degradation of KIFC1 regulates the bipolar spindle formation and proper cell division. A targeted quantitative proteomics experiment showed that KIFC1 degradation is modulated by a stabilizing CDK1-dependent phosphorylation site within the degradation motif of KIFC1. The regulation of KIFC1 (de-)phosphorylation and degradation provides insights into the fidelity and proper ordering of substrate degradation by the APC/C during mitosis. PMID:24510915

  1. Production of geopolymers using glass produced from DC plasma treatment of air pollution control (APC) residues.

    PubMed

    Kourti, Ioanna; Rani, D Amutha; Deegan, D; Boccaccini, A R; Cheeseman, C R

    2010-04-15

    Air pollution control (APC) residues are the hazardous waste produced from cleaning gaseous emissions at energy-from-waste (EfW) facilities processing municipal solid waste (MSW). APC residues have been blended with glass-forming additives and treated using DC plasma technology to produce a high calcium alumino-silicate glass. This research has investigated the optimisation and properties of geopolymers prepared from this glass. Work has shown that high strength geopolymers can be formed and that the NaOH concentration of the activating solution significantly affects the properties. The broad particle size distribution of the APC residue glass used in these experiments results in a microstructure that contains unreacted glass particles included within a geopolymer binder phase. The high calcium content of APC residues may cause the formation of some amorphous calcium silicate hydrate (C-S-H) gel. A mix prepared with S/L=3.4, Si/Al=2.6 and [NaOH]=6M in the activating solution, produced high strength geopolymers with compressive strengths of approximately 130 MPa. This material had high density (2070 kg/m(3)) and low porosity. The research demonstrates for the first time that glass derived from DC plasma treatment of APC residues can be used to form high strength geopolymer-glass composites that have potential for use in a range of applications. PMID:20022170

  2. Modulation of APC Function and Anti-Tumor Immunity by Anti-Cancer Drugs

    PubMed Central

    Martin, Kea; Schreiner, Jens; Zippelius, Alfred

    2015-01-01

    Professional antigen-presenting cells (APCs), such as dendritic cells (DCs), are central to the initiation and regulation of anti-cancer immunity. However, in the immunosuppressive environment within a tumor APCs may antagonize anti-tumor immunity by inducing regulatory T cells (Tregs) or anergy of effector T cells due to lack of efficient costimulation. Hence, in an optimal setting, anti-cancer drugs have the power to reduce tumor size and thereby may induce the release of tumor antigens and, at the same time, modulate APC function toward efficient priming of antigen-specific effector T cells. Selected cytotoxic agents may revert APC dysfunction either by directly maturing DCs or through induction of immunogenic tumor cell death. Furthermore, specific cytotoxic agents may support adaptive immunity by selectively depleting regulatory subsets, such as Tregs or myeloid-derived suppressor cells. Perspectively, this will allow developing effective combination strategies with novel immunotherapies to exert complementary pressure on tumors via direct toxicity as well as immune activation. We, here, review our current knowledge on the capacity of anti-cancer drugs to modulate APC functions to promote durable anti-cancer immune responses. PMID:26483791

  3. Arid1a inactivation in an Apc- and Pten-defective mouse ovarian cancer model enhances epithelial differentiation and prolongs survival.

    PubMed

    Zhai, Yali; Kuick, Rork; Tipton, Courtney; Wu, Rong; Sessine, Michael; Wang, Zhong; Baker, Suzanne J; Fearon, Eric R; Cho, Kathleen R

    2016-01-01

    Inactivation of the ARID1A tumour suppressor gene is frequent in ovarian endometrioid (OEC) and clear cell (OCCC) carcinomas, often in conjunction with mutations activating the PI3K-AKT and/or canonical Wnt signalling pathways. Prior work has shown that conditional bi-allelic inactivation of the Apc and Pten tumour suppressor genes in the mouse ovarian surface epithelium (OSE) promotes outgrowth of tumours that reflect the biological behaviour and gene expression profiles of human OECs harbouring comparable Wnt and PI3K-AKT pathway defects, although the mouse tumours are more poorly differentiated than their human tumour counterparts. We found that conditional inactivation of one or both Arid1a alleles in OSE concurrently with Apc and Pten inactivation unexpectedly prolonged the survival of tumour-bearing mice and promoted striking epithelial differentiation of the cancer cells, resulting in morphological features akin to those in human OECs. Enhanced epithelial differentiation was linked to reduced expression of the mesenchymal markers N-cadherin and vimentin, and increased expression of the epithelial markers Crb3 and E-cadherin. Global gene expression profiling showed enrichment for genes associated with mesenchymal-epithelial transition in the Arid1a-deficient tumours. We also found that an activating (E545K) Pik3ca mutation, unlike Pten inactivation or Pik3ca H1047R mutation, cannot cooperate with Arid1a loss to promote ovarian cancer development in the mouse. Our results indicate that the Arid1a tumour suppressor gene has a key role in regulating OEC differentiation, and paradoxically the mouse cancers with more initiating tumour suppressor gene defects had a less aggressive phenotype than cancers arising from fewer gene alterations. Microarray data have been deposited in NCBI's Gene Expression Omnibus (GSE67695). PMID:26279473

  4. Obesity promotes PhIP-induced small intestinal carcinogenesis in hCYP1A-db/db mice: involvement of mutations and DNA hypermethylation of Apc.

    PubMed

    Wang, Hong; Liu, Anna; Kuo, Yingyi; Chi, Eric; Yang, Xu; Zhang, Lanjing; Yang, Chung S

    2016-07-01

    Obesity is associated with an increased risk of cancer. To study the promotion of dietary carcinogen-induced gastrointestinal cancer by obesity, we employed 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) to induce intestinal tumorigenesis in CYP1A-humanized (hCYP1A) mice, in which mouse Cyp1a1/1a2 was replaced with human CYP1A1/1A2 Obesity was introduced in hCYP1A mice by breeding with Lepr(db/+) mice to establish the genetically induced obese hCYP1A-Lepr(db/db) mice or by feeding hCYP1A mice a high-fat diet. PhIP induced the formation of small intestinal tumors at the ages of weeks 28-40 in obese hCYP1A mice, but not in lean hCYP1A mice. No tumors were found in colon and other gastrointestinal organs in the lean or obese mice. Using immunohistochemistry (IHC), we found strong positive staining of NF-κB p65, pSTAT3 and COX2 as well as elevated levels of nuclear β-catenin (Ctnnb1) in small intestinal tumors, but not in normal tissues. By sequencing Apc and Ctnnb1 genes, we found that most PhIP-induced small intestinal tumors in obese mice carried only a single heterozygous mutation in Apc By bisulfite-sequencing of CpG islands of Apc, we found DNA hypermethylation in a CpG cluster located in its transcription initiation site, which most likely caused the inactivation of the wild-type Apc allele. Our findings demonstrate that PhIP-induced small intestinal carcinogenesis in hCYP1A-db/db mice is promoted by obesity and involves Apc mutation and inactivation by DNA hypermethylation. This experimental result is consistent with the association of obesity and the increased incidence of small intestinal cancer in humans in recent decades. PMID:27207656

  5. Characterization of elastic-plastic properties of AS4/APC-2 thermoplastic composite

    NASA Technical Reports Server (NTRS)

    Sun, C. T.; Yoon, K. J.

    1988-01-01

    Elastic and inelastic properties of AS4/APC-2 composites were characterized with respect to temperature variation by using a one-parameter orthotropic plasticity model and a one parameter failure criterion. Simple uniaxial off-axis tension tests were performed on coupon specimens of unidirectional AS4/APC-2 thermoplastic composite at various temperatures. To avoid the complication caused by the extension-shear coupling effect in off-axis testing, new tabs were designed and used on the test specimens. The experimental results showed that the nonlinear behavior of constitutive relations and the failure strengths can be characterized quite well using the one parameter plasticity model and the failure criterion, respectively.

  6. Orthotropic elasto-plastic behavior of AS4/APC-2 thermoplastic composite in compression

    NASA Technical Reports Server (NTRS)

    Sun, C. T.; Rui, Y.

    1989-01-01

    Uniaxial compression tests were performed on off-axis coupon specimens of unidirectional AS4/APC-2 thermoplastic composite at various temperatures. The elasto-plastic and strength properties of AS4/APC-2 composite were characterized with respect to temperature variation by using a one-parameter orthotropic plasticity model and a one-parameter failure criterion. Experimental results show that the orthotropic plastic behavior can be characterized quite well using the plasticity model, and the matrix-dominant compressive strengths can be predicted very accurately by the one-parameter failure criterion.

  7. Multiple mucin depleted foci, high proliferation and low apoptotic response in the onset of colon carcinogenesis of the PIRC rat, mutated in Apc.

    PubMed

    Femia, Angelo Pietro; Luceri, Cristina; Soares, Paulo Victoria; Lodovici, Maura; Caderni, Giovanna

    2015-03-15

    PIRC rats (F344/NTac-Apc (am1137) ) mutated in the Apc gene spontaneously develop colon tumors thus mimicking familial adenomatous polyposis (FAP) and sporadic colorectal cancer (CRC) more closely than Apc-based rodent models developing tumors mostly in the small intestine. To understand whether microscopic dysplastic lesions precede the development of macroscopic tumors, PIRC rat colon was examined for the presence of mucin depleted foci (MDF), microadenomas of the rodent and human colon. Few MDF (about 4/animal) were already present in 1-month-old rats and their number rapidly increases to about 250 in 8-month-old rats. These lesions showed Wnt signaling activation (nuclear β-catenin accumulation) and were dramatically decreased by sulindac (320 ppm), a drug with chemopreventive activity (MDF/rat at 4 months: 156 ± 8 and 38 ± 6 in controls and sulindac-treated rats, respectively, means ± SE, p < 0.001). Since altered proliferation and apoptosis could underlie the early phases of carcinogenesis, we studied these processes in the apparently normal colon mucosa (NM) of 1-month-old PIRC and wt rats. Colon proliferation (PCNA expression) was significantly higher in PIRC rats. Notably, PIRC rat NM showed resistance to apoptosis since it sustained proliferation and had lower apoptosis after a cytotoxic insult with 1,2 dimethylhydrazine. Gene expression of Myc, p21, Birc5, Ogg1, Apex1 and Sod2 were significantly up-regulated in the NM of PIRC rat. The overall results put forward PIRC rat as useful model of colon carcinogenesis, either to study the process itself or to test in vivo chemopreventive agents in both short- and long-term studies. PMID:25257656

  8. An APC/C-Cdh1 Biosensor Reveals the Dynamics of Cdh1 Inactivation at the G1/S Transition

    PubMed Central

    Ondracka, Andrej; Robbins, Jonathan A.; Cross, Frederick R.

    2016-01-01

    B-type cyclin-dependent kinase activity must be turned off for mitotic exit and G1 stabilization. B-type cyclin degradation is mediated by the anaphase-promoting complex/cyclosome (APC/C); during and after mitotic exit, APC/C is dependent on Cdh1. Cdh1 is in turn phosphorylated and inactivated by cyclin-CDK at the Start transition of the new cell cycle. We developed a biosensor to assess the cell cycle dynamics of APC/C-Cdh1. Nuclear exit of the G1 transcriptional repressor Whi5 is a known marker of Start; APC/C-Cdh1 is inactivated 12 min after Whi5 nuclear exit with little measurable cell-to-cell timing variability. Multiple phosphorylation sites on Cdh1 act in a redundant manner to repress its activity. Reducing the number of phosphorylation sites on Cdh1 can to some extent be tolerated for cell viability, but it increases variability in timing of APC/C-Cdh1 inactivation. Mutants with minimal subsets of phosphorylation sites required for viability exhibit striking stochasticity in multiple responses including budding, nuclear division, and APC/C-Cdh1 activity itself. Multiple cyclin-CDK complexes, as well as the stoichiometric inhibitor Acm1, contribute to APC/C-Cdh1 inactivation; this redundant control is likely to promote rapid and reliable APC/C-Cdh1 inactivation immediately following the Start transition. PMID:27410035

  9. Dual control by Cdk1 phosphorylation of the budding yeast APC/C ubiquitin ligase activator Cdh1.

    PubMed

    Höckner, Sebastian; Neumann-Arnold, Lea; Seufert, Wolfgang

    2016-07-15

    The antagonism between cyclin-dependent kinases (Cdks) and the ubiquitin ligase APC/C-Cdh1 is central to eukaryotic cell cycle control. APC/C-Cdh1 targets cyclin B and other regulatory proteins for degradation, whereas Cdks disable APC/C-Cdh1 through phosphorylation of the Cdh1 activator protein at multiple sites. Budding yeast Cdh1 carries nine Cdk phosphorylation sites in its N-terminal regulatory domain, most or all of which contribute to inhibition. However, the precise role of individual sites has remained unclear. Here, we report that the Cdk phosphorylation sites of yeast Cdh1 are organized into autonomous subgroups and act through separate mechanisms. Cdk sites 1-3 had no direct effect on the APC/C binding of Cdh1 but inactivated a bipartite nuclear localization sequence (NLS) and thereby controlled the partitioning of Cdh1 between cytoplasm and nucleus. In contrast, Cdk sites 4-9 did not influence the cell cycle-regulated localization of Cdh1 but prevented its binding to the APC/C. Cdk sites 4-9 reside near two recently identified APC/C interaction motifs in a pattern conserved with the human Cdh1 orthologue. Thus a Cdk-inhibited NLS goes along with Cdk-inhibited APC/C binding sites in yeast Cdh1 to relay the negative control by Cdk1 phosphorylation of the ubiquitin ligase APC/C-Cdh1. PMID:27226481

  10. APC/C-Cdh1 coordinates neurogenesis and cortical size during development

    NASA Astrophysics Data System (ADS)

    Delgado-Esteban, Maria; García-Higuera, Irene; Maestre, Carolina; Moreno, Sergio; Almeida, Angeles

    2013-12-01

    The morphology of the adult brain is the result of a delicate balance between neural progenitor proliferation and the initiation of neurogenesis in the embryonic period. Here we assessed whether the anaphase-promoting complex/cyclosome (APC/C) cofactor, Cdh1—which regulates mitosis exit and G1-phase length in dividing cells—regulates neurogenesis in vivo. We use an embryo-restricted Cdh1 knockout mouse model and show that functional APC/C-Cdh1 ubiquitin ligase activity is required for both terminal differentiation of cortical neurons in vitro and neurogenesis in vivo. Further, genetic ablation of Cdh1 impairs the ability of APC/C to promote neurogenesis by delaying the exit of the progenitor cells from the cell cycle. This causes replicative stress and p53-mediated apoptotic death resulting in decreased number of cortical neurons and cortex size. These results demonstrate that APC/C-Cdh1 coordinates cortical neurogenesis and size, thus posing Cdh1 in the molecular pathogenesis of congenital neurodevelopmental disorders, such as microcephaly.

  11. The future of APCs: a look at Medicare's 2005 changes and industry trends.

    PubMed

    Leary, Renee S; Farley, Dean

    2005-03-01

    As the healthcare industry continues to struggle with an ever-evolving APC-based hospital outpatient PPS, it's important to be aware of some of the latest changes affecting providers and to recognize areas that may present particular challenges for private payers. PMID:17233243

  12. Control of APC/C-dependent ubiquitin chain elongation by reversible phosphorylation

    PubMed Central

    Craney, Allison; Kelly, Aileen; Jia, Luying; Fedrigo, Indro; Yu, Hongtao; Rape, Michael

    2016-01-01

    Most metazoan E3 ligases contain a signature RING domain that promotes the transfer of ubiquitin from the active site of E2 conjugating enzymes to lysine residues in substrates. Although these RING-E3s depend on E2 enzymes for catalysis, how they turn on their E2s at the right time and place remains poorly understood. Here we report a phosphorylation-dependent mechanism that ensures timely activation of the E2 Ube2S by its RING-E3, the anaphase-promoting complex (APC/C); while phosphorylation of a specific serine residue in the APC/C coactivator Cdc20 prevents delivery of Ube2S to the APC/C, removal of this mark by PP2AB56 allows Ube2S to bind the APC/C and catalyze ubiquitin chain elongation. PP2AB56 also stabilizes kinetochore–microtubule attachments to shut off the spindle checkpoint, suggesting that cells regulate the E2–E3 interplay to coordinate ubiquitination with critical events during cell division. PMID:26811472

  13. Control of APC/C-dependent ubiquitin chain elongation by reversible phosphorylation.

    PubMed

    Craney, Allison; Kelly, Aileen; Jia, Luying; Fedrigo, Indro; Yu, Hongtao; Rape, Michael

    2016-02-01

    Most metazoan E3 ligases contain a signature RING domain that promotes the transfer of ubiquitin from the active site of E2 conjugating enzymes to lysine residues in substrates. Although these RING-E3s depend on E2 enzymes for catalysis, how they turn on their E2s at the right time and place remains poorly understood. Here we report a phosphorylation-dependent mechanism that ensures timely activation of the E2 Ube2S by its RING-E3, the anaphase-promoting complex (APC/C); while phosphorylation of a specific serine residue in the APC/C coactivator Cdc20 prevents delivery of Ube2S to the APC/C, removal of this mark by PP2A(B56) allows Ube2S to bind the APC/C and catalyze ubiquitin chain elongation. PP2A(B56) also stabilizes kinetochore-microtubule attachments to shut off the spindle checkpoint, suggesting that cells regulate the E2-E3 interplay to coordinate ubiquitination with critical events during cell division. PMID:26811472

  14. Application of accelerated carbonation on MSW combustion APC residues for metal immobilization and CO2 sequestration.

    PubMed

    Cappai, G; Cara, S; Muntoni, A; Piredda, M

    2012-03-15

    The present study focuses on the application of an aqueous phase accelerated carbonation treatment on air pollution control (APC) residues from municipal solid waste combustion, aimed at assessing its influence on the environmental behaviour of the residue under concern, as well as the potential of the process in terms of sequestration of the CO2. APC residues are considered hazardous waste and must be treated before final disposal in order to achieve the immobilization/mobilization of critical contaminants such as heavy metals as well as mobilization of soluble salts. The treatment applied proved to be effective in reducing the mobility of Pb, Zn, Cr, Cu and Mo, the optimum final pH for the carbonated APC residues being in a range of 10-10.5, whilst a mobilization effect was noticed for Sb and no effect was assessed for chlorides. The effect of carbonation treatment on the contaminant release was further evaluated by means of a sequential extraction procedure, indicating that the distribution of contaminants on water soluble, exchangeable and carbonate fraction was modified after treatment. The CO2 sequestration potential assessed for the APC residues showed that the carbonation technology could be a technically viable option in order to reduce emissions from WtE plants. PMID:21601357

  15. A novel tankyrase small-molecule inhibitor suppresses APC mutation-driven colorectal tumor growth.

    PubMed

    Lau, Ted; Chan, Emily; Callow, Marinella; Waaler, Jo; Boggs, Jason; Blake, Robert A; Magnuson, Steven; Sambrone, Amy; Schutten, Melissa; Firestein, Ron; Machon, Ondrej; Korinek, Vladimir; Choo, Edna; Diaz, Dolores; Merchant, Mark; Polakis, Paul; Holsworth, Daniel D; Krauss, Stefan; Costa, Mike

    2013-05-15

    Most colorectal cancers (CRC) are initiated by mutations of APC, leading to increased β-catenin-mediated signaling. However, continued requirement of Wnt/β-catenin signaling for tumor progression in the context of acquired KRAS and other mutations is less well-established. To attenuate Wnt/β-catenin signaling in tumors, we have developed potent and specific small-molecule tankyrase inhibitors, G007-LK and G244-LM, that reduce Wnt/β-catenin signaling by preventing poly(ADP-ribosyl)ation-dependent AXIN degradation, thereby promoting β-catenin destabilization. We show that novel tankyrase inhibitors completely block ligand-driven Wnt/β-catenin signaling in cell culture and display approximately 50% inhibition of APC mutation-driven signaling in most CRC cell lines. It was previously unknown whether the level of AXIN protein stabilization by tankyrase inhibition is sufficient to impact tumor growth in the absence of normal APC activity. Compound G007-LK displays favorable pharmacokinetic properties and inhibits in vivo tumor growth in a subset of APC-mutant CRC xenograft models. In the xenograft model most sensitive to tankyrase inhibitor, COLO-320DM, G007-LK inhibits cell-cycle progression, reduces colony formation, and induces differentiation, suggesting that β-catenin-dependent maintenance of an undifferentiated state may be blocked by tankyrase inhibition. The full potential of the antitumor activity of G007-LK may be limited by intestinal toxicity associated with inhibition of Wnt/β-catenin signaling and cell proliferation in intestinal crypts. These results establish proof-of-concept antitumor efficacy for tankyrase inhibitors in APC-mutant CRC models and uncover potential diagnostic and safety concerns to be overcome as tankyrase inhibitors are advanced into the clinic. PMID:23539443

  16. Targeting of Fzr/Cdh1 for timely activation of the APC/C at the centrosome during mitotic exit

    PubMed Central

    Meghini, Francesco; Martins, Torcato; Tait, Xavier; Fujimitsu, Kazuyuki; Yamano, Hiroyuki; Glover, David M.; Kimata, Yuu

    2016-01-01

    A multi-subunit ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C), regulates critical cellular processes including the cell cycle. To accomplish its diverse functions, APC/C activity must be precisely regulated in time and space. The interphase APC/C activator Fizzy-related (Fzr or Cdh1) is localized at centrosomes in animal cells. However, neither the mechanism of its localization nor its importance is clear. Here we identify the centrosome component Spd2 as a major partner of Fzr in Drosophila. The localization of Fzr to the centriole during interphase depends on direct interaction with Spd2. By generating Spd2 mutants unable to bind Fzr, we show that centrosomal localization of Fzr is essential for optimal APC/C activation towards its centrosomal substrate Aurora A. Finally, we show that Spd2 is also a novel APC/CFzr substrate. Our study is the first to demonstrate the critical importance of distinct subcellular pools of APC/C activators in the spatiotemporal control of APC/C activity. PMID:27558644

  17. Regulation of intracellular beta-catenin levels by the adenomatous polyposis coli (APC) tumor-suppressor protein.

    PubMed Central

    Munemitsu, S; Albert, I; Souza, B; Rubinfeld, B; Polakis, P

    1995-01-01

    The APC tumor-suppressor protein associates with beta-catenin, a cell adhesion protein that is upregulated by the WNT1 oncogene. We examined the effects of exogenous APC expression on the distribution and amount of beta-catenin in a colorectal cancer cell containing only mutant APC. Expression of wild-type APC caused a pronounced reduction in total beta-catenin levels by eliminating an excessive supply of cytoplasmic beta-catenin indigenous to the SW480 colorectal cancer cell line. This reduction was due to an enhanced rate of beta-catenin protein degradation. Truncated mutant APC proteins, characteristic of those associated with cancer, lacked this activity. Mutational analysis revealed that the central region of the APC protein, which is typically deleted or severely truncated in tumors, was responsible for the down-regulation of beta-catenin. These results suggest that the tumor-suppressor activity of mutant APC may be compromised due to a defect in its ability to regulate beta-catenin. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:7708772

  18. Cryo-EM of Mitotic Checkpoint Complex-Bound APC/C Reveals Reciprocal and Conformational Regulation of Ubiquitin Ligation.

    PubMed

    Yamaguchi, Masaya; VanderLinden, Ryan; Weissmann, Florian; Qiao, Renping; Dube, Prakash; Brown, Nicholas G; Haselbach, David; Zhang, Wei; Sidhu, Sachdev S; Peters, Jan-Michael; Stark, Holger; Schulman, Brenda A

    2016-08-18

    The mitotic checkpoint complex (MCC) coordinates proper chromosome biorientation on the spindle with ubiquitination activities of CDC20-activated anaphase-promoting complex/cyclosome (APC/C(CDC20)). APC/C(CDC20) and two E2s, UBE2C and UBE2S, catalyze ubiquitination through distinct architectures for linking ubiquitin (UB) to substrates and elongating polyUB chains, respectively. MCC, which contains a second molecule of CDC20, blocks APC/C(CDC20)-UBE2C-dependent ubiquitination of Securin and Cyclins, while differentially determining or inhibiting CDC20 ubiquitination to regulate spindle surveillance, checkpoint activation, and checkpoint termination. Here electron microscopy reveals conformational variation of APC/C(CDC20)-MCC underlying this multifaceted regulation. MCC binds APC/C-bound CDC20 to inhibit substrate access. However, rotation about the CDC20-MCC assembly and conformational variability of APC/C modulate UBE2C-catalyzed ubiquitination of MCC's CDC20 molecule. Access of UBE2C is limiting for subsequent polyubiquitination by UBE2S. We propose that conformational dynamics of APC/C(CDC20)-MCC modulate E2 activation and determine distinctive ubiquitination activities as part of a response mechanism ensuring accurate sister chromatid segregation. PMID:27522463

  19. APC Activation Restores Functional CD4+CD25+ Regulatory T Cells in NOD Mice that Can Prevent Diabetes Development

    PubMed Central

    Manirarora, Jean N.; Kosiewicz, Michele M.; Parnell, Sarah A.; Alard, Pascale

    2008-01-01

    Background Defects in APC and regulatory cells are associated with diabetes development in NOD mice. We have shown previously that NOD APC are not effective at stimulating CD4+CD25+ regulatory cell function in vitro. We hypothesize that failure of NOD APC to properly activate CD4+CD25+ regulatory cells in vivo could compromise their ability to control pathogenic cells, and activation of NOD APC could restore this defect, thereby preventing disease. Methodology/Principal Findings To test these hypotheses, we used the well-documented ability of complete Freund's adjuvant (CFA), an APC activator, to prevent disease in NOD mice. Phenotype and function of CD4+CD25+ regulatory cells from untreated and CFA-treated NOD mice were determined by FACS, and in vitro and in vivo assays. APC from these mice were also evaluated for their ability to activate regulatory cells in vitro. We have found that sick NOD CD4+CD25+ cells expressed Foxp3 at the same percentages, but decreased levels per cell, compared to young NOD or non-NOD controls. Treatment with CFA increased Foxp3 expression in NOD cells, and also increased the percentages of CD4+CD25+Foxp3+ cells infiltrating the pancreas compared to untreated NOD mice. Moreover, CD4+CD25+ cells from pancreatic LN of CFA-treated, but not untreated, NOD mice transferred protection from diabetes. Finally, APC isolated from CFA-treated mice increased Foxp3 and granzyme B expression as well as regulatory function by NOD CD4+CD25+ cells in vitro compared to APC from untreated NOD mice. Conclusions/Significance These data suggest that regulatory T cell function and ability to control pathogenic cells can be enhanced in NOD mice by activating NOD APC. PMID:19011680

  20. Sugar-carrying Polystyrenes Facilitate Harvesting of APCs from MLRs: Possible Application of Sugar-carrying Polystyrenes to Immunotherapy.

    PubMed

    Imaizumi, Akira; Onishi, Hideya; Yamasaki, Akio; Kawamoto, Makoto; Morisaki, Takashi; Goto, Mitsuaki; Iwama, Masamichi; Akaike, Toshihiro; Hasumi, Kenichiro

    2016-02-01

    Antigen-presenting cells (APCs) play a pivotal role in cancer immunotherapy. APCs in conventionally used flasks are harvested by enzymatic digestion or cell scraping for application to cancer immunotherapy. However, these methods may impair functional molecules expressed on the APC surface and reduce their effects in cancer immunotherapy. Recently, we found that APCs could be harvested by shaking at 4°C in flasks coated with poly[N-p-vinylbenzyl-O-2-acetoamide-2-deoxy-β-D-glucopyranosyl-(1→4)-2-acetoamide-2-deoxy-β-D-gluconamide] (PVGlcNAc) or a copolymer consisting of sulfonylurea (SU) linked to poly[N-p-vinyl-benzyl-4-O-β-D-galactopyranosyl-D-gluconamide] [P(VLA-co-SU)]. In the present study, we compared the functions of cytotoxic T-lymphocytes (CTLs) induced by APCs generated in PVGlcNAc- or P(VLA-co-SU)-coated flasks and conventional flasks. APCs from PVGlcNAc- or P(VLA-co-SU)-coated flasks showed higher expression of cluster of differentiation (CD)80/86, CD11c, and major histocompatibility complex class II alloantigen I-A(d), and higher cytotoxicity than APCs from conventional flasks. These results suggest that the use of PVGlcNAc- or P(VLA-co-SU)-coated flasks is optimal for harvesting APCs. The generated APCs also have a higher antigen-presenting ability compared to those generated in conventional flasks. Our results may contribute to the development of effective cancer immunotherapies. PMID:26851023

  1. Analysing the impact of nucleo-cytoplasmic shuttling of β-catenin and its antagonists APC, Axin and GSK3 on Wnt/β-catenin signalling.

    PubMed

    Schmitz, Yvonne; Rateitschak, Katja; Wolkenhauer, Olaf

    2013-11-01

    The canonical Wnt signalling pathway plays a critical role in development and disease. The key player of the pathway is β-catenin. Its activity is mainly regulated by the destruction complex consisting of APC, Axin and GSK3. In the nucleus, the complex formation of β-catenin and TCF initiates target gene expression. Our study provides a comprehensive analysis of the role of nucleo-cytoplasmic shuttling of APC, Axin, and GSK3 and the inactivation of β-catenin by the destruction complex in Wnt/β-catenin signalling. We address the following questions: Can nucleo-cytoplasmic shuttling of APC, Axin and GSK3 increase the [β-catenin/TCF] concentration? And, how is the [β-catenin/TCF] concentration influenced by phosphorylation and subsequent degradation of nuclear β-catenin? Based on experimental findings, we develop a compartmental model and conduct several simulation experiments. Our analysis reveals the following key findings: 1) nucleo-cytoplasmic shuttling of β-catenin and its antagonists can yield a spatial separation between the said proteins, which results in a breakdown of β-catenin degradation, followed by an accumulation of β-catenin and hence leads to an increase of the [β-catenin/TCF] concentration. Our results strongly suggest that Wnt signalling can benefit from nucleo-cytoplasmic shuttling of APC, Axin and GSK3, although they are in general β-catenin antagonising proteins. 2) The total robustness of the [β-catenin/TCF] output is closely linked to its absolute concentration levels. We demonstrate that the compartmental separation of β-catenin and the destruction complex does not only lead to a maximization, but additionally to an increased robustness of [β-catenin/TCF] signalling against perturbations in the cellular environment. 3) A nuclear accumulation of the destruction complex renders the pathway robust against fluctuations in Wnt signalling and against changes in the compartmental distribution of β-catenin. 4) Elucidating the impact of

  2. APC targeting enhances immunogenicity of a novel multistage Fc-fusion tuberculosis vaccine in mice.

    PubMed

    Soleimanpour, Saman; Farsiani, Hadi; Mosavat, Arman; Ghazvini, Kiarash; Eydgahi, Mohammad Reza Akbari; Sankian, Mojtaba; Sadeghian, Hamid; Meshkat, Zahra; Rezaee, Seyed Abdolrahim

    2015-12-01

    Numerous studies have demonstrated that targeting immunogens to FcγR on antigen-presenting cells (APCs) can selectively uptake and increase cellular immunity in vitro and in vivo. Therefore, the present study was conducted to evaluate immunogenicity of a novel multistage tuberculosis vaccine, a combination of an early and a dormant immunogenic protein, ESAT6 and HspX, fused to Fcγ2a fragment of mouse IgG2a to target all forms of tuberculosis. Codon-optimized genes consisting of ESAT6, a linker, and HspX fused either to mouse Fcγ2a (ESAT6:HspX:mFcγ2a) or 6× His-tag (ESAT6:HspX:His) were synthesized. The resulting proteins were then produced in Pichia pastoris. The fusion proteins were separately emulsified in dimethyldioctadecylammonium bromide(DDA)-trehalose-6,6-dibehenate(TDB) adjuvant, and their immunogenicity with and without bacille Calmette-Guérin (BCG) was assessed in C57BL/6 mice. Th1, Th2, Th17, and T-reg cytokine patterns were evaluated using the ELISA method. Both multistage vaccines induced very strong IL-12 and IFN-γ secretion from splenic cells; the Fc-tagged subunit vaccine induced a more effective Th1 immune response (IFN-γ, 910 pg/mL, and IL-12, 854 pg/mL) with a very low increase in IL-17 (∼0.1 pg/mL) and IL-4 (37 pg/mL) and a mild increase in TGF-β (543 pg/mL) compared to the BCG or ESAT6:HspX:His primed and boosted groups. The production of IFN-γ to ESAT6:HspX:Fcγ2a was very consistent and showed an increasing trend for IL-12 compared to the BCG or ESAT6:HspX:His primed and boosted groups. Fcγ2a used as a delivery vehicle supported the idea of selective uptake, inducing cross-presentation and forming a proper anti-tuberculosis response in context of Th1/Th2 and Th17/T-reg balances, which is important for protection and prevention of damage. PMID:26373723

  3. APC/β-catenin-rich complexes at membrane protrusions regulate mammary tumor cell migration and mesenchymal morphology

    PubMed Central

    2013-01-01

    Background The APC tumor suppressor is mutated or downregulated in many tumor types, and is prominently localized to punctate clusters at protrusion tips in migratory cells, such as in astrocytes where it has been implicated in directed cell motility. Although APC loss is considered an initiating event in colorectal cancer, for example, it is less clear what role APC plays in tumor cell motility and whether loss of APC might be an important promoter of tumor progression in addition to initiation. Methods The localization of APC and β-catenin was analyzed in multiple cell lines, including non-transformed epithelial lines treated with a proteasome inhibitor or TGFβ to induce an epithelial-to-mesenchymal transition (EMT), as well as several breast cancer lines, by immunofluorescence. APC expression was knocked down in 4T07 mammary tumor cells using lentiviral-mediated delivery of APC-specific short-hairpin (sh) RNAs, and assessed using quantitative (q) reverse-transcriptase (RT)-PCR and western blotting. Tumor cell motility was analyzed by performing wound-filling assays, and morphology via immunofluorescence (IF) and phase-contrast microscopy. Additionally, proliferation was measured using BrdU incorporation, and TCF reporter assays were performed to determine β-catenin/TCF-mediated transcriptional activity. Results APC/β-catenin-rich complexes were observed at protrusion ends of migratory epithelial cells treated with a proteasome inhibitor or when EMT has been induced and in tumor cells with a mesenchymal, spindle-like morphology. 4T07 tumor cells with reduced APC levels were significantly less motile and had a more rounded morphology; yet, they did not differ significantly in proliferation or β-catenin/TCF transcriptional activity. Furthermore, we found that APC/β-catenin-rich complexes at protrusion ends were dependent upon an intact microtubule cytoskeleton. Conclusions These findings indicate that membrane protrusions with APC/β-catenin-containing puncta

  4. Reversible modification of adenomatous polyposis coli (APC) with K63-linked polyubiquitin regulates the assembly and activity of the β-catenin destruction complex.

    PubMed

    Tran, Hoanh; Polakis, Paul

    2012-08-17

    The adenomatous polyposis coli (APC) tumor suppressor forms a complex with Axin and GSK3β to promote the phosphorylation and degradation of β-catenin, a key co-activator of Wnt-induced transcription. Here, we establish that APC is modified predominantly with K63-linked ubiquitin chains when it is bound to Axin in unstimulated HEK293 cells. Wnt3a stimulation induced a time-dependent loss of K63-polyubiquitin adducts from APC, an effect synchronous with the dissociation of Axin from APC and the stabilization of cytosolic β-catenin. RNAi-mediated depletion of Axin or β-catenin, which negated the association between APC and Axin, resulted in the absence of K63-adducts on APC. Overexpression of wild-type and phosphodegron-mutant β-catenin, combined with analysis of thirteen human cancer cell lines that harbor oncogenic mutations in APC, Axin, or β-catenin, support the hypothesis that a fully assembled APC-Axin-GSK3β-phospho-β-catenin complex is necessary for the K63-polyubiquitylation of APC. Intriguingly, the degree of this modification on APC appears to correlate inversely with the levels of β-catenin in cells. Together, our results indicate that K63-linked polyubiquitin adducts on APC regulate the assembly and/or efficiency of the β-catenin destruction complex. PMID:22761442

  5. CD8 T cell memory recall is enhanced by novel direct interactions with CD4 T cells enabled by MHC class II transferred from APCs.

    PubMed

    Romagnoli, Pablo A; Premenko-Lanier, Mary F; Loria, Gilbert D; Altman, John D

    2013-01-01

    Protection against many intracellular pathogens is provided by CD8 T cells, which are thought to need CD4 T cell help to develop into effective memory CD8 T cells. Because murine CD8 T cells do not transcribe MHC class II (MHC-II) genes, several models have proposed antigen presenting cells (APCs) as intermediaries required for CD4 T cells to deliver their help to CD8 T cells. Here, we demonstrate the presence of MHC-II molecules on activated murine CD8 T cells in vitro as well as in vivo. These MHC-II molecules are acquired via trogocytosis by CD8 T cells from their activating APCs, particularly CD11c positive dendritic cells (DCs). Transferred MHC-II molecules on activated murine CD8 T cells were functionally competent in stimulating specific indicator CD4 T cells. CD8 T cells that were "helped" in vitro and subsequently allowed to rest in vivo showed enhanced recall responses upon challenge compared to "helpless" CD8 T cells; in contrast, no differences were seen upon immediate challenge. These data indicate that direct CD8:CD4 T cell interactions may significantly contribute to help for CD8 T cells. Furthermore, this mechanism may enable CD8 T cells to communicate with different subsets of interacting CD4 T cells that could modulate immune responses. PMID:23441229

  6. Geopolymers prepared from DC plasma treated air pollution control (APC) residues glass: properties and characterisation of the binder phase.

    PubMed

    Kourti, Ioanna; Devaraj, Amutha Rani; Bustos, Ana Guerrero; Deegan, David; Boccaccini, Aldo R; Cheeseman, Christopher R

    2011-11-30

    Air pollution control (APC) residues have been blended with glass-forming additives and treated using DC plasma technology to produce a high calcium aluminosilicate glass (APC glass). This has been used to form geopolymer-glass composites that exhibit high strength and density, low porosity, low water absorption, low leaching and high acid resistance. The composites have a microstructure consisting of un-reacted residual APC glass particles imbedded in a complex geopolymer and C-S-H gel binder phase, and behave as particle reinforced composites. The work demonstrates that materials prepared from DC plasma treated APC residues have potential to be used to form high quality pre-cast products. PMID:21963174

  7. APC/C-Mediated Degradation of dsRNA-Binding Protein 4 (DRB4) Involved in RNA Silencing

    PubMed Central

    Marrocco, Katia; Criqui, Marie-Claire; Zervudacki, Jérôme; Schott, Gregory; Eisler, Herfried; Parnet, Aude; Dunoyer, Patrice; Genschik, Pascal

    2012-01-01

    Background Selective protein degradation via the ubiquitin-26S proteasome is a major mechanism underlying DNA replication and cell division in all Eukaryotes. In particular, the APC/C (Anaphase Promoting Complex or Cyclosome) is a master ubiquitin protein ligase (E3) that targets regulatory proteins for degradation allowing sister chromatid separation and exit from mitosis. Interestingly, recent work also indicates that the APC/C remains active in differentiated animal and plant cells. However, its role in post-mitotic cells remains elusive and only a few substrates have been characterized. Methodology/Principal Findings In order to identify novel APC/C substrates, we performed a yeast two-hybrid screen using as the bait Arabidopsis APC10/DOC1, one core subunit of the APC/C, which is required for substrate recruitment. This screen identified DRB4, a double-stranded RNA binding protein involved in the biogenesis of different classes of small RNA (sRNA). This protein interaction was further confirmed in vitro and in plant cells. Moreover, APC10 interacts with DRB4 through the second dsRNA binding motif (dsRBD2) of DRB4, which is also required for its homodimerization and binding to its Dicer partner DCL4. We further showed that DRB4 protein accumulates when the proteasome is inactivated and, most importantly, we found that DRB4 stability depends on APC/C activity. Hence, depletion of Arabidopsis APC/C activity by RNAi leads to a strong accumulation of endogenous DRB4, far beyond its normal level of accumulation. However, we could not detect any defects in sRNA production in lines where DRB4 was overexpressed. Conclusions/Significance Our work identified a first plant substrate of the APC/C, which is not a regulator of the cell cycle. Though we cannot exclude that APC/C-dependent degradation of DRB4 has some regulatory roles under specific growth conditions, our work rather points to a housekeeping function of APC/C in maintaining precise cellular

  8. Dual RING E3 Architectures Regulate Multiubiquitination and Ubiquitin Chain Elongation by APC/C.

    PubMed

    Brown, Nicholas G; VanderLinden, Ryan; Watson, Edmond R; Weissmann, Florian; Ordureau, Alban; Wu, Kuen-Phon; Zhang, Wei; Yu, Shanshan; Mercredi, Peter Y; Harrison, Joseph S; Davidson, Iain F; Qiao, Renping; Lu, Ying; Dube, Prakash; Brunner, Michael R; Grace, Christy R R; Miller, Darcie J; Haselbach, David; Jarvis, Marc A; Yamaguchi, Masaya; Yanishevski, David; Petzold, Georg; Sidhu, Sachdev S; Kuhlman, Brian; Kirschner, Marc W; Harper, J Wade; Peters, Jan-Michael; Stark, Holger; Schulman, Brenda A

    2016-06-01

    Protein ubiquitination involves E1, E2, and E3 trienzyme cascades. E2 and RING E3 enzymes often collaborate to first prime a substrate with a single ubiquitin (UB) and then achieve different forms of polyubiquitination: multiubiquitination of several sites and elongation of linkage-specific UB chains. Here, cryo-EM and biochemistry show that the human E3 anaphase-promoting complex/cyclosome (APC/C) and its two partner E2s, UBE2C (aka UBCH10) and UBE2S, adopt specialized catalytic architectures for these two distinct forms of polyubiquitination. The APC/C RING constrains UBE2C proximal to a substrate and simultaneously binds a substrate-linked UB to drive processive multiubiquitination. Alternatively, during UB chain elongation, the RING does not bind UBE2S but rather lures an evolving substrate-linked UB to UBE2S positioned through a cullin interaction to generate a Lys11-linked chain. Our findings define mechanisms of APC/C regulation, and establish principles by which specialized E3-E2-substrate-UB architectures control different forms of polyubiquitination. PMID:27259151

  9. The Specificity of Targeted Vaccines for APC Surface Molecules Influences the Immune Response Phenotype

    PubMed Central

    Grødeland, Gunnveig; Mjaaland, Siri; Tunheim, Gro; Fredriksen, Agnete B.; Bogen, Bjarne

    2013-01-01

    Different diseases require different immune responses for efficient protection. Thus, prophylactic vaccines should prime the immune system for the particular type of response needed for protection against a given infectious agent. We have here tested fusion DNA vaccines which encode proteins that bivalently target influenza hemagglutinins (HA) to different surface molecules on antigen presenting cells (APC). We demonstrate that targeting to MHC class II molecules predominantly induced an antibody/Th2 response, whereas targeting to CCR1/3/5 predominantly induced a CD8+/Th1 T cell response. With respect to antibodies, the polarizing effect was even more pronounced upon intramuscular (i.m) delivery as compared to intradermal (i.d.) vaccination. Despite these differences in induced immune responses, both vaccines protected against a viral challenge with influenza H1N1. Substitution of HA with ovalbumin (OVA) demonstrated that polarization of immune responses, as a consequence of APC targeting specificity, could be extended to other antigens. Taken together, the results demonstrate that vaccination can be tailor-made to induce a particular phenotype of adaptive immune responses by specifically targeting different surface molecules on APCs. PMID:24244595

  10. Cdh1-APC/C, cyclin B-Cdc2, and Alzheimer's disease pathology

    SciTech Connect

    Aulia, Selina; Tang, Bor Luen . E-mail: bchtbl@nus.edu.sg

    2006-01-06

    The anaphase-promoting complex/cyclosome (APC/C) is a key E3 ubiquitin ligase complex that functions in regulating cell cycle transitions in proliferating cells and has, as revealed recently, novel roles in postmitotic neurons. Regulated by its activator Cdh1 (or Hct1), whose level is high in postmitotic neurons, APC/C seems to have multiple functions at different cellular locations, modulating diverse processes such as synaptic development and axonal growth. These processes do not, however, appear to be directly connected to cell cycle regulation. It is now shown that Cdh1-APC/C activity may also have a basic role in suppressing cyclin B levels, thus preventing terminally differentiated neurons from aberrantly re-entering the cell cycle. The result of an aberrant cyclin B-induced S-phase entry, at least for some of these neurons, would be death via apoptosis. Cdh1 thus play an active role in maintaining the terminally differentiated, non-cycling state of postmitotic neurons-a function that could become impaired in Alzheimer's and other neurodegenerative diseases.

  11. Increased IκBα expression is essential for the tolerogenic property of TGF-β-exposed APCs

    PubMed Central

    Ghafoori, Paiman; Yoshimura, Takeru; Turpie, Bruce; Masli, Sharmila

    2009-01-01

    IκBα is an inhibitor of the transcriptional factor NF-κB, and it is an essential component of the signaling pathways that lead to expression of inflammatory molecules. These include cytokines and costimulatory molecules associated with antigen presentation in an inflammatory immune response. In this study, we report that antigen-presenting cells exposed to TGF-β induce peripheral tolerance by increasing IκBα expression. Exposure of antigen presenting cells (APCs) to TGF-β is known to impair their ability to secrete IL-12, and such impairment correlated with reduced NF-κB activity as indicated by significantly reduced nuclear levels of p50, an essential subunit of NF-κB for IL-12 transcription. Blockade of increased nuclear IκBα in APCs by expression of small interfering RNA molecules (siRNAs) targeting IκBα transcripts prevented IL-12 impairment and the decline in nuclear p50 levels. Furthermore, such IκBα blockade also interfered with the tolerogenic property of TGF-β- exposed APCs. However, increased expression of IκBα in APCs, independent of TGF-β exposure, reduced nuclear p50 levels and permitted tolerance induction by APCs. Thus, our findings attribute a direct and significant role to IκBα in the tolerogenic potential of APCs. Increased IκBα expression in APCs may therefore offer a therapeutic approach to achieve antigen-specific immunomodulation.—Ghafoori, P., Yoshimura, T., Turpie, B., Masli, S. Increased IκBα expression is essential for the tolerogenic property of TGF-β-exposed APCs. PMID:19237504

  12. Sesamol suppresses cyclooxygenase-2 transcriptional activity in colon cancer cells and modifies intestinal polyp development in ApcMin/+ mice

    PubMed Central

    Shimizu, Satomi; Fujii, Gen; Takahashi, Mami; Nakanishi, Ruri; Komiya, Masami; Shimura, Misato; Noma, Nobuharu; Onuma, Wakana; Terasaki, Masaru; Yano, Tomohiro; Mutoh, Michihiro

    2014-01-01

    Excessive prostaglandin production by cyclooxygenase-2 in stromal and epithelial cells is a causative factor of colorectal carcinogenesis. Thus, compounds which inhibit cyclooxygenase-2 transcriptional activity in colon epithelial cells could be candidates for anti-carcinogenic agents. A cyclooxygenase-2 transcriptional activity in the human colon cancer cell line DLD-1 has been measured using a β-galactosidase reporter gene system. Using this system, we demonstrated that the decrease in basal cyclooxygenase-2 transcriptional activities at 100 µM sesamol, one of the lignans in sesame seeds, was 50%. Other compounds in sesame seeds such as sesamin, sesamolin, ferulic acid, and syringic acid did not exhibit significant suppression of cyclooxygenase-2 transcriptional activity at up to 100 µM. In a following experiment, 6-week-old male Min mice, Apc-deficient mice, were divided into a non-treated and 500 ppm sesamol groups. At the age of 15 weeks, it was found that treatment with sesamol decreased the number of polyps in the middle part of small intestine to 66.1% of the untreated value. Moreover, sesamol suppressed cyclooxygenase-2 and cytosolic prostaglandin E2 synthase mRNA in the polyp parts. The present findings may demonstrate the novel anti-carcinogenetic property of sesamol, and imply that agents that can suppress cyclooxygenase-2 expression may be useful cancer chemopreventive agents. PMID:24688218

  13. Hypermethylation of hMLH1, HPP1, p14(ARF), p16(INK4A) and APC in primary adenocarcinomas of the small bowel.

    PubMed

    Brücher, Björn L D M; Geddert, Helene; Langner, Cord; Höfler, Heinz; Fink, Ulrich; Siewert, Jörg R; Sarbia, Mario

    2006-09-15

    Small bowel adenocarcinoma (SB-AC) is a very rare tumor entity. Epigenetic alterations, including hypermethylation of DNA mismatch repair genes and tumor suppressor genes, seem to be important for carcinogenesis in tumors of the gastrointestinal tract, but have not yet been investigated in SB-AC. In the current study, the prevalence of hypermethylation in a panel of genes involved in gastrointestinal carcinogenesis (hMLH1, HPP1, p14(ARF), p16(INK4A), APC) was determined in a series of SB-AC. Paraffin-embedded tumor samples from 56 patients with SB-AC who underwent surgical resection between January 1985 and December 2003 were investigated for hypermethylation by means of methylation-specific real-time PCR, and compared with our findings in a previously investigated series of 50 gastric adenocarcinomas. In comparison with adenocarcinomas of the stomach, SB-AC revealed a significantly higher rate of hypermethylation of HPP1 (86% versus 54%, p = 0.0003), p16(INK4A) (32% versus 10%, p = 0.0006), and a significantly lower rate of hypermethylation of APC (48% versus 84%, p = 0.0001). Hypermethylation of hMLH1 and p14(ARF) was present in 23% and 9% of SB-AC, respectively. Locally advanced tumor categories (pT3/4) showed a higher rate of hypermethylation of HPP1 (90%) than did early tumor categories (pT1/2 categories, 40%; p = 0.0036). This was also reflected by the correlation between the HPP1 hypermethylation and high UICC stage (p = 0.02). No correlation was found between hypermethylation and other clinicopathologic parameters such as age, tumor grade and nodal status. Our findings suggest that hypermethylation of hMLH1, HPP1, p16(INK4A) and APC is frequent in primary adenocarcinomas of the small bowel. The differences in the hypermethylation spectrum of small bowel and stomach cancer indicate significant epigenetic differences between these tumors. PMID:16619216

  14. APC is an RNA-Binding Protein and its Interactome Provides a Link to Neural Development and Microtubule Assembly

    PubMed Central

    Preitner, Nicolas; Quan, Jie; Nowakowski, Dan W.; Hancock, Melissa L.; Shi, Jianhua; Tcherkezian, Joseph; Young-Pearse, Tracy L.; Flanagan, John G.

    2014-01-01

    SUMMARY Adenomatous polyposis coli (APC) is a microtubule plus-end scaffolding protein important in biology and disease. APC is implicated in RNA localization, although the mechanisms and functional significance remain unclear. We show that APC is an RNA-binding protein, and identify an RNA interactome by HITS-CLIP. Targets were highly enriched for APC-related functions, including microtubule organization, cell motility, cancer and neurologic disease. Among the targets is β2B-tubulin, known to be required in human neuron and axon migration. We show β2B-tubulin is synthesized in axons and localizes preferentially to dynamic microtubules in the growth cone periphery. APC binds the β2B-tubulin 3'UTR; treatments interfering with this interaction reduced β2B-tubulin mRNA axonal localization and expression, depleted dynamic microtubules and the growth cone periphery, and impaired neuron migration. These results identify APC as a platform binding functionally-related protein and RNA networks, and suggest a self-organizing model for the microtubule to localize synthesis of its own subunits. PMID:25036633

  15. Association between aberrant APC promoter methylation and breast cancer pathogenesis: a meta-analysis of 35 observational studies.

    PubMed

    Zhou, Dan; Tang, Weiwei; Wang, Wenyi; Pan, Xiaoyan; An, Han-Xiang; Zhang, Yun

    2016-01-01

    Background. Adenomatous polyposis coli (APC) is widely known as an antagonist of the Wnt signaling pathway via the inactivation of β-catenin. An increasing number of studies have reported that APC methylation contributes to the predisposition to breast cancer (BC). However, recent studies have yielded conflicting results. Methods. Herein, we systematically carried out a meta-analysis to assess the correlation between APC methylation and BC risk. Based on searches of the Cochrane Library, PubMed, Web of Science and Embase databases, the odds ratio (OR) with 95% confidence interval (CI) values were pooled and summarized. Results. A total of 31 articles involving 35 observational studies with 2,483 cases and 1,218 controls met the inclusion criteria. The results demonstrated that the frequency of APC methylation was significantly higher in BC cases than controls under a random effect model (OR = 8.92, 95% CI [5.12-15.52]). Subgroup analysis further confirmed the reliable results, regardless of the sample types detected, methylation detection methods applied and different regions included. Interestingly, our results also showed that the frequency of APC methylation was significantly lower in early-stage BC patients than late-stage ones (OR = 0.62, 95% CI [0.42-0.93]). Conclusion. APC methylation might play an indispensable role in the pathogenesis of BC and could be regarded as a potential biomarker for the diagnosis of BC. PMID:27478702

  16. Interphase APC/C–Cdc20 inhibition by cyclin A2–Cdk2 ensures efficient mitotic entry

    PubMed Central

    Hein, Jamin B.; Nilsson, Jakob

    2016-01-01

    Proper cell-cycle progression requires tight temporal control of the Anaphase Promoting Complex/Cyclosome (APC/C), a large ubiquitin ligase that is activated by one of two co-activators, Cdh1 or Cdc20. APC/C and Cdc20 are already present during interphase but APC/C–Cdc20 regulation during this window of the cell cycle, if any, is unknown. Here we show that cyclin A2–Cdk2 binds and phosphorylates Cdc20 in interphase and this inhibits APC/C–Cdc20 activity. Preventing Cdc20 phosphorylation results in pre-mature activation of the APC/C–Cdc20 and several substrates, including cyclin B1 and A2, are destabilized which lengthens G2 and slows mitotic entry. Expressing non-degradable cyclin A2 but not cyclin B1 restores mitotic entry in these cells. We have thus uncovered a novel positive feedback loop centred on cyclin A2–Cdk2 inhibition of interphase APC/C–Cdc20 to allow further cyclin A2 accumulation and mitotic entry. PMID:26960431

  17. The APC tumor suppressor binds to C-terminal binding protein to divert nuclear beta-catenin from TCF.

    PubMed

    Hamada, Fumihiko; Bienz, Mariann

    2004-11-01

    Adenomatous polyposis coli (APC) is an important tumor suppressor in the colon. APC antagonizes the transcriptional activity of the Wnt effector beta-catenin by promoting its nuclear export and its proteasomal destruction in the cytoplasm. Here, we show that a third function of APC in antagonizing beta-catenin involves C-terminal binding protein (CtBP). APC is associated with CtBP in vivo and binds to CtBP in vitro through its conserved 15 amino acid repeats. Failure of this association results in elevated levels of beta-catenin/TCF complexes and of TCF-mediated transcription. Notably, CtBP is neither associated with TCF in vivo nor does mutation of the CtBP binding motifs in TCF-4 alter its transcriptional activity. This questions the idea that CtBP is a direct corepressor of TCF. Our evidence indicates that APC is an adaptor between beta-catenin and CtBP and that CtBP lowers the availability of free nuclear beta-catenin for binding to TCF by sequestering APC/beta-catenin complexes. PMID:15525529

  18. Association between aberrant APC promoter methylation and breast cancer pathogenesis: a meta-analysis of 35 observational studies

    PubMed Central

    Zhou, Dan; Tang, Weiwei; Wang, Wenyi; Pan, Xiaoyan

    2016-01-01

    Background. Adenomatous polyposis coli (APC) is widely known as an antagonist of the Wnt signaling pathway via the inactivation of β-catenin. An increasing number of studies have reported that APC methylation contributes to the predisposition to breast cancer (BC). However, recent studies have yielded conflicting results. Methods. Herein, we systematically carried out a meta-analysis to assess the correlation between APC methylation and BC risk. Based on searches of the Cochrane Library, PubMed, Web of Science and Embase databases, the odds ratio (OR) with 95% confidence interval (CI) values were pooled and summarized. Results. A total of 31 articles involving 35 observational studies with 2,483 cases and 1,218 controls met the inclusion criteria. The results demonstrated that the frequency of APC methylation was significantly higher in BC cases than controls under a random effect model (OR = 8.92, 95% CI [5.12–15.52]). Subgroup analysis further confirmed the reliable results, regardless of the sample types detected, methylation detection methods applied and different regions included. Interestingly, our results also showed that the frequency of APC methylation was significantly lower in early-stage BC patients than late-stage ones (OR = 0.62, 95% CI [0.42–0.93]). Conclusion. APC methylation might play an indispensable role in the pathogenesis of BC and could be regarded as a potential biomarker for the diagnosis of BC. PMID:27478702

  19. The APC I1307K allele conveys a significant increased risk for cancer.

    PubMed

    Leshno, Ari; Shapira, Shiran; Liberman, Eliezer; Kraus, Sarah; Sror, Miri; Harlap-Gat, Amira; Avivi, Doran; Galazan, Lior; David, Maayan; Maharshak, Nitsan; Moanis, Serhan; Arber, Nadir; Moshkowitz, Menachem

    2016-03-15

    This study is the first attempt to evaluate the association between the APC I1307K variant and overall cancer risk. It is unique in both its large sample size and in the reliability of data in the control group. The findings described in this article have major implications in terms of identifying asymptomatic individuals who are at increased risk to harbor cancer and therefore targeted to be enrolled in specific early detection and prevention programs. The prevalence of the APC I1307K missense mutation among Ashkenazi Jews is ∼ 6%. Carriers are at an increased risk for colorectal neoplasia. In this study, we examined the association of this variant with non-colorectal cancers. Consecutive 13,013 healthy subjects who underwent screening at the Integrated Cancer Prevention Center between 2006 and 2014 were enrolled. This population was supplemented with 1,611 cancer patients from the same institution. Demographics, medical history, and pathological data were recorded. Mortality data were obtained from the Ministry of Health's registry. The prevalence of APC I1307K in cancer patients and healthy subjects was compared. The APC I1307K variant was detected in 189 (11.8%) cancer patients compared to 614 (4.7%) healthy subjects, reflecting an adjusted age and sex odds ratio (OR) of 2.53 (p < 0.0001). History of two or more cancer types was associated with a positive carrier prevalence (OR = 4.38 p < 0.0001). Males had significantly increased carrier prevalence in lung, urologic, pancreatic, and skin cancers. The carrier prevalence among females was significantly higher only in breast and skin cancers. Female carriers developed cancer at a significantly older age compared to non-carriers (average 62.7 years vs. 57.8, respectively, p = 0.027), had better survival rates (HR = 0.58, p = 0.022) and overall increased longevity (average age of death 78.8 vs. 70.4 years, respectively, p = 0.003). In conclusion, the APC I1307K variant is a reliable marker for overall cancer risk

  20. Drosophila Homologues of Adenomatous Polyposis Coli (APC) and the Formin Diaphanous Collaborate by a Conserved Mechanism to Stimulate Actin Filament Assembly*

    PubMed Central

    Jaiswal, Richa; Stepanik, Vince; Rankova, Aneliya; Molinar, Olivia; Goode, Bruce L.; McCartney, Brooke M.

    2013-01-01

    Adenomatous polyposis coli (APC) is a large multidomain protein that regulates the cytoskeleton. Recently, it was shown that vertebrate APC through its Basic domain directly collaborates with the formin mDia1 to stimulate actin filament assembly in the presence of nucleation barriers. However, it has been unclear whether these activities extend to homologues of APC and Dia in other organisms. Drosophila APC and Dia are each required to promote actin furrow formation in the syncytial embryo, suggesting a potential collaboration in actin assembly, but low sequence homology between the Basic domains of Drosophila and vertebrate APC has left their functional and mechanistic parallels uncertain. To address this question, we purified Drosophila APC1 and Dia and determined their individual and combined effects on actin assembly using both bulk fluorescence assays and total internal reflection fluorescence microscopy. Our data show that APC1, similar to its vertebrate homologue, bound to actin monomers and nucleated and bundled filaments. Further, Drosophila Dia nucleated actin assembly and protected growing filament barbed ends from capping protein. Drosophila APC1 and Dia directly interacted and collaborated to promote actin assembly in the combined presence of profilin and capping protein. Thus, despite limited sequence homology, Drosophila and vertebrate APCs exhibit highly related activities and mechanisms and directly collaborate with formins. These results suggest that APC-Dia interactions in actin assembly are conserved and may underlie important in vivo functions in a broad range of animal phyla. PMID:23558679

  1. Study of Cyclin Proteolysis in Anaphase-Promoting Complex (APC) Mutant Cells Reveals the Requirement for APC Function in the Final Steps of the Fission Yeast Septation Initiation Network

    PubMed Central

    Chang, Louise; Morrell, Jennifer L.; Feoktistova, Anna; Gould, Kathleen L.

    2001-01-01

    Cytokinesis in eukaryotic cells requires the inactivation of mitotic cyclin-dependent kinase complexes. An apparent exception to this relationship is found in Schizosaccharomyces pombe mutants with mutations of the anaphase-promoting complex (APC). These conditional lethal mutants arrest with unsegregated chromosomes because they cannot degrade the securin, Cut2p. Although failing at nuclear division, these mutants septate and divide. Since septation requires Cdc2p inactivation in wild-type S. pombe, it has been suggested that Cdc2p inactivation occurs in these mutants by a mechanism independent of cyclin degradation. In contrast to this prediction, we show that Cdc2p kinase activity fluctuates in APC cut mutants due to Cdc13/cyclin B destruction. In APC-null mutants, however, septation and cutting do not occur and Cdc13p is stable. We conclude that APC cut mutants are hypomorphic with respect to Cdc13p degradation. Indeed, overproduction of nondestructible Cdc13p prevents septation in APC cut mutants and the normal reorganization of septation initiation network components during anaphase. PMID:11533255

  2. The Four Canonical TPR Subunits of Human APC/C Form Related Homo-Dimeric Structures and Stack in Parallel to Form a TPR Suprahelix☆

    PubMed Central

    Zhang, Ziguo; Chang, Leifu; Yang, Jing; Conin, Nora; Kulkarni, Kiran; Barford, David

    2013-01-01

    The anaphase-promoting complex or cyclosome (APC/C) is a large E3 RING-cullin ubiquitin ligase composed of between 14 and 15 individual proteins. A striking feature of the APC/C is that only four proteins are involved in directly recognizing target proteins and catalyzing the assembly of a polyubiquitin chain. All other subunits, which account for > 80% of the mass of the APC/C, provide scaffolding functions. A major proportion of these scaffolding subunits are structurally related. In metazoans, there are four canonical tetratricopeptide repeat (TPR) proteins that form homo-dimers (Apc3/Cdc27, Apc6/Cdc16, Apc7 and Apc8/Cdc23). Here, we describe the crystal structure of the N-terminal homo-dimerization domain of Schizosaccharomyces pombe Cdc23 (Cdc23Nterm). Cdc23Nterm is composed of seven contiguous TPR motifs that self-associate through a related mechanism to those of Cdc16 and Cdc27. Using the Cdc23Nterm structure, we generated a model of full-length Cdc23. The resultant “V”-shaped molecule docks into the Cdc23-assigned density of the human APC/C structure determined using negative stain electron microscopy (EM). Based on sequence conservation, we propose that Apc7 forms a homo-dimeric structure equivalent to those of Cdc16, Cdc23 and Cdc27. The model is consistent with the Apc7-assigned density of the human APC/C EM structure. The four canonical homo-dimeric TPR proteins of human APC/C stack in parallel on one side of the complex. Remarkably, the uniform relative packing of neighboring TPR proteins generates a novel left-handed suprahelical TPR assembly. This finding has implications for understanding the assembly of other TPR-containing multimeric complexes. PMID:23583778

  3. Kinesin-2 and Apc function at dendrite branch points to resolve microtubule collisions.

    PubMed

    Weiner, Alexis T; Lanz, Michael C; Goetschius, Daniel J; Hancock, William O; Rolls, Melissa M

    2016-01-01

    In Drosophila neurons, kinesin-2, EB1 and Apc are required to maintain minus-end-out dendrite microtubule polarity, and we previously proposed they steer microtubules at branch points. Motor-mediated steering of microtubule plus ends could be accomplished in two ways: 1) by linking a growing microtubule tip to the side of an adjacent microtubule as it navigates the branch point (bundling), or 2) by directing a growing microtubule after a collision with a stable microtubule (collision resolution). Using live imaging to distinguish between these two mechanisms, we found that reduction of kinesin-2 did not alter the number of microtubules that grew along the edge of the branch points where stable microtubules are found. However, reduction of kinesin-2 or Apc did affect the number of microtubules that slowed down or depolymerized as they encountered the side of the branch opposite to the entry point. These results are consistent with kinesin-2 functioning with Apc to resolve collisions. However, they do not pinpoint stable microtubules as the collision partner as stable microtubules are typically very close to the membrane. To determine whether growing microtubules were steered along stable ones after a collision, we analyzed the behavior of growing microtubules at dendrite crossroads where stable microtubules run through the middle of the branch point. In control neurons, microtubules turned in the middle of the crossroads. However, when kinesin-2 was reduced some microtubules grew straight through the branch point and failed to turn. We propose that kinesin-2 functions to steer growing microtubules along stable ones following collisions. PMID:26785384

  4. PLK1 regulates spindle formation kinetics and APC/C activation in mouse zygote.

    PubMed

    Baran, Vladimir; Brzakova, Adela; Rehak, Pavol; Kovarikova, Veronika; Solc, Petr

    2016-06-01

    Polo-like kinase 1 (PLK1) is involved in essential events of cell cycle including mitosis in which it participates in centrosomal microtubule nucleation, spindle bipolarity establishment and cytokinesis. Although PLK1 function has been studied in cycling cancer cells, only limited data are known about its role in the first mitosis of mammalian zygotes. During the 1-cell stage of mouse embryo development, the acentriolar spindle is formed and the shift from acentriolar to centrosomal spindle formation progresses gradually throughout the preimplantation stage, thus providing a unique possibility to study acentriolar spindle formation. We have shown previously that PLK1 activity is not essential for entry into first mitosis, but is required for correct spindle formation and anaphase onset in 1-cell mouse embryos. In the present study, we extend this knowledge by employing quantitative confocal live cell imaging to determine spindle formation kinetics in the absence of PLK1 activity and answer the question whether metaphase arrest at PLK1-inhibited embryos is associated with low anaphase-promoting complex/cyclosome (APC/C) activity and consequently high securin level. We have shown that inhibition of PLK1 activity induces a delay in onset of acentriolar spindle formation during first mitosis. Although these PLK1-inhibited 1-cell embryos were finally able to form a bipolar spindle, not all chromosomes were aligned at the metaphase equator. PLK1-inhibited embryos were arrested in metaphase without any sign of APC/C activation with high securin levels. Our results document that PLK1 controls the onset of spindle assembly and spindle formation, and is essential for APC/C activation before anaphase onset in mouse zygotes. PMID:26174739

  5. STS-34 crewmembers sit in M1-13 APC during emergency egress training at KSC

    NASA Technical Reports Server (NTRS)

    1989-01-01

    STS-34 crewmembers sit in M1-13 Armored Personnel Carrier (APC) during emergency egress training at KSC's shuttle landing facility (SLF) prior to terminal countdown demonstration test (TCDT) activities. Wearing launch and entry suits (LESs), are (from left) Mission Specialist (MS) Ellen S. Baker, MS Shannon W. Lucid, Commander Donald E. Williams (right side, in back), MS Franklin R. Chang-Diaz, and Pilot Michael J. McCulley (holding headset). View provided by KSC with alternate number KSC-89PC-871.

  6. The ubiquitin ligase APC/CCdh1 puts the brakes on DNA-end resection

    PubMed Central

    Lafranchi, Lorenzo; Sartori, Alessandro A

    2015-01-01

    DNA double-strand breaks (DSBs) are highly deleterious lesions and their misrepair can promote genomic instability, a hallmark of cancer. DNA-end resection is a cell cycle-regulated mechanism that is required for the faithful repair of DSBs. We recently discovered that the anaphase-promoting complex/cyclosome-Cdh1 (APC/CCdh1) ubiquitin ligase is responsible for the timely degradation of CtBP-interacting protein (CtIP), a key DNA-end resection factor, providing a new layer of regulation of DSB repair in human cells. PMID:27308488

  7. Intestinal APCs of the endogenous nanomineral pathway fail to express PD-L1 in Crohn’s disease

    PubMed Central

    Robertson, Jack; Haas, Carolin T.; Pele, Laetitia C.; Monie, Tom P.; Charalambos, Charles; Parkes, Miles; Hewitt, Rachel E.; Powell, Jonathan J.

    2016-01-01

    Crohn’s disease is a chronic inflammatory condition most commonly affecting the ileum and colon. The aetiology of Crohn’s disease is complex and may include defects in peptidoglycan recognition, and/or failures in the establishment of intestinal tolerance. We have recently described a novel constitutive endogenous delivery system for the translocation of nanomineral-antigen-peptidoglycan (NAP) conjugates to antigen presenting cells (APCs) in intestinal lymphoid patches. In mice NAP conjugate delivery to APCs results in high surface expression of the immuno-modulatory molecule programmed death receptor ligand 1 (PD-L1). Here we report that NAP conjugate positive APCs in human ileal tissues from individuals with ulcerative colitis and intestinal carcinomas, also have high expression of PD-L1. However, NAP-conjugate positive APCs in intestinal tissue from patients with Crohn’s disease show selective failure in PD-L1 expression. Therefore, in Crohn’s disease intestinal antigen taken up by lymphoid patch APCs will be presented without PD-L1 induced tolerogenic signalling, perhaps initiating disease. PMID:27226337

  8. Aggregate material formulated with MSWI bottom ash and APC fly ash for use as secondary building material

    SciTech Connect

    Valle-Zermeño, R. del; Formosa, J.; Chimenos, J.M.; Martínez, M.; Fernández, A.I.

    2013-03-15

    Highlights: ► A concrete formulation was optimized using Bottom Ash and APC ash. ► 10% of APC ash achieves good compromise between economic and performance aspects. ► The crushed concrete was evaluated as secondary building granular material. ► The environmental behavior allows its use as secondary material. ► The abrasion resistance is not good enough for its use as a road sub-base material. - Abstract: The main goal of this paper is to obtain a granular material formulated with Municipal Solid Waste Incineration (MSWI) bottom ash (BA) and air pollution control (APC) fly ash to be used as secondary building material. Previously, an optimum concrete mixture using both MSWI residues as aggregates was formulated. A compromise between the environmental behavior whilst maximizing the reuse of APC fly ash was considered and assessed. Unconfined compressive strength and abrasion resistance values were measured in order to evaluate the mechanical properties. From these results, the granular mixture was not suited for certain applications owing to the high BA/APC fly ash content and low cement percentages used to reduce the costs of the final product. Nevertheless, the leaching test performed showed that the concentrations of all heavy metals were below the limits established by the current Catalan legislation for their reutilization. Therefore, the material studied might be mainly used in embankments, where high mechanical properties are not needed and environmental safety is assured.

  9. Irreversible APC(Cdh1) Inactivation Underlies the Point of No Return for Cell-Cycle Entry.

    PubMed

    Cappell, Steven D; Chung, Mingyu; Jaimovich, Ariel; Spencer, Sabrina L; Meyer, Tobias

    2016-06-30

    Proliferating cells must cross a point of no return before they replicate their DNA and divide. This commitment decision plays a fundamental role in cancer and degenerative diseases and has been proposed to be mediated by phosphorylation of retinoblastoma (Rb) protein. Here, we show that inactivation of the anaphase-promoting complex/cyclosome (APC(Cdh1)) has the necessary characteristics to be the point of no return for cell-cycle entry. Our study shows that APC(Cdh1) inactivation is a rapid, bistable switch initiated shortly before the start of DNA replication by cyclin E/Cdk2 and made irreversible by Emi1. Exposure to stress between Rb phosphorylation and APC(Cdh1) inactivation, but not after APC(Cdh1) inactivation, reverted cells to a mitogen-sensitive quiescent state, from which they can later re-enter the cell cycle. Thus, APC(Cdh1) inactivation is the commitment point when cells lose the ability to return to quiescence and decide to progress through the cell cycle. PMID:27368103

  10. New origin firing is inhibited by APC/CCdh1 activation in S-phase after severe replication stress

    PubMed Central

    Ercilla, Amaia; Llopis, Alba; Feu, Sonia; Aranda, Sergi; Ernfors, Patrik; Freire, Raimundo; Agell, Neus

    2016-01-01

    Defects in DNA replication and repair are known to promote genomic instability, a hallmark of cancer cells. Thus, eukaryotic cells have developed complex mechanisms to ensure accurate duplication of their genomes. While DNA damage response has been extensively studied in tumour cells, the pathways implicated in the response to replication stress are less well understood especially in non-transformed cells. Here we show that in non-transformed cells, APC/CCdh1 is activated upon severe replication stress. Activation of APC/CCdh1 prevents new origin firing and induces permanent arrest in S-phase. Moreover, Rad51-mediated homologous recombination is also impaired under these conditions. APC/CCdh1 activation in S-phase occurs after replication forks have been processed into double strand breaks. Remarkably, this activation, which correlates with decreased Emi1 levels, is not prevented by ATR/ATM inhibition, but it is abrogated in cells depleted of p53 or p21. Importantly, we found that the lack of APC/CCdh1 activity correlated with an increase in genomic instability. Taken together, our results define a new APC/CCdh1 function that prevents cell cycle resumption after prolonged replication stress by inhibiting origin firing, which may act as an additional mechanism in safeguarding genome integrity. PMID:26939887

  11. The ubiquitin ligase APC(Cdh1) is required to maintain genome integrity in primary human cells.

    PubMed

    Engelbert, D; Schnerch, D; Baumgarten, A; Wäsch, R

    2008-02-01

    Ensuring precise DNA replication and chromosome segregation is essential during cell division in order to provide genomic stability and avoid malignant growth. Proteolytic control of cell cycle regulators by the anaphase-promoting complex, activated by Cdh1 (APC(Cdh1)), is responsible for a stable G1 phase after mitotic exit allowing accurate preparation for DNA replication in the following S phase. APC(Cdh1) target proteins are frequently upregulated in tumor cells and the inactivation of human Cdh1 might interfere with genome integrity by target stabilization. Here we show that APC(Cdh1) is required for maintaining genomic integrity in primary human cells. Lentiviral-delivered strong and stable suppression of Cdh1 by RNA interference (RNAi) causes aberrant accumulation of several APC(Cdh1) target proteins, such as cyclin A, B, Aurora A or Plk1, which control accurate and equal distribution of the genetic information to daughter cells. This induces a premature and prolonged S phase, mitotic-entry delay and defects in chromosome separation and cytokinesis. Cell cycle deregulation by stable knockdown of Cdh1 leads to activation of p53/p21 and genomic instability, which is further increased by codepletion of p53. Thus, stabilization of APC(Cdh1) targets may initiate aberrant DNA replication and chromosome separation, and trigger a p53 response by deregulating G1 in primary human cells. PMID:17700535

  12. The Cdk1-APC/C cell cycle oscillator circuit functions as a time-delayed, ultrasensitive switch

    PubMed Central

    Yang, Qiong; Ferrell, James E.

    2013-01-01

    Despite the complexity and variety of biological oscillators, their core design invariably includes an essential negative feedback loop. In the Xenopus laevis embryonic cell cycle oscillator, this loop consists of the kinase cyclin B-Cdk1 and the ubiquitin ligase APC/CCdc20; active Cdk1 activates APC/CCdc20, which then brings about cyclin B degradation and inactivates Cdk1. Here we ask how this negative feedback loop functions quantitatively, with the aim of understanding what mechanisms keep the Cdk1-APC/CCdc20 system from settling into a stable steady state with intermediate levels of Cdk1 and APC/CCdc20 activity. We found that the system operates as a time-delayed, digital switch, with a time lag of ~15 min between Cdk1 and APC/CCdc20 activation and a tremendously high degree of ultrasensitivity (nH ≈ 17). Computational modeling shows how these attributes contribute to the generation of robust, clock-like oscillations. Principles uncovered here may also apply to other activator-repressor oscillators and help in designing robust synthetic clocks. PMID:23624406

  13. New origin firing is inhibited by APC/CCdh1 activation in S-phase after severe replication stress.

    PubMed

    Ercilla, Amaia; Llopis, Alba; Feu, Sonia; Aranda, Sergi; Ernfors, Patrik; Freire, Raimundo; Agell, Neus

    2016-06-01

    Defects in DNA replication and repair are known to promote genomic instability, a hallmark of cancer cells. Thus, eukaryotic cells have developed complex mechanisms to ensure accurate duplication of their genomes. While DNA damage response has been extensively studied in tumour cells, the pathways implicated in the response to replication stress are less well understood especially in non-transformed cells. Here we show that in non-transformed cells, APC/C(Cdh1) is activated upon severe replication stress. Activation of APC/C(Cdh1) prevents new origin firing and induces permanent arrest in S-phase. Moreover, Rad51-mediated homologous recombination is also impaired under these conditions. APC/C(Cdh1) activation in S-phase occurs after replication forks have been processed into double strand breaks. Remarkably, this activation, which correlates with decreased Emi1 levels, is not prevented by ATR/ATM inhibition, but it is abrogated in cells depleted of p53 or p21. Importantly, we found that the lack of APC/C(Cdh1) activity correlated with an increase in genomic instability. Taken together, our results define a new APC/C(Cdh1) function that prevents cell cycle resumption after prolonged replication stress by inhibiting origin firing, which may act as an additional mechanism in safeguarding genome integrity. PMID:26939887

  14. Intestinal APCs of the endogenous nanomineral pathway fail to express PD-L1 in Crohn's disease.

    PubMed

    Robertson, Jack; Haas, Carolin T; Pele, Laetitia C; Monie, Tom P; Charalambos, Charles; Parkes, Miles; Hewitt, Rachel E; Powell, Jonathan J

    2016-01-01

    Crohn's disease is a chronic inflammatory condition most commonly affecting the ileum and colon. The aetiology of Crohn's disease is complex and may include defects in peptidoglycan recognition, and/or failures in the establishment of intestinal tolerance. We have recently described a novel constitutive endogenous delivery system for the translocation of nanomineral-antigen-peptidoglycan (NAP) conjugates to antigen presenting cells (APCs) in intestinal lymphoid patches. In mice NAP conjugate delivery to APCs results in high surface expression of the immuno-modulatory molecule programmed death receptor ligand 1 (PD-L1). Here we report that NAP conjugate positive APCs in human ileal tissues from individuals with ulcerative colitis and intestinal carcinomas, also have high expression of PD-L1. However, NAP-conjugate positive APCs in intestinal tissue from patients with Crohn's disease show selective failure in PD-L1 expression. Therefore, in Crohn's disease intestinal antigen taken up by lymphoid patch APCs will be presented without PD-L1 induced tolerogenic signalling, perhaps initiating disease. PMID:27226337

  15. American ginseng significantly reduced the progression of high-fat-diet-enhanced colon carcinogenesis in ApcMin/+mice

    PubMed Central

    Yu, Chunhao; Wen, Xiao-Dong; Zhang, Zhiyu; Zhang, Chun-Feng; Wu, Xiaohui; He, Xin; Liao, Yang; Wu, Ningning; Wang, Chong-Zhi; Du, Wei; He, Tong-Chuan; Yuan, Chun-Su

    2015-01-01

    Background Colorectal cancer (CRC) is a leading cause of death worldwide. Chronic gut inflammation is recognized as a risk factor for tumor development, including CRC. American ginseng is a very commonly used ginseng species in the West. Methods A genetically engineered ApcMin/+ mouse model was used in this study. We analyzed the saponin composition of American ginseng used in this project, and evaluated its effects on the progression of high-fat-diet-enhanced CRC carcinogenesis. Results After oral ginseng administration (10–20 mg/kg/d for up to 32 wk), experimental data showed that, compared with the untreated mice, ginseng very significantly reduced tumor initiation and progression in both the small intestine (including the proximal end, middle end, and distal end) and the colon (all p < 0.01). This tumor number reduction was more obvious in those mice treated with a low dose of ginseng. The tumor multiplicity data were supported by body weight changes and gut tissue histology examinations. In addition, quantitative real-time polymerase chain reaction analysis showed that compared with the untreated group, ginseng very significantly reduced the gene expression of inflammatory cytokines, including interleukin-1α (IL-1α), IL-1β, IL-6, tumor necrosis factor-α, granulocyte-colony stimulating factor, and granulocyte-macrophage colony-stimulating factor in both the small intestine and the colon (all p < 0.01). Conclusion Further studies are needed to link our observed effects to the actions of the gut microbiome in converting the parent ginsenosides to bioactive ginseng metabolites. Our data suggest that American ginseng may have potential value in CRC chemoprevention. PMID:26199554

  16. Berberine Inhibits Intestinal Polyps Growth in Apc (min/+) Mice via Regulation of Macrophage Polarization.

    PubMed

    Piao, Meiyu; Cao, Hailong; He, NaNa; Yang, Boli; Dong, Wenxiao; Xu, Mengque; Yan, Fang; Zhou, Bing; Wang, Bangmao

    2016-01-01

    Antitumor effect of berberine has been reported in a wide spectrum of cancer, however, the mechanisms of which are not fully understood. The aim of this study was to investigate the hypothesis that berberine suppresses tumorigenesis in the familial adenomatous polyposis (FAP) by regulating the macrophage polarization in Apc (min/+) mouse model. Berberine was given to Apc (min/+) mice for 12 weeks. Primary macrophages were isolated; after berberine treatment, the change in signaling cascade was determined. The total number and size of polyps were reduced remarkably in berberine group, compared with control group. A significant decrease in protein levels of F4/80, mannose receptor (MR), and COX-2 in stroma of intestinal polyps and an increase in the level of iNOS were observed after berberine treatment. The mRNA level of MR and Arg-1 in berberine group was significantly lower than those in IL-10 or IL-4 group, while no significant difference in mRNA levels of iNOS and CXCL10 was observed. The migration and invasiveness assays in vitro showed that berberine could reduce the capability of migration and invasiveness. These findings suggest that berberine attenuates intestinal tumorigenesis by inhibiting the migration and invasion of colorectal tumor cells via regulation of macrophage polarization. PMID:27493671

  17. Increased variability in ApcMin/+ intestinal tissue can be measured with microultrasound

    NASA Astrophysics Data System (ADS)

    Fatehullah, A.; Sharma, S.; Newton, I. P.; Langlands, A. J.; Lay, H.; Nelson, S. A.; McMahon, R. K.; McIlvenny, N.; Appleton, P. L.; Cochran, S.; Näthke, I. S.

    2016-07-01

    Altered tissue structure is a feature of many disease states and is usually measured by microscopic methods, limiting analysis to small areas. Means to rapidly and quantitatively measure the structure and organisation of large tissue areas would represent a major advance not just for research but also in the clinic. Here, changes in tissue organisation that result from heterozygosity in Apc, a precancerous situation, are comprehensively measured using microultrasound and three-dimensional high-resolution microscopy. Despite its normal appearance in conventionally examined cross-sections, both approaches revealed a significant increase in the variability of tissue organisation in Apc heterozygous tissue. These changes preceded the formation of aberrant crypt foci or adenoma. Measuring these premalignant changes using microultrasound provides a potential means to detect microscopically abnormal regions in large tissue samples, independent of visual examination or biopsies. Not only does this provide a powerful tool for studying tissue structure in experimental settings, the ability to detect and monitor tissue changes by microultrasound could be developed into a powerful adjunct to screening endoscopy in the clinic.

  18. Increased variability in ApcMin/+ intestinal tissue can be measured with microultrasound

    PubMed Central

    Fatehullah, A.; Sharma, S.; Newton, I. P.; Langlands, A. J.; Lay, H.; Nelson, S. A.; McMahon, R. K.; McIlvenny, N.; Appleton, P. L.; Cochran, S.; Näthke, I. S.

    2016-01-01

    Altered tissue structure is a feature of many disease states and is usually measured by microscopic methods, limiting analysis to small areas. Means to rapidly and quantitatively measure the structure and organisation of large tissue areas would represent a major advance not just for research but also in the clinic. Here, changes in tissue organisation that result from heterozygosity in Apc, a precancerous situation, are comprehensively measured using microultrasound and three-dimensional high-resolution microscopy. Despite its normal appearance in conventionally examined cross-sections, both approaches revealed a significant increase in the variability of tissue organisation in Apc heterozygous tissue. These changes preceded the formation of aberrant crypt foci or adenoma. Measuring these premalignant changes using microultrasound provides a potential means to detect microscopically abnormal regions in large tissue samples, independent of visual examination or biopsies. Not only does this provide a powerful tool for studying tissue structure in experimental settings, the ability to detect and monitor tissue changes by microultrasound could be developed into a powerful adjunct to screening endoscopy in the clinic. PMID:27406832

  19. Berberine Inhibits Intestinal Polyps Growth in Apc (min/+) Mice via Regulation of Macrophage Polarization

    PubMed Central

    Piao, Meiyu; Cao, Hailong; He, NaNa; Yang, Boli; Dong, Wenxiao; Xu, Mengque; Yan, Fang; Zhou, Bing

    2016-01-01

    Antitumor effect of berberine has been reported in a wide spectrum of cancer, however, the mechanisms of which are not fully understood. The aim of this study was to investigate the hypothesis that berberine suppresses tumorigenesis in the familial adenomatous polyposis (FAP) by regulating the macrophage polarization in Apc (min/+) mouse model. Berberine was given to Apc (min/+) mice for 12 weeks. Primary macrophages were isolated; after berberine treatment, the change in signaling cascade was determined. The total number and size of polyps were reduced remarkably in berberine group, compared with control group. A significant decrease in protein levels of F4/80, mannose receptor (MR), and COX-2 in stroma of intestinal polyps and an increase in the level of iNOS were observed after berberine treatment. The mRNA level of MR and Arg-1 in berberine group was significantly lower than those in IL-10 or IL-4 group, while no significant difference in mRNA levels of iNOS and CXCL10 was observed. The migration and invasiveness assays in vitro showed that berberine could reduce the capability of migration and invasiveness. These findings suggest that berberine attenuates intestinal tumorigenesis by inhibiting the migration and invasion of colorectal tumor cells via regulation of macrophage polarization. PMID:27493671

  20. Increased variability in Apc(Min)/+ intestinal tissue can be measured with microultrasound.

    PubMed

    Fatehullah, A; Sharma, S; Newton, I P; Langlands, A J; Lay, H; Nelson, S A; McMahon, R K; McIlvenny, N; Appleton, P L; Cochran, S; Näthke, I S

    2016-01-01

    Altered tissue structure is a feature of many disease states and is usually measured by microscopic methods, limiting analysis to small areas. Means to rapidly and quantitatively measure the structure and organisation of large tissue areas would represent a major advance not just for research but also in the clinic. Here, changes in tissue organisation that result from heterozygosity in Apc, a precancerous situation, are comprehensively measured using microultrasound and three-dimensional high-resolution microscopy. Despite its normal appearance in conventionally examined cross-sections, both approaches revealed a significant increase in the variability of tissue organisation in Apc heterozygous tissue. These changes preceded the formation of aberrant crypt foci or adenoma. Measuring these premalignant changes using microultrasound provides a potential means to detect microscopically abnormal regions in large tissue samples, independent of visual examination or biopsies. Not only does this provide a powerful tool for studying tissue structure in experimental settings, the ability to detect and monitor tissue changes by microultrasound could be developed into a powerful adjunct to screening endoscopy in the clinic. PMID:27406832

  1. The Poly(ADP-ribose) Polymerase Enzyme Tankyrase Antagonizes Activity of the β-Catenin Destruction Complex through ADP-ribosylation of Axin and APC2.

    PubMed

    Croy, Heather E; Fuller, Caitlyn N; Giannotti, Jemma; Robinson, Paige; Foley, Andrew V A; Yamulla, Robert J; Cosgriff, Sean; Greaves, Bradford D; von Kleeck, Ryan A; An, Hyun Hyung; Powers, Catherine M; Tran, Julie K; Tocker, Aaron M; Jacob, Kimberly D; Davis, Beckley K; Roberts, David M

    2016-06-10

    Most colon cancer cases are initiated by truncating mutations in the tumor suppressor, adenomatous polyposis coli (APC). APC is a critical negative regulator of the Wnt signaling pathway that participates in a multi-protein "destruction complex" to target the key effector protein β-catenin for ubiquitin-mediated proteolysis. Prior work has established that the poly(ADP-ribose) polymerase (PARP) enzyme Tankyrase (TNKS) antagonizes destruction complex activity by promoting degradation of the scaffold protein Axin, and recent work suggests that TNKS inhibition is a promising cancer therapy. We performed a yeast two-hybrid (Y2H) screen and uncovered TNKS as a putative binding partner of Drosophila APC2, suggesting that TNKS may play multiple roles in destruction complex regulation. We find that TNKS binds a C-terminal RPQPSG motif in Drosophila APC2, and that this motif is conserved in human APC2, but not human APC1. In addition, we find that APC2 can recruit TNKS into the β-catenin destruction complex, placing the APC2/TNKS interaction at the correct intracellular location to regulate β-catenin proteolysis. We further show that TNKS directly PARylates both Drosophila Axin and APC2, but that PARylation does not globally regulate APC2 protein levels as it does for Axin. Moreover, TNKS inhibition in colon cancer cells decreases β-catenin signaling, which we find cannot be explained solely through Axin stabilization. Instead, our findings suggest that TNKS regulates destruction complex activity at the level of both Axin and APC2, providing further mechanistic insight into TNKS inhibition as a potential Wnt pathway cancer therapy. PMID:27068743

  2. Testing models of the APC tumor suppressor/β-catenin interaction reshapes our view of the destruction complex in Wnt signaling.

    PubMed

    Yamulla, Robert J; Kane, Eric G; Moody, Alexandra E; Politi, Kristin A; Lock, Nicole E; Foley, Andrew V A; Roberts, David M

    2014-08-01

    The Wnt pathway is a conserved signal transduction pathway that contributes to normal development and adult homeostasis, but is also misregulated in human diseases such as cancer. The tumor suppressor adenomatous polyposis coli (APC) is an essential negative regulator of Wnt signaling inactivated in >80% of colorectal cancers. APC participates in a multiprotein "destruction complex" that targets the proto-oncogene β-catenin for ubiquitin-mediated proteolysis; however, the mechanistic role of APC in the destruction complex remains unknown. Several models of APC function have recently been proposed, many of which have emphasized the importance of phosphorylation of high-affinity β-catenin-binding sites [20-amino-acid repeats (20Rs)] on APC. Here we test these models by generating a Drosophila APC2 mutant lacking all β-catenin-binding 20Rs and performing functional studies in human colon cancer cell lines and Drosophila embryos. Our results are inconsistent with current models, as we find that β-catenin binding to the 20Rs of APC is not required for destruction complex activity. In addition, we generate an APC2 mutant lacking all β-catenin-binding sites (including the 15Rs) and find that a direct β-catenin/APC interaction is also not essential for β-catenin destruction, although it increases destruction complex efficiency in certain developmental contexts. Overall, our findings support a model whereby β-catenin-binding sites on APC do not provide a critical mechanistic function per se, but rather dock β-catenin in the destruction complex to increase the efficiency of β-catenin destruction. Furthermore, in Drosophila embryos expressing some APC2 mutant transgenes we observe a separation of β-catenin destruction and Wg/Wnt signaling outputs and suggest that cytoplasmic retention of β-catenin likely accounts for this difference. PMID:24931405

  3. Cloning and sequencing of the allophycocyanin genes from Spirulina maxima (Cyanophyta)

    NASA Astrophysics Data System (ADS)

    Qin, Song; Hiroyuki, Kojima; Yoshikazu, Kawata; Shin-Ichi, Yano; Zeng, Cheng-Kui

    1998-03-01

    The genes coding for the α-and β-subunit of allophycocyanin ( apcA and apcB) from the cyanophyte Spirulina maxima were cloned and sequenced. The results revealed 44.4% of nucleotide sequence similarity and 30.4% of similarity of deduced amino acid sequence between them. The amino acid sequence identities between S. maxima and S. platensis are 99.4% for α subunit and 100% for β subunit.

  4. Lycopene synergistically enhances quinacrine action to inhibit Wnt-TCF signaling in breast cancer cells through APC.

    PubMed

    Preet, Ranjan; Mohapatra, Purusottam; Das, Dipon; Satapathy, Shakti R; Choudhuri, Tathagata; Wyatt, Michael D; Kundu, Chanakya N

    2013-02-01

    We previously reported that quinacrine (QC) has anticancer activity against breast cancer cells. Here, we examine the mechanism of action of QC and its ability to inhibit Wnt-TCF signaling in two independent breast cancer cell lines. QC altered Wnt-TCF signaling components by increasing the levels of adenomatous polyposis coli (APC), DAB2, GSK-3β and axin and decreasing the levels of β-catenin, p-GSK3β (ser 9) and CK1. QC also reduced the activity of the Wnt transcription factor TCF/LEF and its downstream targets cyclin D1 and c-MYC. Using a luciferase-based Wnt-TCF transcription factor assay, it was shown that APC levels were inversely associated with TCF/LEF activity. Induction of apoptosis and DNA damage was observed after treatment with QC, which was associated with increased expression of APC. The effects induced by QC depend on APC because the inhibition of Wnt-TCF signaling by QC is lost in APC-knockdown cells, and consequently, the extent of apoptosis and DNA damage caused by QC is reduced compared with parental cells. Because we previously showed that QC inhibits topoisomerase, we examined the effect of another topoisomerase inhibitor, etoposide, on Wnt signaling. Interestingly, etoposide treatment also reduced TCF/LEF activity, β-catenin and cyclin D1 levels commensurate with induction of DNA damage and apoptosis. Lycopene, a plant-derived antioxidant, synergistically increased QC activity and inhibited Wnt-TCF signaling in cancer cells without affecting the MCF-10A normal breast cell line. Collectively, the data suggest that QC-mediated Wnt-TCF signal inhibition depends on APC and that the addition of lycopene synergistically increases QC anticancer activity. PMID:23129580

  5. Peptide-MHC-I from Endogenous Antigen Outnumber Those from Exogenous Antigen, Irrespective of APC Phenotype or Activation

    PubMed Central

    Sei, Janet J.; Haskett, Scott; Kaminsky, Lauren W.; Lin, Eugene; Truckenmiller, Mary E.; Bellone, Clifford J.; Buller, R. Mark; Norbury, Christopher C.

    2015-01-01

    Naïve anti-viral CD8+ T cells (TCD8+) are activated by the presence of peptide-MHC Class I complexes (pMHC-I) on the surface of professional antigen presenting cells (pAPC). Increasing the number of pMHC-I in vivo can increase the number of responding TCD8+. Antigen can be presented directly or indirectly (cross presentation) from virus-infected and uninfected cells, respectively. Here we determined the relative importance of these two antigen presenting pathways in mousepox, a natural disease of the mouse caused by the poxvirus, ectromelia (ECTV). We demonstrated that ECTV infected several pAPC types (macrophages, B cells, and dendritic cells (DC), including DC subsets), which directly presented pMHC-I to naïve TCD8+ with similar efficiencies in vitro. We also provided evidence that these same cell-types presented antigen in vivo, as they form contacts with antigen-specific TCD8+. Importantly, the number of pMHC-I on infected pAPC (direct presentation) vastly outnumbered those on uninfected cells (cross presentation), where presentation only occurred in a specialized subset of DC. In addition, prior maturation of DC failed to enhance antigen presentation, but markedly inhibited ECTV infection of DC. These results suggest that direct antigen presentation is the dominant pathway in mice during mousepox. In a broader context, these findings indicate that if a virus infects a pAPC then the presentation by that cell is likely to dominate over cross presentation as the most effective mode of generating large quantities of pMHC-I is on the surface of pAPC that endogenously express antigens. Recent trends in vaccine design have focused upon the introduction of exogenous antigens into the MHC Class I processing pathway (cross presentation) in specific pAPC populations. However, use of a pantropic viral vector that targets pAPC to express antigen endogenously likely represents a more effective vaccine strategy than the targeting of exogenous antigen to a limiting pAPC

  6. Peptide-MHC-I from Endogenous Antigen Outnumber Those from Exogenous Antigen, Irrespective of APC Phenotype or Activation.

    PubMed

    Sei, Janet J; Haskett, Scott; Kaminsky, Lauren W; Lin, Eugene; Truckenmiller, Mary E; Bellone, Clifford J; Buller, R Mark; Norbury, Christopher C

    2015-06-01

    Naïve anti-viral CD8+ T cells (TCD8+) are activated by the presence of peptide-MHC Class I complexes (pMHC-I) on the surface of professional antigen presenting cells (pAPC). Increasing the number of pMHC-I in vivo can increase the number of responding TCD8+. Antigen can be presented directly or indirectly (cross presentation) from virus-infected and uninfected cells, respectively. Here we determined the relative importance of these two antigen presenting pathways in mousepox, a natural disease of the mouse caused by the poxvirus, ectromelia (ECTV). We demonstrated that ECTV infected several pAPC types (macrophages, B cells, and dendritic cells (DC), including DC subsets), which directly presented pMHC-I to naïve TCD8+ with similar efficiencies in vitro. We also provided evidence that these same cell-types presented antigen in vivo, as they form contacts with antigen-specific TCD8+. Importantly, the number of pMHC-I on infected pAPC (direct presentation) vastly outnumbered those on uninfected cells (cross presentation), where presentation only occurred in a specialized subset of DC. In addition, prior maturation of DC failed to enhance antigen presentation, but markedly inhibited ECTV infection of DC. These results suggest that direct antigen presentation is the dominant pathway in mice during mousepox. In a broader context, these findings indicate that if a virus infects a pAPC then the presentation by that cell is likely to dominate over cross presentation as the most effective mode of generating large quantities of pMHC-I is on the surface of pAPC that endogenously express antigens. Recent trends in vaccine design have focused upon the introduction of exogenous antigens into the MHC Class I processing pathway (cross presentation) in specific pAPC populations. However, use of a pantropic viral vector that targets pAPC to express antigen endogenously likely represents a more effective vaccine strategy than the targeting of exogenous antigen to a limiting pAPC

  7. A role for cyclin-dependent kinase(s) in the modulation of fast anterograde axonal transport: effects defined by olomoucine and the APC tumor suppressor protein

    NASA Technical Reports Server (NTRS)

    Ratner, N.; Bloom, G. S.; Brady, S. T.

    1998-01-01

    Proteins that interact with both cytoskeletal and membrane components are candidates to modulate membrane trafficking. The tumor suppressor proteins neurofibromin (NF1) and adenomatous polyposis coli (APC) both bind to microtubules and interact with membrane-associated proteins. The effects of recombinant NF1 and APC fragments on vesicle motility were evaluated by measuring fast axonal transport along microtubules in axoplasm from squid giant axons. APC4 (amino acids 1034-2844) reduced only anterograde movements, whereas APC2 (aa 1034-2130) or APC3 (aa 2130-2844) reduced both anterograde and retrograde transport. NF1 had no effect on organelle movement in either direction. Because APC contains multiple cyclin-dependent kinase (CDK) consensus phosphorylation motifs, the kinase inhibitor olomoucine was examined. At concentrations in which olomoucine is specific for cyclin-dependent kinases (5 microM), it reduced only anterograde transport, whereas anterograde and retrograde movement were both affected at concentrations at which other kinases are inhibited as well (50 microM). Both anterograde and retrograde transport also were inhibited by histone H1 and KSPXK peptides, substrates for proline-directed kinases, including CDKs. Our data suggest that CDK-like axonal kinases modulate fast anterograde transport and that other axonal kinases may be involved in modulating retrograde transport. The specific effect of APC4 on anterograde transport suggests a model in which the binding of APC to microtubules may limit the activity of axonal CDK kinase or kinases in restricted domains, thereby affecting organelle transport.

  8. A novel GSK3-regulated APC:Axin interaction regulates Wnt signaling by driving a catalytic cycle of efficient βcatenin destruction

    PubMed Central

    Pronobis, Mira I; Rusan, Nasser M; Peifer, Mark

    2015-01-01

    APC, a key negative regulator of Wnt signaling in development and oncogenesis, acts in the destruction complex with the scaffold Axin and the kinases GSK3 and CK1 to target βcatenin for destruction. Despite 20 years of research, APC's mechanistic function remains mysterious. We used FRAP, super-resolution microscopy, functional tests in mammalian cells and flies, and other approaches to define APC's mechanistic role in the active destruction complex when Wnt signaling is off. Our data suggest APC plays two roles: (1) APC promotes efficient Axin multimerization through one known and one novel APC:Axin interaction site, and (2) GSK3 acts through APC motifs R2 and B to regulate APC:Axin interactions, promoting high-throughput of βcatenin to destruction. We propose a new dynamic model of how the destruction complex regulates Wnt signaling and how this goes wrong in cancer, providing insights into how this multiprotein signaling complex is assembled and functions via multivalent interactions. DOI: http://dx.doi.org/10.7554/eLife.08022.001 PMID:26393419

  9. Regulation of the phosphorylation and nuclear import and export of β-catenin by APC and its cancer-related truncated form.

    PubMed

    Wang, Lili; Liu, Xiaoyong; Gusev, Ekaterina; Wang, Chuanxin; Fagotto, François

    2014-04-15

    We report the first direct analysis of the endogenous β-catenin phosphorylation activity in colon cancer SW480 cells. By comparing parental SW480 cells that harbor a typical truncated adenomatous polyposis coli (APC) form, cells expressing full-length APC and APC-depleted cells, we provide the formal demonstration that APC is necessary for β-catenin phosphorylation, both for priming of the protein at residue serine 45 and for the subsequent phosphorylation of residues 33, 37 and 41. Truncated APC still sustains a surprisingly high phosphorylation activity, which requires the protein to bind to β-catenin through the APC 20-amino-acid (20AA) repeats, thus providing a biochemical explanation for the precise truncations found in cancer cells. We also show that most of the β-catenin phosphorylation activity is associated with a dense insoluble fraction. We finally examine the impact of full-length and truncated APC on β-catenin nuclear transport. We observe that β-catenin is transported much faster than previously thought. Although this fast translocation is largely insensitive to the presence of wild-type or truncated APC, the two forms appear to limit the pool of β-catenin that is available for transport, which could have an impact on β-catenin nuclear activities in normal and cancer cells. PMID:24496450

  10. A novel GSK3-regulated APC:Axin interaction regulates Wnt signaling by driving a catalytic cycle of efficient βcatenin destruction.

    PubMed

    Pronobis, Mira I; Rusan, Nasser M; Peifer, Mark

    2015-01-01

    APC, a key negative regulator of Wnt signaling in development and oncogenesis, acts in the destruction complex with the scaffold Axin and the kinases GSK3 and CK1 to target βcatenin for destruction. Despite 20 years of research, APC's mechanistic function remains mysterious. We used FRAP, super-resolution microscopy, functional tests in mammalian cells and flies, and other approaches to define APC's mechanistic role in the active destruction complex when Wnt signaling is off. Our data suggest APC plays two roles: (1) APC promotes efficient Axin multimerization through one known and one novel APC:Axin interaction site, and (2) GSK3 acts through APC motifs R2 and B to regulate APC:Axin interactions, promoting high-throughput of βcatenin to destruction. We propose a new dynamic model of how the destruction complex regulates Wnt signaling and how this goes wrong in cancer, providing insights into how this multiprotein signaling complex is assembled and functions via multivalent interactions. PMID:26393419

  11. Missense Polymorphisms in the Adenomatous Polyposis Coli Gene and Colorectal Cancer Risk

    PubMed Central

    Cleary, Sean P.; Kim, Hyeja; Croitoru, Marina E.; Redston, Mark; Knight, Julia A.; Gallinger, Steven; Gryfe, Robert

    2009-01-01

    PURPOSE Whereas truncating germline mutations of the adenomatous polyposis coli (APC) gene give rise to familial adenomatous polyposis, missense polymorphisms of APC may confer a weaker risk for colorectal cancer. METHODS We sequenced the entire open reading frame of the APC gene and tested for two common MYH mutations in a population-based series of patients with colorectal cancer and 5 to 99 adenomas. Missense adenomatous polyposis coli alterations identified in this colorectal cancer multiple-polyp population were analyzed in a population-based series of patients with colorectal cancer and healthy control subjects. RESULTS Germline APC or mutY human homologue (MYH) alterations were identified in 16 of 39 colorectal cancer-multiple polyp patients. Four missense APC gene alterations (S130G, E1317Q, Dl822V, G2502S) were observed in 13 individuals and 3 additional patients carried presumed pathogenic (APC Y94X, biallelic MYH Y165C and heterozygous MYH G382D) mutations. When independently assessed in 971 patients with colorectal cancer and 954 healthy control subjects, none of the identified missense APC alterations conferred a significantly increased risk for colorectal cancer, odds ratio (95 percent confidence intervals): S130G=3.1 (0.29–32.25), E1317Q= 1.08 (0.59–2.74), G2502S= 1 (0.65–1.63), D1822V (heterozygous)=0.79 (0.64–0.98), D1822V (homozygous) =0.82 (0.63–1.27). CONCLUSIONS Germline missense APC alterations observed in 33 percent of patients with multiple colorectal neoplasms seemed to play a limited role in colorectal cancer risk when independently assessed by a population-based, case-control analysis. PMID:18612690

  12. Creation of an engineered APC system to explore and optimize the presentation of immunodominant peptides of major allergens.

    PubMed

    Rosskopf, Sandra; Jutz, Sabrina; Neunkirchner, Alina; Candia, Martín R; Jahn-Schmid, Beatrice; Bohle, Barbara; Pickl, Winfried F; Steinberger, Peter

    2016-01-01

    We have generated engineered APC to present immunodominant peptides derived from the major aero-allergens of birch and mugwort pollen, Bet v 1142-153 and Art v 125-36, respectively. Jurkat-based T cell reporter lines expressing the cognate allergen-specific T cell receptors were used to read out the presentation of allergenic peptides on the engineered APC. Different modalities of peptide loading and presentation on MHC class II molecules were compared. Upon exogenous loading with allergenic peptides, the engineered APC elicited a dose-dependent response in the reporter T cells and the presence of chemical loading enhancers strongly increased reporter activation. Invariant chain-based MHC class II targeting strategies of endogenously expressed peptides resulted in stronger activation of the reporters than exogenous loading. Moreover, we used Bet v 1 as model allergen to study the ability of K562 cells to present antigenic peptides derived from whole proteins either taken up or endogenously expressed as LAMP-1 fusion protein. In both cases the ability of these cells to process and present peptides derived from whole proteins critically depended on the expression of HLA-DM. We have identified strategies to achieve efficient presentation of allergenic peptides on engineered APC and demonstrate their use to stimulate T cells from allergic individuals. PMID:27539532

  13. Application of the microbiological method DEFT/APC to detect minimally processed vegetables treated with gamma radiation

    NASA Astrophysics Data System (ADS)

    Araújo, M. M.; Duarte, R. C.; Silva, P. V.; Marchioni, E.; Villavicencio, A. L. C. H.

    2009-07-01

    Marketing of minimally processed vegetables (MPV) are gaining impetus due to its convenience, freshness and apparent health effect. However, minimal processing does not reduce pathogenic microorganisms to safe levels. Food irradiation is used to extend the shelf life and to inactivate food-borne pathogens. In combination with minimal processing it could improve safety and quality of MPV. A microbiological screening method based on the use of direct epifluorescent filter technique (DEFT) and aerobic plate count (APC) has been established for the detection of irradiated foodstuffs. The aim of this study was to evaluate the applicability of this technique in detecting MPV irradiation. Samples from retail markets were irradiated with 0.5 and 1.0 kGy using a 60Co facility. In general, with a dose increment, DEFT counts remained similar independent of the irradiation while APC counts decreased gradually. The difference of the two counts gradually increased with dose increment in all samples. It could be suggested that a DEFT/APC difference over 2.0 log would be a criteria to judge if a MPV was treated by irradiation. The DEFT/APC method could be used satisfactorily as a screening method for indicating irradiation processing.

  14. Colon epithelial cell differentiation is inhibited by constitutive c-myb expression or mutant APC plus activated RAS.

    PubMed

    Ramsay, Robert G; Ciznadija, Daniel; Sicurella, Catherine; Reyes, Nancy; Mitchelhill, Ken; Darcy, Phillip K; D'Abaco, Giovanna; Mantamadiotis, Theo

    2005-01-01

    Blocked differentiation is a hallmark of cancer cells and the restoration of differentiation programs in vivo is an actively pursued clinical aim. Understanding the key regulators of cyto-differentiation may focus therapies on molecules that reactivate this process. c-myb expression declines rapidly when human colon cancer epithelial cells are induced to differentiate with the physiologically relevant short-chain fatty acid, sodium butyrate. These cells show increased expression of alkaline phosphatase and cytokeratin 8. Similarly, murine Immorto-epithelial cells derived from wild-type colon cells also show c-myb mRNA declines when induced to differentiate with sodium butyrate. Immorto-cells harboring a single APC mutation are indistinguishable from wild-type cells with regard to differentiation, while addition of activated RAS alone markedly enhances differentiation. In marked contrast, complete differentiation arrest occurs when both APC and RAS are mutated. Expression of MybER, a 4-hydroxytamoxifen-activatable form of c-Myb, blocks differentiation in wildtype and APC mutant Immorto-cell lines as well as LIM1215 human colon carcinoma cells. These data identify two pathways of oncogenic change that lead to retarded epithelial cell differentiation, one involving the presence of a single APC mutation in conjunction with activated RAS or alternatively constitutive c-myb expression. PMID:15684716

  15. 77 FR 34455 - In the Matter of Aegis Assessments, Inc., APC Group, Inc., Aurelio Resource Corp., BioAuthorize...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-06-11

    ... From the Federal Register Online via the Government Publishing Office SECURITIES AND EXCHANGE COMMISSION In the Matter of Aegis Assessments, Inc., APC Group, Inc., Aurelio Resource Corp., BioAuthorize... securities of BioAuthorize Holdings, Inc. because it has not filed any periodic reports since the...

  16. Positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis in budding yeast.

    PubMed

    Hatano, Yuhki; Naoki, Koike; Suzuki, Asuka; Ushimaru, Takashi

    2016-10-01

    The mitotic inhibitor securin is degraded via the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C)-Cdc20 after anaphase onset. This triggers activation of the mitotic protease separase and thereby sister chromatid separation. However, only a proportion of securin molecules are degraded at metaphase-anaphase transition and the remaining molecules are still present in anaphase. The roles of securin and separase in late mitosis remain elusive. Here, we show that securin still inhibits separase to repress mitotic exit in anaphase in budding yeast. APC/C-Cdh1-mediated securin degradation at telophase further liberated separase, which promotes Cdc14 release and mitotic exit. Separase executed these events via its proteolytic action and that in the Cdc14 early release (FEAR) network. Cdc14 release further activated APC/C-Cdh1 in the manner of a positive feedback loop. Thus, the positive feedback promotes mitotic exit via the APC/C-Cdh1-separase-Cdc14 axis. This study shows the importance of the two-step degradation mode of securin and the role of separase in mitotic exit. PMID:27418100

  17. FIP200 inhibits β-catenin-mediated transcription by promoting APC-independent β-catenin ubiquitination.

    PubMed

    Choi, J D; Ryu, M; Ae Park, M; Jeong, G; Lee, J-S

    2013-05-01

    Focal adhesion kinase-family-interacting protein of 200 kDa (FIP200) has been shown to regulate multiple cellular functions, including cell adhesion, autophagy, development and proliferation. Furthermore, FIP200 is considered to have tumor-suppressive activity, which may be correlated with its inactivation in human breast cancers, in addition to its role as an important signal transduction node. Herein, we report that FIP200 interacts with the oncoprotein β-catenin. Moreover, FIP200 promotes destabilization of wild-type β-catenin, but not a cancer-causing form of β-catenin, and as a result represses the β-catenin-mediated transcription. FIP200-induced degradation of β-catenin is independent of adenomatous polyposis coli (APC) of the well-established β-catenin destruction complex (glycogen synthase kinase-3β/axin/APC), in a component of β-catenin E3 ubiquitin ligase, β-TrCP-dependent manner. Thus, the APC-independent β-catenin degradation by FIP200 suggests a role for FIP200 in tumor suppression in the presence of APC dysfunction. These findings reveal a new and important function of FIP200 in regulation of the Wnt/β-catenin pathway. PMID:22751121

  18. EM Structure of human APC/CCDH1-EMI1 reveals multimodal mechanism of E3 ligase shutdown

    PubMed Central

    Frye, Jeremiah J.; Brown, Nicholas G.; Petzold, Georg; Watson, Edmond R.; Grace, Christy R. R.; Nourse, Amanda; Jarvis, Marc A.; Kriwacki, Richard W.; Peters, Jan-Michael; Stark, Holger; Schulman, Brenda A.

    2013-01-01

    The Anaphase Promoting Complex/Cyclosome (APC/C) is a ~1.5 MDa multiprotein E3 ligase enzyme that regulates cell division by promoting timely ubiquitin-mediated proteolysis of key cell cycle regulatory proteins. Inhibition of human APC/CCDH1 during interphase by Early Mitotic Inhibitor 1 (EMI1) is essential for accurate coordination of DNA synthesis and mitosis. Here, we report a hybrid structural approach involving NMR, electron microscopy, and enzymology, which reveal that EMI1’s 143-residue C-terminal domain inhibits multiple APC/CCDH1 functions. The intrinsically disordered D-box, Linker, and Tail elements, together with a structured zinc-binding domain, bind distinct regions of APC/CCDH1 to synergistically both block the substrate-binding site and inhibit ubiquitin chain elongation. The functional importance of intrinsic structural disorder is explained by enabling a small inhibitory domain to bind multiple sites to shut down multiple functions of a “molecular machine” nearly 100 times its size. PMID:23708605

  19. Creation of an engineered APC system to explore and optimize the presentation of immunodominant peptides of major allergens

    PubMed Central

    Rosskopf, Sandra; Jutz, Sabrina; Neunkirchner, Alina; Candia, Martín R.; Jahn-Schmid, Beatrice; Bohle, Barbara; Pickl, Winfried F.; Steinberger, Peter

    2016-01-01

    We have generated engineered APC to present immunodominant peptides derived from the major aero-allergens of birch and mugwort pollen, Bet v 1142–153 and Art v 125–36, respectively. Jurkat-based T cell reporter lines expressing the cognate allergen-specific T cell receptors were used to read out the presentation of allergenic peptides on the engineered APC. Different modalities of peptide loading and presentation on MHC class II molecules were compared. Upon exogenous loading with allergenic peptides, the engineered APC elicited a dose-dependent response in the reporter T cells and the presence of chemical loading enhancers strongly increased reporter activation. Invariant chain-based MHC class II targeting strategies of endogenously expressed peptides resulted in stronger activation of the reporters than exogenous loading. Moreover, we used Bet v 1 as model allergen to study the ability of K562 cells to present antigenic peptides derived from whole proteins either taken up or endogenously expressed as LAMP-1 fusion protein. In both cases the ability of these cells to process and present peptides derived from whole proteins critically depended on the expression of HLA-DM. We have identified strategies to achieve efficient presentation of allergenic peptides on engineered APC and demonstrate their use to stimulate T cells from allergic individuals. PMID:27539532

  20. Dual-mode regulation of the APC/C by CDK1 and MAPK controls meiosis I progression and fidelity.

    PubMed

    Nabti, Ibtissem; Marangos, Petros; Bormann, Jenny; Kudo, Nobuaki R; Carroll, John

    2014-03-17

    Female meiosis is driven by the activities of two major kinases, cyclin-dependent kinase 1 (Cdk1) and mitogen-activated protein kinase (MAPK). To date, the role of MAPK in control of meiosis is thought to be restricted to maintaining metaphase II arrest through stabilizing Cdk1 activity. In this paper, we find that MAPK and Cdk1 play compensatory roles to suppress the anaphase-promoting complex/cyclosome (APC/C) activity early in prometaphase, thereby allowing accumulation of APC/C substrates essential for meiosis I. Furthermore, inhibition of MAPK around the onset of APC/C activity at the transition from meiosis I to meiosis II led to accelerated completion of meiosis I and an increase in aneuploidy at metaphase II. These effects appear to be mediated via a Cdk1/MAPK-dependent stabilization of the spindle assembly checkpoint, which when inhibited leads to increased APC/C activity. These findings demonstrate new roles for MAPK in the regulation of meiosis in mammalian oocytes. PMID:24637322

  1. Dual-mode regulation of the APC/C by CDK1 and MAPK controls meiosis I progression and fidelity

    PubMed Central

    Nabti, Ibtissem; Bormann, Jenny; Kudo, Nobuaki R.

    2014-01-01

    Female meiosis is driven by the activities of two major kinases, cyclin-dependent kinase 1 (Cdk1) and mitogen-activated protein kinase (MAPK). To date, the role of MAPK in control of meiosis is thought to be restricted to maintaining metaphase II arrest through stabilizing Cdk1 activity. In this paper, we find that MAPK and Cdk1 play compensatory roles to suppress the anaphase-promoting complex/cyclosome (APC/C) activity early in prometaphase, thereby allowing accumulation of APC/C substrates essential for meiosis I. Furthermore, inhibition of MAPK around the onset of APC/C activity at the transition from meiosis I to meiosis II led to accelerated completion of meiosis I and an increase in aneuploidy at metaphase II. These effects appear to be mediated via a Cdk1/MAPK-dependent stabilization of the spindle assembly checkpoint, which when inhibited leads to increased APC/C activity. These findings demonstrate new roles for MAPK in the regulation of meiosis in mammalian oocytes. PMID:24637322

  2. Swelling behavior of O-alkylated APCS coals as examined by the EPR spin probe method

    SciTech Connect

    Ding, Ruisong; Tucker, D.; Kispert, L.D.

    1995-12-31

    Known O-alkylation procedures have been used to derivatize the carboxyl and hydroxyl groups in the APCS coals Lewiston-Stockton, Wyodak-Anderson, Beulah-Zap, Illinois {number_sign}6, Upper Freeport, and Pittsburgh {number_sign}8. In general the resulting decrease in hydrogen bonding reduced the cyclical variation in nitroxide spin probe retention observed for nonalkylated coals when small amounts (<1%) of pyridine are present in the toluene swelling solvent. An increase in spin probe retention by the O-alkylated coals relative to the underivatized coals indicates a more open arrangement in the coal due to a decrease in attractive forces, confirming that microporosity increases with increasing rank.

  3. Statin Use and Colorectal Adenoma Risk: Results from the Adenoma Prevention with Celecoxib (APC) Trial

    PubMed Central

    Bertagnolli, Monica M.; Hsu, Meier; Hawk, Ernest T.; Eagle, Craig J.; Zauber, Ann G.

    2010-01-01

    Background Statins are widely prescribed for cardiovascular disease prevention, and also commonly used in patients at high risk for colorectal cancer (CRC). We report the results of a planned secondary analysis of the relationship between statin use and colorectal adenoma risk in a large chemoprevention trial. Methods The Adenoma Prevention with Celecoxib (APC) trial randomized 2035 adenoma patients to receive placebo (679 patients), 200 mg celecoxib twice daily (685 patients), or 400 mg celecoxib twice daily (671 patients). The study collected complete medical history and medication use data, and performed colonoscopic surveillance to 5 years after study enrollment. Effects of statin use on newly detected adenomas and cardiovascular adverse events were analyzed as time-dependent variables by multivariable Cox regression. Results Statins were used by 36% (N=730) of APC trial participants. When adjusted for covariates including cardioprotective aspirin use, age, and sex, participants on the placebo arm who used statins at any time had no benefit over 5 years compared to never users (Risk Ratio (RR) 1.24; 95% confidence interval (CI) (0.99-1.56); p=0.065). Statin use for >3 years increased adenoma risk over 5 years (RR 1.39; 95%CI 1.04-1.86; p=0.024). For all comparisons of patients treated with celecoxib, adenoma detection rates for statin users and non-users were equivalent. Consistent with their use in patients at high risk, cardiovascular serious adverse events were more common among statin users. Conclusions For patients at high risk of CRC, statins do not protect against colorectal neoplasms and may even increase the risk of developing colorectal adenomas. PMID:20403998

  4. Self-association of the APC tumor suppressor is required for the assembly, stability, and activity of the Wnt signaling destruction complex.

    PubMed

    Kunttas-Tatli, Ezgi; Roberts, David M; McCartney, Brooke M

    2014-11-01

    The tumor suppressor adenomatous polyposis coli (APC) is an essential negative regulator of Wnt signaling through its activity in the destruction complex with Axin, GSK3β, and CK1 that targets β-catenin/Armadillo (β-cat/Arm) for proteosomal degradation. The destruction complex forms macromolecular particles we termed the destructosome. Whereas APC functions in the complex through its ability to bind both β-cat and Axin, we hypothesize that APC proteins play an additional role in destructosome assembly through self-association. Here we show that a novel N-terminal coil, the APC self-association domain (ASAD), found in vertebrate and invertebrate APCs, directly mediates self-association of Drosophila APC2 and plays an essential role in the assembly and stability of the destructosome that regulates β-cat degradation in Drosophila and human cells. Consistent with this, removal of the ASAD from the Drosophila embryo results in β-cat/Arm accumulation and aberrant Wnt pathway activation. These results suggest that APC proteins are required not only for the activity of the destructosome, but also for the assembly and stability of this macromolecular machine. PMID:25208568

  5. Mps1Mph1 Kinase Phosphorylates Mad3 to Inhibit Cdc20Slp1-APC/C and Maintain Spindle Checkpoint Arrests

    PubMed Central

    Syred, Heather M.; van der Sar, Sjaak; Patel, Hitesh; Moresco, James J.; Sarkeshik, Ali; Yates, John R.; Rappsilber, Juri; Hardwick, Kevin G.

    2016-01-01

    The spindle checkpoint is a mitotic surveillance system which ensures equal segregation of sister chromatids. It delays anaphase onset by inhibiting the action of the E3 ubiquitin ligase known as the anaphase promoting complex or cyclosome (APC/C). Mad3/BubR1 is a key component of the mitotic checkpoint complex (MCC) which binds and inhibits the APC/C early in mitosis. Mps1Mph1 kinase is critical for checkpoint signalling and MCC-APC/C inhibition, yet few substrates have been identified. Here we identify Mad3 as a substrate of fission yeast Mps1Mph1 kinase. We map and mutate phosphorylation sites in Mad3, producing mutants that are targeted to kinetochores and assembled into MCC, yet display reduced APC/C binding and are unable to maintain checkpoint arrests. We show biochemically that Mad3 phospho-mimics are potent APC/C inhibitors in vitro, demonstrating that Mad3p modification can directly influence Cdc20Slp1-APC/C activity. This genetic dissection of APC/C inhibition demonstrates that Mps1Mph1 kinase-dependent modifications of Mad3 and Mad2 act in a concerted manner to maintain spindle checkpoint arrests. PMID:26882497

  6. Interaction of APC/C-E3 Ligase with Swi6/HP1 and Clr4/Suv39 in Heterochromatin Assembly in Fission Yeast*S⃞♦

    PubMed Central

    Dubey, Rudra Narayan; Nakwal, Nandni; Bisht, Kamlesh Kumar; Saini, Ashok; Haldar, Swati; Singh, Jagmohan

    2009-01-01

    Heterochromatin assembly in fission yeast is initiated by binding of Swi6/HP1 to the Lys-9-dimethylated H3 followed by spreading via cooperative recruitment of Swi6/HP1. Recruitment of Cohesin by Swi6/HP1 further stabilizes the heterochromatin structure and integrity. Subsequently, polyubiquitylation of Cut2 by anaphase-promoting complex-cyclosome (APC/C)-ubiquitin-protein isopeptide ligase (E3 ligase) followed by degradation of Cut2 releases Cut1, which cleaves the Rad21 subunit of Cohesin, facilitating sister chromatid separation during mitosis. Here, we demonstrate a surprising role of APC/C in assembly of heterochromatin and silencing at mating type, centromere, and ribosomal DNA loci. Coincidentally with the loss of silencing, recruitment of Swi6, H3-Lys-9-Me2, and Clr4 at dg-dh repeats at cen1 and the K region of mat locus is abrogated in mutants cut4, cut9, and nuc2. Surprisingly, both Cut4 and Cut9 are also highly enriched at these regions in wild type and depleted in swi6Δ mutant. Cut4 and Cut9 interact directly with Swi6/HP1 and Clr4, whereas the mutant Cut4 does not, suggesting that a direct physical interaction of APC subunits Cut4 and Cut9 with Swi6 and Clr4 is instrumental in heterochromatin assembly. The silencing defect in APC mutants is causally related to ubiquitylation activity of APC-E3 ligase. Like swi6 mutant, APC mutants are also defective in Cohesin recruitment and exhibit defects like lagging chromosomes, chromosome loss, and aberrant recombination in the mat region. In addition, APC mutants exhibit a bidirectional expression of dh repeats, suggesting a role in the RNA interference pathway. Thus, APC and heterochromatin proteins Swi6 and Clr4 play a mutually cooperative role in heterochromatin assembly, thereby ensuring chromosomal integrity, inheritance, and segregation during mitosis and meiosis. PMID:19117951

  7. Increase in a distinct pulmonary macrophage subset possessing an antigen-presenting cell phenotype and in vitro APC activity following silica exposure

    SciTech Connect

    Migliaccio, Christopher T. . E-mail: christopher.migliaccio@umontana.edu; Hamilton, Raymond F.; Holian, Andrij

    2005-06-01

    Silica inhalation results in chronic lung inflammation and fibrosis. While the role of the alveolar macrophage (AM) is considered key to the effects of silica on lung pathology, the etiology is not completely understood. Evidence suggests an increase in antigen presenting cell (APC) activity as a contributing factor to this process, as well as potential roles for both AM and interstitial macrophages (IM) in silicosis. In order to study the effects of crystalline silica on the APC activity of pulmonary macrophages, mice were exposed intranasally and changes in pulmonary macrophage populations were assessed using flow cytometry. Following intranasal instillation of silica, a significant increase in the APC activity of AM was observed, as well as a significant increase in a subset of IM expressing classic APC markers (MHC class II, CD11c). In addition, an in vitro system using bone marrow-derived macrophages (BMDM) was generated to assess the effects of silica on the APC activity of macrophages in vitro. Data using BMDM in the in vitro APC assay demonstrated a significant increase in APC activity following silica exposure, but not following exposure to saline or a control particle (TiO{sub 2}). Using a combination of in vivo and in vitro experiments, the current study describes a significant increase in an interstitial macrophage subset with an APC phenotype, as well as an increase in the APC activity of both AM and BMDM, as a direct result of exposure to crystalline silica. These studies suggest a specific mechanism, macrophage subset activation, by which crystalline silica exposure results in chronic pulmonary inflammation and, eventually, fibrosis.

  8. Exercise effects on polyp burden and immune markers in the ApcMin/+ mouse model of intestinal tumorigenesis.

    PubMed

    McClellan, Jamie L; Steiner, Jennifer L; Day, Stani D; Enos, Reilly T; Davis, Mark J; Singh, Udai P; Murphy, E Angela

    2014-08-01

    Many observational epidemiologic studies suggest an association between exercise and colon cancer risk. The mechanisms contributing to a preventative effect of exercise on colon cancer are complex and multifaceted. Altered immune system function is one possible mechanism that has been largely unexplored. Therefore, the purpose of this study was to examine the effects of exercise on markers associated with macrophages and select T cell populations in a mouse model of intestinal tumorigenesis and to relate this to polyp characteristics. Male Apc(Min/+) mice were randomly assigned to either sedentary (Sed) or exercise (Ex) treatment (n=6-9/group). The exercise treatment consisted of treadmill running for 1 h/day and 6 days a week at 15 m/min from 4 until 16 weeks of age. Intestinal polyps were counted and categorized by size. Mucosal tissue was analyzed for mRNA expression of overall macrophages (F4/80), for genes associated with M1 (IL-12, IL-23 and Nos2) and M2 (CD206, IL-10, IL-4, CCL17, CCL22 and Arg-1) macrophages and the macrophage chemoattractants MCP-1, fetuin A and CXCL14. Markers for cytotoxic T cells (CTLs) and regulatory T cells were also examined by measuring mRNA expression of CD8 and Foxp3, respectively. While there was no significant difference in overall polyp number between the groups (Sed, 23.3±4.3; and Ex, 16.5±4.3), Ex did have a reduction in the number of large polyps (Sed, 6.1±1.1; and Ex, 3.0±0.6) (P<0.05). This was consistent with a decrease in spleen weight (P<0.05). Similarly, Ex reduced mRNA expression of overall macrophages (F4/80) as well as markers associated with both M1 (IL-12) and M2 (CD206, CCL22 and Arg-1) subtypes (P<0.05) but there was no significant decrease in macrophage chemoattractants. CD8 expression was increased while Foxp3 expression was decreased with Ex (P<0.05). Overall the data provide important new information on immune regulation as a possible mechanism for the documented benefits of exercise training on reducing

  9. Five Year Efficacy and Safety Analysis of the Adenoma Prevention with Celecoxib (APC) Trial

    PubMed Central

    Bertagnolli, Monica M.; Eagle, Craig J.; Zauber, Ann G.; Redston, Mark; Breazna, Aurora; Kim, KyungMann; Tang, Jie; Rosenstein, Rebecca B.; Umar, Asad; Bagheri, Donya; Collins, Neal T.; Burn, John; Chung, Daniel C.; Dewar, Thomas; Foley, T. Raymond; Hoffman, Neville; Macrae, Finlay; Pruitt, Ronald E.; Saltzman, John R.; Salzberg, Bruce; Sylwestrowicz, Thomas; Hawk, Ernest T.

    2010-01-01

    The Adenoma Prevention with Celecoxib (APC) Trial examined the efficacy and safety of the Cox-2 inhibitor, celecoxib, for sporadic colorectal adenoma prevention in patients at high risk for colorectal cancer (CRC). The trial randomized 2035 subjects to receive either placebo, celecoxib 200mg twice daily, or celecoxib 400mg twice daily. The primary study safety and efficacy analyses involved three years of treatment. The results showed significant anti-tumor effect, but also indicated increased cardiovascular adverse events in patients treated with celecoxib compared to placebo. A total of 933 patients participated in an extension of the APC Trial, with a planned total treatment and surveillance duration of 5 years. Study medication was stopped early, resulting in a median treatment duration of 3.1 years. Patients treated on the placebo arm had a cumulative adenoma incidence of 68.4% over 5 years of observation. This figure was 59.0% (p<0.0001) for those receiving low dose celecoxib, and 60.1% (p<0.0001) for those receiving high dose celecoxib. The cumulative incidence of advanced adenomas over 5 years was 21.3% of those taking placebo, 12.5% (p<0.0001) of those taking low dose celecoxib and 15.8% (p<0.0001) of those taking high dose celecoxib. Investigator reported treatment emergent adverse events were similar across all treatment groups for categories including renal and hypertensive events and gastrointestinal ulceration and hemorrhage events. For a category composed of cardiovascular and thrombotic events, the risk relative to placebo was 1.6 (95%CI 1.0, 2.5) for those using 200mg BID celecoxib and 1.9 (95%CI 1.2, 3.1) for those using 400mg BID celecoxib. Secondary analysis showed an interaction between a baseline history of atherosclerotic heart disease and study drug use with respect to cardiovascular and thrombotic adverse events (p=0.004). These results confirm the inhibitory effect of celecoxib on colorectal adenoma formation, and provide additional safety

  10. Metabonomic Profiling Reveals Cancer Chemopreventive Effects of American Ginseng on Colon Carcinogenesis in Apc(Min/+) Mice.

    PubMed

    Xie, Guoxiang; Wang, Chong-Zhi; Yu, Chunhao; Qiu, Yunping; Wen, Xiao-Dong; Zhang, Chun-Feng; Yuan, Chun-Su; Jia, Wei

    2015-08-01

    American ginseng (Panax quinquefolius L.) is one of the most commonly used herbal medicines in the West. It has been reported to possess significant antitumor effects that inhibit the process of carcinogenesis. However, the mechanisms underlying its anticancer effects remain largely unresolved. In this study, we investigated the cancer chemopreventive effects of American ginseng on the progression of high fat (HF) diet-enhanced colorectal carcinogenesis with a genetically engineered Apc(Min/+) mouse model. The metabolic alterations in sera of experimental mice perturbed by HF diet intervention as well as the American ginseng treatment were measured by gas chromatography time-of-flight mass spectrometry (GC-TOFMS) and liquid chromatography time-of-flight mass spectrometry (LC-TOFMS) analysis. American ginseng treatment significantly extended the life span of the Apc(Min/+) mouse. Significant alterations of metabolites involving amino acids, organic acids, fatty acids, and carbohydrates were observed in Apc(Min/+) mouse in sera, which were attenuated by American ginseng treatment and concurrent with the histopathological improvement with significantly reduced tumor initiation, progression and gut inflammation. These metabolic changes suggest that the preventive effect of American ginseng is associated with attenuation of impaired amino acid, carbohydrates, and lipid metabolism. It also appears that American ginseng induced significant metabolic alterations independent of the Apc(Min/+) induced metabolic changes. The significantly altered metabolites induced by American ginseng intervention include arachidonic acid, linolelaidic acid, glutamate, docosahexaenoate, tryptophan, and fructose, all of which are associated with inflammation and oxidation. This suggests that American ginseng exerts the chemopreventive effects by anti-inflammatory and antioxidant mechanisms. PMID:26136108

  11. A l-Lysine Transporter of High Stereoselectivity of the Amino Acid-Polyamine-Organocation (APC) Superfamily

    PubMed Central

    Kaur, Jagdeep; Olkhova, Elena; Malviya, Viveka Nand; Grell, Ernst; Michel, Hartmut

    2014-01-01

    Membrane proteins of the amino acid-polyamine-organocation (APC) superfamily transport amino acids and amines across membranes and play an important role in the regulation of cellular processes. We report the heterologous production of the LysP-related transporter STM2200 from Salmonella typhimurium in Escherichia coli, its purification, and functional characterization. STM2200 is assumed to be a proton-dependent APC transporter of l-lysine. The functional interaction between basic amino acids and STM2200 was investigated by thermoanalytical methods, i.e. differential scanning and isothermal titration calorimetry. Binding of l-lysine to STM2200 in its solubilized monomer form is entropy-driven. It is characterized by a dissociation constant of 40 μm at pH 5.9 and is highly selective; no evidence was found for the binding of l-arginine, l-ornithine, l-2,4-diaminobutyric acid, and l-alanine. d-Lysine is bound 45 times more weakly than its l-chiral form. We thus postulate that STM2200 functions as a specific transport protein. Based on the crystal structure of ApcT (Shaffer, P. L., Goehring, A., Shankaranarayanan, A., and Gouaux, E. (2009) Science 325, 1010–1014), a proton-dependent amino acid transporter of the APC superfamily, a homology model of STM2200 was created. Docking studies allowed identification of possible ligand binding sites. The resulting predictions indicated that Glu-222 and Arg-395 of STM2200 are markedly involved in ligand binding, whereas Lys-163 is suggested to be of structural and functional relevance. Selected variants of STM2200 where these three amino acid residues were substituted using single site-directed mutagenesis showed no evidence for l-lysine binding by isothermal titration calorimetry, which confirmed the predictions. Molecular aspects of the observed ligand specificity are discussed. PMID:24257746

  12. Racial variations in frequency and phenotypes of APC and MUTYH mutations in 6,169 individuals undergoing genetic testing

    PubMed Central

    Inra, Jennifer A.; Steyerberg, Ewout W.; Grover, Shilpa; McFarland, Ashley; Syngal, Sapna; Kastrinos, Fay

    2016-01-01

    Purpose To assess whether differences in frequency and phenotype of APC and MUTYH mutations exist among racially/ethnically diverse populations. Methods 6169 individuals with personal and/or family history of colorectal cancer (CRC) and polyps were studied. APC testing involved full sequencing/large rearrangement analysis (FS/LRA); MUTYH involved “panel testing” (for Y165C, G382D mutations), or FS/LRA, performed by Myriad Genetics, a commercial laboratory. Subjects were identified as Caucasian, Asian, African American (AA), or Other. Statistical tests included Chi-Square, Fisher’s Exact, ANOVA and z-approximation. Results 17.5% had pathogenic APC mutations. 4.8% were biallelic MUTYH carriers. 18% were non-Caucasian with >100 adenomas and younger ages of adenoma or CRC diagnosis (p<0.0001) than Caucasians. The overall APC mutation rate was higher in Asians, AAs and Others compared to Caucasians (25.2%, 30.9%, 24%, 15.5%;p<0.0001) but similar in all groups when adjusted for polyp burden. More MUTYH biallelic carriers were Caucasian or Other than Asian or AA (5%, 7%, 2.7%, 0.3%;p<0.0001). Among Caucasians, 5% were biallelic carriers identified by panel testing versus 2% by sequencing/LRA (p=0.002). Among non-Caucasians, 3% undergoing panel testing were biallelic carriers versus 10% identified by sequencing/LRA(p<0.0002). Conclusion Non-Caucasians undergo genetic testing at more advanced stages of polyposis and/or younger ages of CRC/polyp diagnosis. Restricted MUTYH analysis may miss significant numbers of biallelic carriers, particularly in non-Caucasians. PMID:25590978

  13. Sulindac increases the expression of APC mRNA in malignant colonic epithelial cells: an in vitro study.

    PubMed Central

    Schnitzler, M; Dwight, T; Robinson, B G

    1996-01-01

    BACKGROUND--Sulindac is a non-steroidal anti-inflammatory drug which induces regression of colonic polyps in patients with familial adenomatous polyposis. Animal and in vitro studies have shown that both the sulphide metabolite of sulindac, which is able to inhibit cyclo-oxygenase, and the sulphone metabolite, which lacks this ability, are able to inhibit the growth of colonic carcinoma cells. The exact mechanism by which these effects occurs is not known. AIMS--To examine the effect of sulindac sulphide and sulindac sulphone on the expression of APC messenger RNA (mRNA), and on the proliferation of colonic carcinoma cells in vitro. METHODS--The colonic carcinoma cell line LIM 1215 was treated with sulindac sulphide and sulindac sulphone (10 microM or 100 microM) for 24 hours. Total RNA was extracted and APC mRNA was quantitated using competitive reverse transcription polymerase chain reaction. Measurements of cell number, cell proliferation, and prostaglandin E2 concentrations were also made. RESULTS--A significant increase in APC mRNA was observed after treatment with 10 microM of both sulindac sulphide and sulindac sulphone (control: 37.2 (19.7); 10 microM sulindac sulphide: 129 (112.8); 10 microM sulindac sulphone: 207.7 (102.9) pg/(g total RNA) (p < 0.05). Prostaglandin E2 concentrations were significantly reduced after treatment with sulindac sulphide, but not after sulindac sulphone. Both agents produced a dose dependent reduction in cell numbers and cell proliferation, which was more noticeable after treatment with sulindac sulphide. CONCLUSIONS--Both sulindac sulphide and sulindac sulphone inhibit the growth of carcinoma cells in vitro and cause an increase in APC mRNA. The effect of these agents on colonic carcinogenesis is not mediated entirely by means of an inhibition of prostaglandin biosynthesis. Images Figure 1 PMID:8707116

  14. Stabilisation/solidification of APC residues from MSW incineration with hydraulic binders and chemical additives.

    PubMed

    Quina, Margarida J; Bordado, João C M; Quinta-Ferreira, Rosa M

    2014-01-15

    This study focuses on the stabilisation/solidification (S/S) treatment of air pollution control (APC) residues from municipal solid waste (MSW) incineration. Six formulations (T1-T6) were tested based on different cements as binders, for the immobilisation of pollutants and to prevent their entering into the environment at unacceptable rates. Soluble phosphates and silicates were considered in some cases to fix heavy metals. The performance of T1-T6 products was measured in terms of initial and final setting times, mechanical strength, total availability and leaching from S/S products. Two monolithic leaching tests were used to estimate emissions of pollutants over 48h and 64 days. The results showed that the setting time was reduced when soluble phosphates were used. Moreover, although all the treatments have met the threshold of 1MPa for unconfined compressive strength, this parameter was significantly reduced due to matrix dissolution during immersion. After three cycles of leaching, the limit of 10% for solubilisation was exceeded for all treatments with the exception of T5 (with phosphates). This study demonstrated that the S/S treatment used at the industrial level can be improved with respect to toxic heavy metals, by using soluble silicates or phosphates, but not regarding soluble salts. PMID:24291664

  15. Effect of improving flue gas cleaning on characteristics and immobilisation of APC residues from MSW incineration.

    PubMed

    Geysen, D; Vandecasteele, C; Jaspers, M; Brouwers, E; Wauters, G

    2006-01-16

    The flue gas cleaning system of a MSW incinerator with a capacity of 350 kt/year was changed to improve the HCl elimination efficiency. Instead of the semi-wet operating spray reactor and subsequent baghouse, a two-step wet flue gas cleaning was added behind the baghouse. Elemental composition, X-ray powder diffraction patterns and TGA measurements showed that the resulting APC residue was totally different from the former residue. As a consequence, leaching characteristics of both residues also differed and another treatment was required prior to disposal. For the former residue, mainly leaching of Pb (>100 mg/l), necessitated treatment prior to landfilling. The lower alkalinity of the new residue resulted in a leachate pH of 9.7 and a Pb concentration of 0.8 mg/l. The leachate pH of the former residue was 12.4. The leaching of Pb and Zn increased above 100 mg/l when immobilising the new residue with cement. Better results were obtained when immobilising with micro silica. The high CaCl2 x 2H2O content of the new residue brought along clogging of the bag filter system. Adding 1.4% of CaO (or 1.9% of Ca(OH)2) to the residue already improved these inconveniences but again significantly changed the leaching behaviour of the residue. PMID:16386367

  16. Regulation of mitochondrial morphology by APC/CCdh1-mediated control of Drp1 stability

    PubMed Central

    Horn, Sarah R.; Thomenius, Michael J.; Johnson, Erika Segear; Freel, Christopher D.; Wu, Judy Q.; Coloff, Jonathan L.; Yang, Chih-Sheng; Tang, Wanli; An, Jie; Ilkayeva, Olga R.; Rathmell, Jeffrey C.; Newgard, Christopher B.; Kornbluth, Sally

    2011-01-01

    Homeostatic maintenance of cellular mitochondria requires a dynamic balance between fission and fusion, and controlled changes in morphology are important for processes such as apoptosis and cellular division. Interphase mitochondria have been described as an interconnected network that fragments as cells enter mitosis, and this mitotic mitochondrial fragmentation is known to be regulated by the dynamin-related GTPase Drp1 (dynamin-related protein 1), a key component of the mitochondrial division machinery. Loss of Drp1 function and the subsequent failure of mitochondrial division during mitosis lead to incomplete cytokinesis and the unequal distribution of mitochondria into daughter cells. During mitotic exit and interphase, the mitochondrial network reforms. Here we demonstrate that changes in mitochondrial dynamics as cells exit mitosis are driven in part through ubiquitylation of Drp1, catalyzed by the APC/CCdh1 (anaphase-promoting complex/cyclosome and its coactivator Cdh1) E3 ubiquitin ligase complex. Importantly, inhibition of Cdh1-mediated Drp1 ubiquitylation and proteasomal degradation during interphase prevents the normal G1 phase regrowth of mitochondrial networks following cell division. PMID:21325626

  17. Uncovering the role of APC-Cdh1 in generating the dynamics of S-phase onset

    PubMed Central

    Yuan, Xi; Srividhya, Jeyaraman; De Luca, Thomas; Lee, Ju-hyong E.; Pomerening, Joseph R.

    2014-01-01

    Cdh1, a coactivator of the anaphase-promoting complex (APC), is a potential tumor suppressor. Cdh1 ablation promotes precocious S-phase entry, but it was unclear how this affects DNA replication dynamics while contributing to genomic instability and tumorigenesis. We find that Cdh1 depletion causes early S-phase onset in conjunction with increase in Rb/E2F1-mediated cyclin E1 expression, but reduced levels of cyclin E1 protein promote this transition. We hypothesize that this is due to a weakened cyclin-dependent kinase inhibitor (CKI)–cyclin-dependent kinase 2 positive-feedback loop, normally generated by APC-Cdh1–mediated proteolysis of Skp2. Indeed, Cdh1 depletion increases Skp2 abundance while diminishing levels of the CKI p27. This lowers the level of cyclin E1 needed for S-phase entry and delays cyclin E1 proteolysis during S-phase progression while corresponding to slowed replication fork movement and reduced frequency of termination events. In summary, using both experimental and computational approaches, we show that APC-Cdh1 establishes a stimulus–response relationship that promotes S phase by ensuring that proper levels of p27 accumulate during G1 phase, and defects in its activation accelerate the timing of S-phase onset while prolonging its progression. PMID:24356446

  18. The I1307K APC mutation in a high-risk clinic setting: a follow-up study.

    PubMed

    Regev, M; Barzilai, S-Eisenberg; Figer, A; Zidan, J; Fidder, H H; Friedman, E

    2005-04-01

    While the I1307K APC mutation clearly confers an increased lifetime risk for colorectal cancer, there is a paucity of data on the natural history of colonic neoplasia in symptomatic and asymptomatic mutation carriers. In this study, 51 Jewish I1307K APC mutation carriers were identified in a high-risk familial cancer clinic over a 4-year period, of whom 29 (56.8%) (four males and 25 females) were successfully telephone interviewed for 0.5-5 years (mean 2.4 +/- 1.4) after initial genetic testing. Of these 29 cases, one individual was diagnosed with colon cancer at the age of 45 years, five had adenomatous polyps (mean number of polyps = 1.8), 11 had breast cancer (mean age at diagnosis 49.5 +/- 10.5 years), and 12 were asymptomatic, at the time of the testing. During the follow-up period, new colonic polyps were diagnosed in three mutation carriers, two with previously diagnosed colon cancer and polyps and only one of the asymptomatic mutation carriers, and two additional previously affected patients had new cancer diagnoses: gastric cancer and melanoma. From this descriptive study, it seems that the short-term risk for colonic polyps in I1307K APC mutation is low, primarily affecting patients with previously diagnosed colon tumors. PMID:15733272

  19. A Case of Recurrent Respiratory Papillomatosis Successfully Removed Via Endoscopic Argon Plasma Coagulation (APC) With No Evidence of Recurrence.

    PubMed

    Wong, J L; Tie, S T; Lee, J; Kannan, S K; Rashid Ali, M R; Ibrahim, A; Abdul Rahman, J A

    2014-08-01

    Recurrent respiratory papillomatosis (RRP) is a benign disease caused by the human papilloma virus (HPV), characterized by the formation of recurrent, epithelial neoplastic lesions in the airways. While benign, they can cause significant airway obstruction in some cases. Difficulties in treatment arise from the recurrent nature of the lesions despite repeated procedures. Other known procedures that result in deep tissue damage also cause unacceptable collateral damage to the underlying airway mucosa. We describe a case of recurrent papillomatosis that was successfully treated with argon plasma coagulation ( APC) when laser and electrocautery ablation had failed in the past. After the papillomatasis was treated with APC, there is no recurrence on repeat scope at 4 months and 9 months after the initial procedure. The procedure was done as a day case and there is no complication from the procedure. The property of the APC that allows it to cause only superficial thermal damage to the tissue makes it a suitable adjunct therapy to the treatment of papillomas, which are usually superficial lesions. PMID:25500852

  20. Aggregate material formulated with MSWI bottom ash and APC fly ash for use as secondary building material.

    PubMed

    del Valle-Zermeño, R; Formosa, J; Chimenos, J M; Martínez, M; Fernández, A I

    2013-03-01

    The main goal of this paper is to obtain a granular material formulated with Municipal Solid Waste Incineration (MSWI) bottom ash (BA) and air pollution control (APC) fly ash to be used as secondary building material. Previously, an optimum concrete mixture using both MSWI residues as aggregates was formulated. A compromise between the environmental behavior whilst maximizing the reuse of APC fly ash was considered and assessed. Unconfined compressive strength and abrasion resistance values were measured in order to evaluate the mechanical properties. From these results, the granular mixture was not suited for certain applications owing to the high BA/APC fly ash content and low cement percentages used to reduce the costs of the final product. Nevertheless, the leaching test performed showed that the concentrations of all heavy metals were below the limits established by the current Catalan legislation for their reutilization. Therefore, the material studied might be mainly used in embankments, where high mechanical properties are not needed and environmental safety is assured. PMID:23102641

  1. Liver fatty acid binding protein (L-Fabp) modifies intestinal fatty acid composition and adenoma formation in ApcMin/+ mice

    PubMed Central

    Dharmarajan, Sekhar; Newberry, Elizabeth P.; Montenegro, Grace; Nalbantoglu, ILKe; Davis, Victoria R.; Clanahan, Michael J.; Blanc, Valerie; Xie, Yan; Luo, Jianyang; Fleshman, James W.; Kennedy, Susan; Davidson, Nicholas O.

    2013-01-01

    Evidence suggests a relationship between dietary fat intake, obesity and colorectal cancer, implying a role for fatty acid (FA) metabolism in intestinal tumorigenesis that is incompletely understood. Liver fatty acid binding protein (L-Fabp), a dominant intestinal FA binding protein, regulates intestinal FA trafficking and metabolism and L-Fabp deletion attenuates diet-induced obesity. Here we examined whether changes in intestinal FA metabolism following L-Fabp deletion modify adenoma development in ApcMin/+ mice. Compound L-Fabp−/−ApcMin/+ mice were generated and fed a 10% fat diet balanced equally between saturated, monounsaturated and polyunsaturated fat. L-Fabp−/−ApcMin/+ mice displayed significant reductions in adenoma number and total polyp area compared to ApcMin/+controls, reflecting a significant shift in distribution toward smaller polyps. Adenomas from L-Fabp−/−ApcMin/+ mice exhibited reductions in cellular proliferation, high-grade dysplasia and nuclear β-catenin translocation. Intestinal FA content was increased in L-Fabp−/−ApcMin/+ mice and lipidomic profiling of intestinal mucosa revealed significant shifts to polyunsaturated FA species with reduced saturated FA species. L-Fabp−/−ApcMin/+mice also demonstrated corresponding changes in mRNA expression of enzymes involved in FA elongation and desaturation. Furthermore, adenomas from L-Fabp−/−ApcMin/+mice displayed significant reductions in mRNA abundance of nuclear hormone receptors involved in cellular proliferation and in enzymes involved in lipogenesis. These findings collectively implicate L-Fabp as an important genetic modifier of intestinal tumorigenesis and identify FA trafficking and metabolic compartmentalization as an important pathway linking dietary fat intake, obesity and intestinal tumor formation. PMID:23921281

  2. Loss of APC protein expressed by human colonic epithelial cells and the appearance of a specific low-molecular-weight form is associated with apoptosis in vitro.

    PubMed

    Browne, S J; Williams, A C; Hague, A; Butt, A J; Paraskeva, C

    1994-10-01

    APC (adenomatous polyposis coli) protein is differentially expressed in the normal colonic crypt and believed to be involved in colonic cell maturation. In this work we investigated whether expression of the APC protein is associated with cell death in colonic epithelial cells. We have previously reported an in vitro system to study apoptosis. Briefly, cells attached to the flask have a low frequency of apoptosis (1-3%), whereas cells that detach from the flask and float in the medium have a high proportion of apoptotic cells (36-96% depending on the cell line). The full-length 300-kDa or truncated APC protein, normally expressed by the attached cells (detected using the FE9 antibody), was found to be lost in the floating apoptotic cells in 8/11 colon tumour cell lines examined. In addition, the APC antibody FE9 detected a 90-kDa protein in the floating apoptotic cells of all cell lines investigated, which was not present in attached cells. Furthermore, loss of full-length APC and gain of the 90-kDa protein was observed in the apoptotic cells of 2 cell lines derived from other tissues: the SV40-transformed fibroblast cell line CMSV40fib and the lymphoblastoid B-cell line BJA-B. In cells repeatedly frozen and thawed, believed to induce necrotic cell death, full-length or truncated APC was also lost, though a 95-kDa protein distinct from that in apoptotic cells was observed. Specific loss of full-length or truncated APC (resulting in a 90-kDa protein in apoptotic cells but a 95-kDa protein in necrotic cells) is therefore associated with cell death. Our findings suggest a possible role for APC in cell survival. PMID:7927905

  3. The E3 Ligase APC/C-Cdh1 Is Required for Associative Fear Memory and Long-Term Potentiation in the Amygdala of Adult Mice

    ERIC Educational Resources Information Center

    Pick, Joseph E.; Malumbres, Marcos; Klann, Eric

    2013-01-01

    The anaphase promoting complex/cyclosome (APC/C) is an E3 ligase regulated by Cdh1. Beyond its role in controlling cell cycle progression, APC/C-Cdh1 has been detected in neurons and plays a role in long-lasting synaptic plasticity and long-term memory. Herein, we further examined the role of Cdh1 in synaptic plasticity and memory by generating…

  4. Environmental impact of APC residues from municipal solid waste incineration: Reuse assessment based on soil and surface water protection criteria

    SciTech Connect

    Quina, Margarida J.; Bordado, Joao C.M.; Quinta-Ferreira, Rosa M.

    2011-09-15

    Highlights: > The Dutch Building Material Decree (BMD) was used to APC residues from MSWI. > BMD is a straightforward tool to calculate expectable loads to the environment of common pollutants. > Chloride load to the environment lead to classification of building material not allowed. > At least a pre-treatment (e.g. washing) is required in order to remove soluble salts. > The stabilization with phosphates or silicates eliminate the problem of heavy metals. - Abstract: Waste management and environmental protection are mandatory requirements of modern society. In our study, air pollution control (APC) residues from municipal solid waste incinerators (MSWI) were considered as a mixture of fly ash and fine particulate solids collected in scrubbers and fabric filters. These are hazardous wastes and require treatment before landfill. Although there are a number of treatment options, it is highly recommended to find practical applications rather than just dump them in landfill sites. In general, for using a construction material, beyond technical specifications also soil and surface water criteria may be used to ensure environmental protection. The Dutch Building Materials Decree (BMD) is a valuable tool in this respect and it was used to investigate which properties do not meet the threshold criteria so that APC residues can be further used as secondary building material. To this end, some scenarios were evaluated by considering release of inorganic species from unmoulded and moulded applications. The main conclusion is that the high amount of soluble salts makes the APC residues a building material prohibited in any of the conditions tested. In case of moulding materials, the limits of heavy metals are complied, and their use in Category 1 would be allowed. However, also in this case, the soluble salts lead to the classification of 'building material not allowed'. The treatments with phosphates or silicates are able to solve the problem of heavy metals, but

  5. Absence of somatic alterations of the EB1 gene adenomatous polyposis coli-associated protein in human sporadic colorectal cancers.

    PubMed Central

    Jaïs, P.; Sabourin, J. C.; Bombled, J.; Rougier, P.; Lasser, P.; Duvillard, P.; Bénard, J.; Bressac-de Paillerets, B.

    1998-01-01

    The human EB1 gene product was recently found, by a yeast two-hybrid screening, to be associated with the carboxy terminus of the APC (adenomatous polyposis coli) protein, the product of a tumour-suppressor gene thought to act as a gatekeeper in colorectal carcinogenesis. Because virtually all of the APC mutations result in the synthesis of carboxy-terminal truncated proteins, mutant APC proteins are expected to lose their ability to interact with EB1 gene product. Thus, the interaction between APC and EB1 proteins may be important for the tumour-suppressor activity of APC protein, and raises the hypothesis that EB1 is also involved in sporadic colorectal tumorigenesis. To investigate this hypothesis, somatic mutations in the entire coding sequence of EB1 cDNA were searched by reverse transcriptase single-strand conformational polymorphism (SSCP) analysis in 21 sporadic colorectal cancers and seven adenomas. None of these tumours contained somatic mutation, whereas a silent cDNA variant was identified in 14% of alleles. Furthermore, to investigate whether EB1 locus was included within a region subjected to losses of heterozygosity, four polymorphism markers surrounding EB1 locus were surveyed. Only one out of 28 colorectal tumours contained a loss of heterozygosity at the D20S107 marker. In conclusion, the present findings strongly suggest that EB1 gene is not involved in somatic colorectal carcinogenesis. Images Figure 2 Figure 3 PMID:9823979

  6. EBV-Infection in Cardiac and Non-Cardiac Gastric Adenocarcinomas is Associated with Promoter Methylation of p16, p14 and APC, but not hMLH1

    PubMed Central

    Geddert, Helene; zur Hausen, Axel; Gabbert, Helmut E.; Sarbia, Mario

    2010-01-01

    Background: Epstein–Barr virus (EBV)-associated gastric carcinomas (GC) constitute a distinct clinicopathological entity of gastric cancer. In order to determine underlying distinct aberrant promoter methylation we tested cardiac and non-cardiac GC with regard to the presence of EBV. Methods: One hundred GC were tested by RNA-in situ hybridization for the presence of EBV by EBV-encoded small RNA (EBER). Aberrant promoter methylation was investigated by methylation-specific real-time PCR for p16, p14, APC and hMLH1. P16 protein expression was assessed by immunohistochemistry. Results: In our selected study cohort, EBER-transcripts were detected in 19.6% (18/92) of GC. EBV-positive GC revealed significantly more often gene hypermethylation of p16, p14 and APC (p < 0.0001, p < 0.0001 and p = 0.02, respectively) than EBV-negative GC. The majority of GC with p16 hypermethylation showed a p16 protein loss (22/28). In contrast, no correlation between the presence of EBV and hMLH1 hypermethylation was found (p = 0.7). EBV-positive GC showed a trend towards non-cardiac location (p = 0.06) and lower stages (I/II) according to the WHO (p = 0.05). Conclusion: Hypermethylation of tumor suppressor genes is significantly more frequent in EBV-associated GC compared to EBV-negative GC. Our data add new insights to the role of EBV in gastric carcinogenesis and underline that EBV-associated GC comprise a distinct molecular-pathologic as well as a distinct clinicopathological entity of GC. PMID:20978327

  7. A gene-centric analysis of activated partial thromboplastin time and activated protein C resistance using the HumanCVD focused genotyping array

    PubMed Central

    Gaunt, Tom R; Lowe, Gordon DO; Lawlor, Debbie A; Casas, Juan-Pablo; Day, Ian NM

    2013-01-01

    Activated partial thromboplastin time (aPTT) is an important routine measure of intrinsic blood coagulation. Addition of activated protein C (APC) to the aPTT test to produce a ratio, provides one measure of APC resistance. The associations of some genetic mutations (eg, factor V Leiden) with these measures are established, but associations of other genetic variations remain to be established. The objective of this work was to test for association between genetic variants and blood coagulation using a high-density genotyping array. Genetic association with aPTT and APC resistance was analysed using a focused genotyping array that tests approximately 50 000 single-nucleotide polymorphisms (SNPs) in nearly 2000 cardiovascular candidate genes, including coagulation pathway genes. Analyses were conducted on 2544 European origin women from the British Women's Heart and Health Study. We confirm associations with aPTT at the coagulation factor XII (F12)/G protein-coupled receptor kinase 6 (GRK6) and kininogen 1 (KNG1)/histidine-rich glycoprotein (HRG) loci, and identify novel SNPs at the ABO locus and novel locus kallikrein B (KLKB1)/F11. In addition, we confirm association between APC resistance and factor V Leiden mutation, and identify novel SNP associations with APC resistance in the HRG and F5/solute carrier family 19 member 2 (SLC19A2) regions. In conclusion, variation at several genetic loci influences intrinsic blood coagulation as measured by both aPTT and APC resistance. PMID:23188048

  8. Asymmetric 1-Alkyl-2-acyl Phosphatidylcholine: a Helper Lipid for Enhanced Non-viral Gene Delivery

    PubMed Central

    Huang, Zhaohua; Li, Weijun; Szoka, Francis C.

    2011-01-01

    Rationally designed asymmetrical alkylacyl phosphatidylcholines (APC) have been synthesized and evaluated as helper lipids for non-viral gene delivery. A long aliphatic chain (C22~C24) was introduced at the 1-position of glycerol backbone, a branched lipid chain (C18) at the 2-position, and a phosphocholine head group at the 3-position. The fusogenicity of APC depends on the length and degree of saturation of the alkyl chain. Cationic lipids were formulated with APC as either lipoplexes or nanolipoparticles, and evaluated for their stability, transfection efficiency, and cytotoxicity. APC mediated high in vitro transfection efficiency, and had low cytotoxicity. Small nanolipoparticles (less than 100 nm) can be obtained with APC by applying as low as 0.1% PEG-lipid. Our study extends the type of helper lipids that are suitable for gene transfer and points the way to improve non-viral nucleic acid delivery system other than the traditional cationic lipids optimization. This work is supported by NIH grant EB003008. PMID:21718766

  9. Gender-specific modulation of tumorigenesis by folic acid supply in the Apc mouse during early neonatal life.

    PubMed

    McKay, Jill A; Williams, Elizabeth A; Mathers, John C

    2008-03-01

    Epidemiological studies suggest an inverse association between folic acid intake and colorectal cancer risk. Conversely, conventional treatment of existing tumours includes the use of folate antagonists. This suggests that the level of exposure to folate and its timing in relation to stage of tumorigenesis may be critical in determining outcomes. We hypothesised that folic acid depletion in utero and during early neonatal life may affect tumorigenesis in offspring. To investigate this hypothesis, female C57Bl6/J mice were randomised to a folic acid adequate (2 mg folic acid/kg diet) or folic acid depleted diet (0.4 mg folic acid/kg) from mating with Apc+/Min sires and throughout pregnancy and lactation. At weaning the Apc+/Min offspring were randomised to a folic acid adequate (2 mg folic acid/kg diet) or depleted (0.26 mg folic acid/kg diet) diet, creating four in utero/post-weaning dietary regimens. At 10 weeks post-weaning, mice were killed and the intestinal tumour number and size were recorded. Folic acid depletion during pregnancy and post-weaning reduced erythrocyte folate concentrations in offspring significantly. Folic acid depletion during pregnancy and lactation did not affect tumour multiplicity or size. However, female mice fed normal folic acid diets post-weaning had more, and larger, tumours when compared with depleted females and both depleted and adequate folic acid fed males. These data suggest that folate depletion post-weaning was protective against neoplasia in female Apc+/Min mice and highlights the need for further investigation of the optimal timing and dose of folic acid supplementation with regard to colorectal cancer risk. PMID:17868491

  10. Leaching of APC residues from secondary Pb metallurgy using single extraction tests: the mineralogical and the geochemical approach.

    PubMed

    Ettler, Vojtech; Mihaljevic, Martin; Sebek, Ondrej; Strnad, Ladislav

    2005-05-20

    Two air-pollution-control (APC) residues--one from flue gas cooling with alkaline water and one from deionized water cooling--from secondary lead metallurgy were submitted to two different standardized short-term leaching protocols: US EPA toxicity characteristic leaching procedure (TCLP) and static leaching according to Czech/European norm EN 12457-2. The experimental procedure was coupled with detailed mineralogical investigation of the solid material (SEM, XRPD) and speciation-solubility calculations using the PHREEQC-2 geochemical code. Both types of residues were considered as hazardous materials exhibiting substantial leaching of Pb (up to 7130 mg/l) and other inorganic contaminants. However, the APC residue produced by flue gas cooling with alkaline water (sample B) exhibits more favourable leaching and environmental characteristics than that produced by simple deionised water cooling (sample A). At pH < 5, primary caracolite (Na3Pb2(SO4)3Cl) and potassium lead chloride (KCl.2PbCl2) are completely or partially dissolved and transformed to residual anglesite (PbSO4), cotunnite (PbCl2) and laurionite (Pb(OH)Cl). At pH 5-6, anglesite is still the principal residual product, whereas at pH > 6, phosgenite (PbCl2.PbCO3) became the dominant secondary phase. The results are consistent with the mineralogical and geochemical studies focused on acidic forest soils highly polluted by smelter emissions, where anglesite, as a unique Pb-bearing phase, has been detected. From the technological point of view, the mixing of APC residue with alkaline water, followed by an increase in the suspension pH and equilibration with atmospheric CO2, may be used to ensure the precipitation of less soluble Pb carbonates, which are more easily recycled in the Pb recovery process in the metallurgical plant. PMID:15885416

  11. The regulation of skeletal muscle protein turnover during the progression of cancer cachexia in the Apc(Min/+) mouse.

    PubMed

    White, James P; Baynes, John W; Welle, Stephen L; Kostek, Matthew C; Matesic, Lydia E; Sato, Shuichi; Carson, James A

    2011-01-01

    Muscle wasting that occurs with cancer cachexia is caused by an imbalance in the rates of muscle protein synthesis and degradation. The Apc(Min/+) mouse is a model of colorectal cancer that develops cachexia that is dependent on circulating IL-6. However, the IL-6 regulation of muscle protein turnover during the initiation and progression of cachexia in the Apc(Min/+) mouse is not known. Cachexia progression was studied in Apc(Min/+) mice that were either weight stable (WS) or had initial (≤5%), intermediate (6-19%), or extreme (≥20%) body weight loss. The initiation of cachexia reduced %MPS 19% and a further ∼50% with additional weight loss. Muscle IGF-1 mRNA expression and mTOR targets were suppressed with the progression of body weight loss, while muscle AMPK phosphorylation (Thr 172), AMPK activity, and raptor phosphorylation (Ser 792) were not increased with the initiation of weight loss, but were induced as cachexia progressed. ATP dependent protein degradation increased during the initiation and progression of cachexia. However, ATP independent protein degradation was not increased until cachexia had progressed beyond the initial phase. IL-6 receptor antibody administration prevented body weight loss and suppressed muscle protein degradation, without any effect on muscle %MPS or IGF-1 associated signaling. In summary, the %MPS reduction during the initiation of cachexia is associated with IGF-1/mTOR signaling repression, while muscle AMPK activation and activation of ATP independent protein degradation occur later in the progression of cachexia. IL-6 receptor antibody treatment blocked cachexia progression through the suppression of muscle protein degradation, while not rescuing the suppression of muscle protein synthesis. Attenuation of IL-6 signaling was effective in blocking the progression of cachexia, but not sufficient to reverse the process. PMID:21949739

  12. Sex-dependent Differences in Intestinal Tumorigenesis Induced in Apc1638N/+ Mice by Exposure to {gamma} Rays

    SciTech Connect

    Trani, Daniela; Moon, Bo-Hyun; Kallakury, Bhaskar; Hartmann, Dan P.; Datta, Kamal; Fornace, Albert J.

    2013-01-01

    Purpose: The purpose of the present study was to assess the effect of 1 and 5 Gy radiation doses and to investigate the interplay of gender and radiation with regard to intestinal tumorigenesis in an adenomatous polyposis coli (APC) mutant mouse model. Methods and Materials: Apc1638N/+ female and male mice were exposed whole body to either 1 Gy or 5 Gy of {gamma} rays and euthanized when most of the treated mice became moribund. Small and large intestines were processed to determine tumor burden, distribution, and grade. Expression of proliferation marker Ki-67 and estrogen receptor (ER)-{alpha} were also assessed by immunohistochemistry. Results: We observed that, with both 1 Gy and 5 Gy of {gamma} rays, females displayed reduced susceptibility to radiation-induced intestinal tumorigenesis compared with males. As for radiation effect on small intestinal tumor progression, although no substantial differences were found in the relative frequency and degree of dysplasia of adenomas in irradiated animals compared with controls, invasive carcinomas were found in 1-Gy- and 5-Gy-irradiated animals. Radiation exposure was also shown to induce an increase in protein levels of proliferation marker Ki-67 and sex-hormone receptor ER-{alpha} in both non tumor mucosa and intestinal tumors from irradiated male mice. Conclusions: We observed important sex-dependent differences in susceptibility to radiation-induced intestinal tumorigenesis in Apc1638N/+ mutants. Furthermore, our data provide evidence that exposure to radiation doses as low as 1 Gy can induce a significant increase in intestinal tumor multiplicity as well as enhance tumor progression in vivo.

  13. Mechanistic study of the anti-cancer effect of Gynostemma pentaphyllum saponins in the Apc(Min/+) mouse model.

    PubMed

    Tai, William Chi-Shing; Wong, Wing-Yan; Lee, Magnolia Muk-Lan; Chan, Brandon Dow; Lu, Cheng; Hsiao, Wen-Luan Wendy

    2016-05-01

    Gynostemma pentaphyllum saponins (GpS) have been shown to have anti-cancer activity. However, the underlying mechanisms remain unclear. In this study, we used the Apc(Min) (/+) colorectal cancer (CRC) mouse model to investigate the anti-cancer effect of GpS and we demonstrated that GpS treatment could significantly reduce the number and size of intestinal polyps in Apc(Min) (/+) mice. In order to identify the potential targets and mechanisms involved, a comparative proteomics analysis was performed and 40 differentially expressed proteins after GpS treatment were identified. Bioinformatics analyses suggested a majority of these proteins were involved in processes related to cellular redox homeostasis, and predicted Raf-1 as a potential target of GpS. The upregulation of two proteins known to be involved in redox homeostasis, peroxiredoxin-1 (Prdx1) and peroxiredoxin-2 (Prdx2), and the downregulation of Raf-1 were validated using Western blot analysis. After further investigation of the associated signaling networks, we postulated that the anti-cancer effect of GpS was mediated through the upregulation of Prdx1 and Prdx2, suppression of Ras, RAF/MEK/ERK/STAT, PI3K/AKT/mTOR signaling and modulation of JNK/p38 MAPK signaling. We also examined the potential combinatorial effect of GpS with the chemotherapeutic 5-fluorouracil (5-FU) and found that GpS could enhance the anti-cancer efficacy of 5-FU, further suppressing the number of polyps in Apc(Min/+) mice. Our findings highlight the potential of GpS as an anti-cancer agent, the potential mechanisms of its anti-cancer activities, and its effect as an adjuvant of 5-FU in the chemotherapy of CRC. PMID:26970558

  14. Source apportionment of gaseous atmospheric pollutants by means of an absolute principal component scores (APCS) receptor model.

    PubMed

    Bruno, P; Caselli, M; de Gennaro, G; Traini, A

    2001-12-01

    A multivariate statistical method has been applied to apportion the atmospheric pollutant concentrations measured by automatic gas analyzers placed on a mobile laboratory for air quality monitoring in Taranto (Italy). In particular, Principal Component Analysis (PCA) followed by Absolute Principal Component Scores (APCS) technique was performed to identify the number of emission sources and their contribution to measured concentrations of CO, NOx, benzene toluene m+p-Xylene (BTX). This procedure singled out two different sources that explain about 85% of collected data variance. PMID:11798109

  15. A Cdh1-APC/FMRP Ubiquitin Signaling Link Drives mGluR-Dependent Synaptic Plasticity in the Mammalian Brain

    PubMed Central

    Huang, Ju; Ikeuchi, Yoshiho; Malumbres, Marcos; Bonni, Azad

    2015-01-01

    SUMMARY Deregulation of synaptic plasticity may contribute to the pathogenesis of developmental cognitive disorders. In particular, exaggerated mGluR-dependent LTD is featured in fragile X syndrome, but the mechanisms that regulate mGluR-LTD remain incompletely understood. We report that conditional knockout of Cdh1, the key regulatory subunit of the ubiquitin ligase Cdh1-anaphase promoting complex (Cdh1-APC), profoundly impairs mGluR-LTD in the hippocampus. Mechanistically, we find that Cdh1-APC operates in the cytoplasm to drive mGluR-LTD. We also identify the fragile X syndrome protein FMRP as a substrate of Cdh1-APC. Endogenous Cdh1-APC forms a complex with endogenous FMRP, and knockout of Cdh1 impairs mGluR-induced ubiquitination and degradation of FMRP in the hippocampus. Knockout of FMRP suppresses, and expression of an FMRP mutant protein that fails to interact with Cdh1 phenocopies, the Cdh1 knockout phenotype of impaired mGluR-LTD. These findings define Cdh1-APC and FMRP as components of a novel ubiquitin-signaling pathway that regulates mGluR-LTD in the brain. PMID:25913861

  16. APC/CCdh1 Targets Brain-Specific Kinase 2 (BRSK2) for Degradation via the Ubiquitin-Proteasome Pathway

    PubMed Central

    Zhou, Jun; Wang, Yingli; Luo, Ting; Gu, Xiuting; Chen, Fang; Yu, Long

    2012-01-01

    Studies of brain-specific kinase 2 (BRSK2), an AMP-activated protein kinase (AMPK)-related kinase, and its homologs suggest that they are multifunctional regulators of cell-cycle progression. BRSK2, which contains a ubiquitin-associated (UBA) domain, is polyubiquitinated in cells. However, the regulatory mechanisms and exact biological function of BRSK2 remain unclear. Herein, we show that BRSK2 co-localizes with the centrosomes during mitosis. We also demonstrate that BRSK2 protein levels fluctuate during the cell cycle, peaking during mitosis and declining in G1 phase. Furthermore, Cdh1, rather than Cdc20, promotes the degradation of BRSK2 in vivo. Consistent with this finding, knock-down of endogenous Cdh1 blocks BRSK2 degradation during the G1 phase. The conserved KEN box of BRSK2 is required for anaphase-promoting complex/cyclosome-Cdh1 (APC/CCdh1)-dependent degradation. Additionally, overexpression of either BRSK2(WT) or BRSK2(ΔKEN) increases the percentage of cells in G2/M. Thus, our results provide the first evidence that BRSK2 regulates cell-cycle progression controlled by APC/CCdh1 through the ubiquitin-proteasome pathway. PMID:23029325

  17. Reducing overlay sampling for APC-based correction per exposure by replacing measured data with computational prediction

    NASA Astrophysics Data System (ADS)

    Noyes, Ben F.; Mokaberi, Babak; Oh, Jong Hun; Kim, Hyun Sik; Sung, Jun Ha; Kea, Marc

    2016-03-01

    One of the keys to successful mass production of sub-20nm nodes in the semiconductor industry is the development of an overlay correction strategy that can meet specifications, reduce the number of layers that require dedicated chuck overlay, and minimize measurement time. Three important aspects of this strategy are: correction per exposure (CPE), integrated metrology (IM), and the prioritization of automated correction over manual subrecipes. The first and third aspects are accomplished through an APC system that uses measurements from production lots to generate CPE corrections that are dynamically applied to future lots. The drawback of this method is that production overlay sampling must be extremely high in order to provide the system with enough data to generate CPE. That drawback makes IM particularly difficult because of the throughput impact that can be created on expensive bottleneck photolithography process tools. The goal is to realize the cycle time and feedback benefits of IM coupled with the enhanced overlay correction capability of automated CPE without impacting process tool throughput. This paper will discuss the development of a system that sends measured data with reduced sampling via an optimized layout to the exposure tool's computational modelling platform to predict and create "upsampled" overlay data in a customizable output layout that is compatible with the fab user CPE APC system. The result is dynamic CPE without the burden of extensive measurement time, which leads to increased utilization of IM.

  18. B lymphoblastoid cell lines as efficient APC to elicit CD8+ T cell responses against a cytomegalovirus antigen.

    PubMed

    Sun, Q; Burton, R L; Dai, L J; Britt, W J; Lucas, K G

    2000-10-01

    Potent and readily accessible APC are critical for development of immunotherapy protocols to treat viral disease and cancer. We have shown that B lymphoblastoid cell lines (BLCL) that stably express CMV phosphoprotein 65 (BLCLpp65), as a result of retroviral transduction, can be used to generate ex vivo CTL cultures that possess cytotoxicity against CMV and EBV. In this report, we demonstrate that the EBV-specific cytotoxicity in the BLCLpp65-primed culture had a spectrum of EBV-Ag recognition similar to that of the BLCL-primed counterpart, suggesting that retroviral transduction and expression of the CMV Ag would not compromise the Ag-presenting capacity of BLCL. In addition, BLCLpp65 appeared to present multiple natural pp65 epitopes, because pp65-specific CTL, which recognized different CMV clinical isolates, were generated in BLCLpp65-primed cultures from individuals with various HLA backgrounds. Consistent with a polyclonal expansion of virus-specific CTL, T cell lines established from the BLCLpp65-primed CTL cultures expressed different TCR-Vbeta Although most of the virus-specific T cell isolates were CD8+, EBV-specific CD4+ lines were also established from BLCLpp65-primed cultures. Western blot analysis revealed that the CD8+ lines, but not the CD4+ line, expressed granzyme B, consistent with features of classic CTL. Thus, our results suggested that BLCL stably expressing a foreign Ag might be used as a practical APC to elicit CD8+ T cell responses. PMID:11034422

  19. Metformin Suppresses MHC-Restricted Antigen Presentation by Inhibiting Co-Stimulatory Factors and MHC Molecules in APCs

    PubMed Central

    Shin, Seulmee; Hyun, Bobae; Lee, Aeri; Kong, Hyunseok; Han, Shinha; Lee, Chong-Kil; Ha, Nam-Joo; Kim, Kyungjae

    2013-01-01

    Metformin is widely used for T2D therapy but its cellular mechanism of action is undefined. Recent studies on the mechanism of metformin in T2D have demonstrated involvement of the immune system. Current immunotherapies focus on the potential of immunomodulatory strategies for the treatment of T2D. In this study, we examined the effects of metformin on the antigen-presenting function of antigen-presenting cells (APCs). Metformin decreased both MHC class I and class II-restricted presentation of OVA and suppressed the expression of both MHC molecules and co-stimulatory factors such as CD54, CD80, and CD86 in DCs, but did not affect the phagocytic activity toward exogenous OVA. The class II-restricted OVA presentation-regulating activity of metformin was also confirmed using mice that had been injected with metformin followed by soluble OVA. These results provide an understanding of the mechanisms of the T cell response-regulating activity of metformin through the inhibition of MHC-restricted antigen presentation in relation to its actions on APCs. PMID:24009856

  20. MASTL(Greatwall) regulates DNA damage responses by coordinating mitotic entry after checkpoint recovery and APC/C activation

    PubMed Central

    Wong, Po Yee; Ma, Hoi Tang; Lee, Hyun-jung; Poon, Randy Y. C.

    2016-01-01

    The G2 DNA damage checkpoint is one of the most important mechanisms controlling G2–mitosis transition. The kinase Greatwall (MASTL in human) promotes normal G2–mitosis transition by inhibiting PP2A via ARPP19 and ENSA. In this study, we demonstrate that MASTL is critical for maintaining genome integrity after DNA damage. Although MASTL did not affect the activation of DNA damage responses and subsequent repair, it determined the timing of entry into mitosis and the subsequent fate of the recovering cells. Constitutively active MASTL promoted dephosphorylation of CDK1Tyr15 and accelerated mitotic entry after DNA damage. Conversely, downregulation of MASTL or ARPP19/ENSA delayed mitotic entry. Remarkably, APC/C was activated precociously, resulting in the damaged cells progressing from G2 directly to G1 and skipping mitosis all together. Collectively, these results established that precise control of MASTL is essential to couple DNA damage to mitosis through the rate of mitotic entry and APC/C activation. PMID:26923777

  1. PP2A delays APC/C-dependent degradation of separase-associated but not free securin

    PubMed Central

    Hellmuth, Susanne; Böttger, Franziska; Pan, Cuiping; Mann, Matthias; Stemmann, Olaf

    2014-01-01

    The universal triggering event of eukaryotic chromosome segregation is cleavage of centromeric cohesin by separase. Prior to anaphase, most separase is kept inactive by association with securin. Protein phosphatase 2A (PP2A) constitutes another binding partner of human separase, but the functional relevance of this interaction has remained enigmatic. We demonstrate that PP2A stabilizes separase-associated securin by dephosphorylation, while phosphorylation of free securin enhances its polyubiquitylation by the ubiquitin ligase APC/C and proteasomal degradation. Changing PP2A substrate phosphorylation sites to alanines slows degradation of free securin, delays separase activation, lengthens early anaphase, and results in anaphase bridges and DNA damage. In contrast, separase-associated securin is destabilized by introduction of phosphorylation-mimetic aspartates or extinction of separase-associated PP2A activity. G2- or prometaphase-arrested cells suffer from unscheduled activation of separase when endogenous securin is replaced by aspartate-mutant securin. Thus, PP2A-dependent stabilization of separase-associated securin prevents precocious activation of separase during checkpoint-mediated arrests with basal APC/C activity and increases the abruptness and fidelity of sister chromatid separation in anaphase. PMID:24781523

  2. Role of RIS/APC for manufacturing RFG/LSD. [Refinery Information Systems/Advanced Process Control, ReFormulated Gasoline/Low Sulfur Diesels

    SciTech Connect

    Latour, P.R. )

    1994-01-01

    Revolutionary changes in quality specifications (number, complexity, uncertainty, economic sensitivity) for reformulated gasolines (RFG) and low-sulfur diesels (LSD) are being addressed by powerful, new, computer-integrated manufacturing technology for Refinery Information Systems and Advanced Process Control (RIS/APC). This paper shows how the five active RIS/APC functions: performance measurement, optimization, scheduling, control and integration are used to manufacture new, clean fuels competitively. With current industry spending for this field averaging 2 to 3 cents/bbl crude, many refineries can capture 50 to 100 cents/bbl if the technology is properly employed and sustained throughout refining operations, organizations, and businesses.

  3. Estimation of sediment friction coefficient from heating upon APC penetration during the IODP NanTroSEIZE

    NASA Astrophysics Data System (ADS)

    Kinoshita, M.; Kawamura, K.; Lin, W.

    2015-12-01

    During the Nankai Trough Seismogenic Zone Experiments (NanTroSEIZE) of the Integrated Ocean Drilling Program (IODP), the advanced piston corer temperature (APC-T) tool was used to determine in situ formation temperatures while piston coring down to ~200 m below sea floor. When the corer is fired into the formation, temperature around the shoe abruptly increases due to the frictional heating. The temperature rise due to the frictional heat at the time of penetration is 10 K or larger. We found that the frictional temperature rise (=maximum temperature) increases with increasing depth, and that its intersection at the seafloor seems non-zero. Frictional heat energy is proportional to the maximum temperature rise, which is confirmed by a FEM numerical simulation of 2D cylindrical system. Here we use the result of numerical simulation to convert the observed temperature rise into the frictional heat energy. The frictional heat energy is represented as the product of the shooting length D and the shear stress (τ) between the pipe and the sediment. Assuming a coulomb slip regime, the shear stress is shows as: τ= τ0 + μ*(Sv-Pp), where τ0 is the cohesive stress, μ the dynamic frictional coefficient between the pipe and the sediment, Sv the normal stress at the pipe, and Pp the pore pressure. This can explain the non-zero intersection as well as depth-dependent increase for the frictional heating observed in the APC-T data. Assuming a hydrostatic state and by using the downhole bulk density data, we estimated the friction coefficient for each APC-T measurement. For comparison, we used the vane-shear strength measured on core samples to estimate the friction coefficients. The frictional coefficients μ were estimated as ranging 0.01 - 0.06, anomalously lower than expected for shallow marine sediments. They were lower than those estimated from vane-shear data, which range 0.05 to 0.2. Still, both estimates exhibit a significant increase in the friction coefficient at

  4. The influence of pH on the leaching behaviour of inorganic components from municipal solid waste APC residues

    SciTech Connect

    Quina, Margarida J. Bordado, Joao C.M.; Quinta-Ferreira, Rosa M.

    2009-09-15

    The influence of pH on the leaching behaviour of air pollution control (APC) residues produced in municipal solid waste incineration (MSWI) is addressed in this study. The residue is considered hazardous waste, and in accordance with their chemical properties, the leaching of contaminants into the environment is the main concern. Several leaching tests can be used for research studies or regulatory purposes, where a wide variety of conditions may be tested. Our work deals mainly with the leaching behaviour of toxic heavy metals (Pb, Cd, Zn, Cr, Ni, Cu) and inorganics associated with soluble salts (Na, K, Ca, Cl). The main goal is to obtain an overview of the leachability of APC residues produced in a Portuguese MSWI process. Among the different variables that may have influence on the leaching behaviour, pH of the leachant solution is the most important one, and was evaluated through pH static tests. The acid neutralization capacity (ANC) of the residue was also determined, which is in the range of 6.2-6.8 meq g{sup -1} (for pH = 7) and 10.1-11.6 meq g{sup -1} (for pH = 4). The analysis of the leaching behaviour is particularly important when the leaching is solubility controlled. The amphoteric behaviour of some elements was observed, namely for Pb and Zn, which is characterized through high solubilization at low and high pH and moderate or low solubility at neutral or moderate high pH. The solubility curves for Pb, Cd, Zn, Cr, Ni and Cu as a function of pH were obtained, which are very useful for predicting the leaching behaviour in different scenarios. The solubility of K and Na reveals to be nearly independent of the solution pH and the released amount is mainly availability-controlled. Moreover, the pH static test showed that Cl{sup -} is the most pH-independent species. The APC residue turns out to be a hazardous waste because of the high leaching of lead and chloride. On the other hand, leaching of elements like cadmium, nickel and copper is limited by the

  5. The polyglutamine-expanded androgen receptor responsible for spinal and bulbar muscular atrophy inhibits the APC/CCdh1 ubiquitin ligase complex

    PubMed Central

    Bott, Laura C.; Salomons, Florian A.; Maric, Dragan; Liu, Yuhong; Merry, Diane; Fischbeck, Kenneth H.; Dantuma, Nico P.

    2016-01-01

    Polyglutamine expansion in the androgen receptor (AR) causes spinal and bulbar muscular atrophy (SBMA), an X-linked neuromuscular disease that is fully manifest only in males. It has been suggested that proteins with expanded polyglutamine tracts impair ubiquitin-dependent proteolysis due to their propensity to aggregate, but recent studies indicate that the overall activity of the ubiquitin-proteasome system is preserved in SBMA models. Here we report that AR selectively interferes with the function of the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C), which, together with its substrate adaptor Cdh1, is critical for cell cycle arrest and neuronal architecture. We show that both wild-type and mutant AR physically interact with the APC/CCdh1 complex in a ligand-dependent fashion without being targeted for proteasomal degradation. Inhibition of APC/CCdh1 by mutant but not wild-type AR in PC12 cells results in enhanced neurite outgrowth which is typically followed by rapid neurite retraction and mitotic entry. Our data indicate a role of AR in neuronal differentiation through regulation of APC/CCdh1 and suggest abnormal cell cycle reactivation as a pathogenic mechanism in SBMA. PMID:27312068

  6. The SnRK2-APC/CTE regulatory module mediates the antagonistic action of gibberellic acid and abscisic acid pathways

    PubMed Central

    Lin, Qibing; Wu, Fuqing; Sheng, Peike; Zhang, Zhe; Zhang, Xin; Guo, Xiuping; Wang, Jiulin; Cheng, Zhijun; Wang, Jie; Wang, Haiyang; Wan, Jianmin

    2015-01-01

    Abscisic acid (ABA) and gibberellic acid (GA) antagonistically regulate many developmental processes and responses to biotic or abiotic stresses in higher plants. However, the molecular mechanism underlying this antagonism is still poorly understood. Here, we show that loss-of-function mutation in rice Tiller Enhancer (TE), an activator of the APC/CTE complex, causes hypersensitivity and hyposensitivity to ABA and GA, respectively. We find that TE physically interacts with ABA receptor OsPYL/RCARs and promotes their degradation by the proteasome. Genetic analysis also shows OsPYL/RCARs act downstream of TE in mediating ABA responses. Conversely, ABA inhibits APC/CTE activity by phosphorylating TE through activating the SNF1-related protein kinases (SnRK2s), which may interrupt the interaction between TE and OsPYL/RCARs and subsequently stabilize OsPYL/RCARs. In contrast, GA can reduce the level of SnRK2s and may promote APC/CTE-mediated degradation of OsPYL/RCARs. Thus, we propose that the SnRK2-APC/CTE regulatory module represents a regulatory hub underlying the antagonistic action of GA and ABA in plants. PMID:26272249

  7. Aβ Induces Excitotoxicity Mediated by APC/C-Cdh1 Depletion That Can Be Prevented by Glutaminase Inhibition Promoting Neuronal Survival

    PubMed Central

    Fuchsberger, T.; Martínez-Bellver, S.; Giraldo, E.; Teruel-Martí, V.; Lloret, A.; Viña, J.

    2016-01-01

    The E3 ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C) is activated by the fizzy-related protein homolog/CDC20-like protein 1 (cdh1) in post-mitotic neurons. Growing evidence suggests that dysregulation of APC/C-Cdh1 is involved in neurodegenerative diseases. Here we show in neurons that oligomers of amyloid beta (Aβ), a peptide related to Alzheimer’s disease, cause proteasome-dependent degradation of cdh1. This leads to a subsequent increase in glutaminase (a degradation target of APC/C-Cdh1), which causes an elevation of glutamate levels and further intraneuronal Ca2+ dysregulation, resulting in neuronal apoptosis. Glutaminase inhibition prevents glutamate excitotoxicity and apoptosis in Aβ treated neurons. Furthermore, glutamate also decreases cdh1 and leads to accumulation of glutaminase, suggesting that there may be a positive feedback loop of cdh1 inactivation. We confirmed the main findings in vivo using microinjection of either Aβ or glutamate in the CA1 region of the rat hippocampus. We show here for the first time in vivo that both Aβ and glutamate cause nuclear exclusion of cdh1 and an increase in glutaminase. These results show that maintaining normal APC/C-Cdh1 activity may be a useful target in Alzheimer’s disease treatment. PMID:27514492

  8. Inducible loss of one Apc allele in Lrig1-expressing progenitor cells results in multiple distal colonic tumors with features of familial adenomatous polyposis

    PubMed Central

    Powell, Anne E.; Vlacich, Gregory; Zhao, Zhen-Yang; McKinley, Eliot T.; Washington, M. Kay; Manning, H. Charles

    2014-01-01

    Individuals with familial adenomatous polyposis (FAP) harbor a germline mutation in adenomatous polyposis coli (APC). The major clinical manifestation is development of multiple colonic tumors at a young age due to stochastic loss of the remaining APC allele. Extracolonic features, including periampullary tumors, gastric abnormalities, and congenital hypertrophy of the retinal pigment epithelium, may occur. The objective of this study was to develop a mouse model that simulates these features of FAP. We combined our Lrig1-CreERT2/+ mice with Apcfl/+ mice, eliminated one copy of Apc in leucine-rich repeats and immunoglobulin-like domains protein 1 (Lrig1)-positive (Lrig1+) progenitor cells with tamoxifen injection, and monitored tumor formation in the colon by colonoscopy and PET. Initial loss of one Apc allele in Lrig1+ cells results in a predictable pattern of preneoplastic changes, culminating in multiple distal colonic tumors within 50 days of induction, as well as the extracolonic manifestations of FAP mentioned above. We show that tumor formation can be monitored by noninvasive PET imaging. This inducible stem cell-driven model recapitulates features of FAP and offers a tractable platform on which therapeutic interventions can be monitored over time by colonoscopy and noninvasive imaging. PMID:24833705

  9. Metals accumulations during thermal processing of sewage sludge - characterization of bottom ash and air pollution control (APC) residues

    NASA Astrophysics Data System (ADS)

    Kasina, Monika; Kowalski, Piotr R.; Michalik, Marek

    2016-04-01

    Due to increasing mass of sewage sludge, problems in its management have appeared. Over years sewage sludge was landfilled, however due to EU directives concerning environmental issues this option is no longer possible. This type of material is considered hazardous due to highly concentrated metals and harmful elements, toxic organic substances and biological components (e.g. parasites, microbes). Currently in Europe, incineration is considered to be the most reasonable method for sewage sludge treatment. As a result of sludge incineration significant amount of energy is recovered due to high calorific value of sewage sludge but bottom ash and APC residues are being produced. In this study we show the preliminary results of chemical and mineral analyses of both bottom ash and APC residues produced in fluidized bed boiler in sewage sludge incineration plant in Poland, with a special emphasis on metals which, as a part of incombustible fraction can accumulate in the residual materials after thermal processing. The bottom ash was a SiO2-P2O5-Fe2O3-CaO-Al2O3 dominated material. Main mineral phases identified in X-ray diffraction patterns were: quartz, feldspar, hematite, and phosphates (apatite and scholzite). The bottom ash was characterized by high content of Zn - 4472 mg kg‑1, Cu - 665.5 mg kg‑1, Pb - 138 mg kg‑1, Ni - 119.5 mg kg‑1, and interestingly high content of Au - 0.858 mg kg‑1 The APC residues composition was dominated by soluble phases which represent more than 90% of the material. The XRD patterns indicated thenardite, halite, anhydrite, calcite and apatite as main mineral phases. The removal of soluble phases by dissolution in deionised water caused a significant mass reduction (ca. 3% of material remained on the filters). Calcite, apatite and quartz were main identified phases. The content of metals in insoluble material is relatively high: Zn - 6326 mg kg‑1, Pb - 514.3 mg kg‑1, Cu - 476.6 mg kg‑1, Ni - 43.3 mg kg‑1. The content of Cd

  10. Role of microRNAs in Resveratrol-Mediated Mitigation of Colitis-Associated Tumorigenesis in ApcMin/+ Mice

    PubMed Central

    Altamemi, Ibrahim; Murphy, E. Angela; Catroppo, James F.; Zumbrun, Elizabeth E.; Zhang, Jiajia; McClellan, Jamie L.; Singh, Udai P.; Nagarkatti, Prakash S.

    2014-01-01

    The pleiotropic effects of resveratrol include anti-inflammatory, antioxidant, and anticancer activities, and thus unique possibilities exist to explore mechanistic pathways of chemoprevention. The aim of this study was to investigate the role of microRNA (miRNA) alterations induced by resveratrol in the context of chemopreventive mechanisms against dextran sodium sulfate (DSS)–induced colitis-associated tumorigenesis in the ApcMin/+ mouse. To that end, ApcMin/+ mice were exposed to 2% DSS to enhance intestinal inflammation and polyp development. Concurrently, mice received either vehicle or resveratrol treatment via oral gavage for 5 weeks. Interestingly, treatment of DSS-exposed mice with resveratrol resulted in decreased number and size of polyps, fewer histologic signs of cell damage, and decreased proliferating epithelial cells in intestinal mucosa compared with vehicle. Resveratrol treatment dramatically reversed the effects of DSS on the numbers of specific inflammatory CD4+ T cells, CD8+ T cells, B cells, natural killer T cells, and myeloid-derived suppressor cells in mesenteric lymph nodes. Resveratrol treatment also decreased interleukin-6 (IL-6) and tumor necrosis factor-α protein levels and reduced IL-6 and cyclooxygenase-2 mRNA expression. Microarray analysis revealed 104 miRNAs exhibiting >1.5-fold differences in expression in the intestinal tissue of resveratrol-treated mice. Among them, two miRNAs with anti-inflammatory properties, miRNA-101b and miRNA-455, were validated to be upregulated with resveratrol treatment by reverse-transcription polymerase chain reaction. Pathway analysis revealed that numerous differentially regulated miRNAs targeted mRNAs associated with inflammatory processes with known roles in intestinal tumorigenesis. These results suggest that resveratrol mediates anti-inflammatory properties and suppresses intestinal tumorigenesis through miRNA modulation. PMID:24817032

  11. Palmatine from Mahonia bealei attenuates gut tumorigenesis in ApcMin/+ mice via inhibition of inflammatory cytokines

    PubMed Central

    MA, WEI-KUN; LI, HUI; DONG, CUI-LAN; HE, XIN; GUO, CHANG-RUN; ZHANG, CHUN-FENG; YU, CHUN-HAO; WANG, CHONG-ZHI; YUAN, CHUN-SU

    2016-01-01

    Mahonia bealei is a Chinese folk medicine used to treat various ailments, in particular gastrointestinal inflammation-related illnesses, and palmatine is one of its active constituents. In this study, ApcMin/+ mice, a genetically engineered model, were used to investigate the effects of palmatine on the initiation and progression of gut inflammation and tumorigenesis enhanced by a high-fat diet. The in vitro antiproliferation and anti-inflammation effects of palmatine were evaluated on HT-29 and SW-480 human colorectal cancer cell lines. The concentration-related antiproliferative effects of palmatine on both cell lines (P<0.01) were observed. Palmatine significantly inhibited lipopolysaccharide-induced increase in cytokine interleukin (IL)-8 levels in the HT-29 cells (P<0.01). In the in vivo studies with ApcMin/+ mice, after 10 or 20 mg/kg/day oral palmatine treatment, tumor numbers were significantly reduced in the small intestine and colon in a dose-dependent manner (P<0.01 compared with the model group). The results were supported by tumor distribution data, body weight changes and organ index. The effect on survival was also dose-dependent. Both the low- and high-dose palmatine treatments significantly increased the life span of the mice (P<0.01). The gut histology from the model group showed a prominent adenomatous change along with inflammatory lesions. With palmatine treatment, however, the dysplastic changes were greatly reduced in the small intestine and colon tissue. Reverse transcription-quantitative polymerase chain reaction analysis of interleukin (IL)-1α, IL1-β, IL-8, granulocyte-colony stimulating factor and granulocyte macrophage colony-stimulating factor in the gut tissue showed that these inflammatory cytokines were reduced significantly following treatment (all P<0.01); serum cytokine levels were also decreased. Data suggests that palmatine has a clinical value in colorectal cancer therapeutics, and this action is likely linked to the

  12. Variants in RBP4 and AR genes modulate age at onset in familial amyloid polyneuropathy (FAP ATTRV30M).

    PubMed

    Santos, Diana; Coelho, Teresa; Alves-Ferreira, Miguel; Sequeiros, Jorge; Mendonça, Denisa; Alonso, Isabel; Lemos, Carolina; Sousa, Alda

    2016-05-01

    Familial amyloid polyneuropathy (FAP) ATTRV30M is a neurodegenerative disorder due to point mutations in the transthyretin gene, with V30M being the commonest. FAP ATTRV30M shows a wide variation in age at onset (AO) between clusters, families and generations. Portuguese patients also show remarkable AO differences between genders. Genes found to be associated with FAP ATTRV30M pathways may act as AO modifiers. Our aim was to further explore the role of APCS and RBP4 genes and to study for the first time the involvement of sex-linked genetic modifiers - AR and HSD17B1 genes - in AO variation in Portuguese families. We collected DNA from a sample of 318 patients, currently under follow-up. A total of 18 tagging SNPs from APCS, RBP4, AR and HSD17B1 and 5 additional SNPs from APCS and RBP4 previously studied were genotyped. To account for nonindependency of AO between members of the same family, we used generalized estimating equations (GEEs). We found that APCS and RBP4 were associated with late AO. In addition, rs11187545 of the RBP4 was associated with an early AO. For the AR, in the male group three SNPs were associated with an early AO, whereas in the female group four were associated with both an early and later AO. These results strengthened the role of APCS and RBP4 genes and revealed for the first time the contribution of AR genes as an AO modifier in both males and females. These findings may have important implications in genetic counseling and for new therapeutic strategies. PMID:26286643

  13. Palmatine from Mahonia bealei attenuates gut tumorigenesis in ApcMin/+ mice via inhibition of inflammatory cytokines.

    PubMed

    Ma, Wei-Kun; Li, Hui; Dong, Cui-Lan; He, Xin; Guo, Chang-Run; Zhang, Chun-Feng; Yu, Chun-Hao; Wang, Chong-Zhi; Yuan, Chun-Su

    2016-07-01

    Mahonia bealei is a Chinese folk medicine used to treat various ailments, in particular gastrointestinal inflammation‑related illnesses, and palmatine is one of its active constituents. In this study, ApcMin/+ mice, a genetically engineered model, were used to investigate the effects of palmatine on the initiation and progression of gut inflammation and tumorigenesis enhanced by a high‑fat diet. The in vitro antiproliferation and anti‑inflammation effects of palmatine were evaluated on HT‑29 and SW‑480 human colorectal cancer cell lines. The concentration‑related antiproliferative effects of palmatine on both cell lines (P<0.01) were observed. Palmatine significantly inhibited lipopolysaccharide‑induced increase in cytokine interleukin (IL)‑8 levels in the HT‑29 cells (P<0.01). In the in vivo studies with ApcMin/+ mice, after 10 or 20 mg/kg/day oral palmatine treatment, tumor numbers were significantly reduced in the small intestine and colon in a dose‑dependent manner (P<0.01 compared with the model group). The results were supported by tumor distribution data, body weight changes and organ index. The effect on survival was also dose‑dependent. Both the low‑ and high‑dose palmatine treatments significantly increased the life span of the mice (P<0.01). The gut histology from the model group showed a prominent adenomatous change along with inflammatory lesions. With palmatine treatment, however, the dysplastic changes were greatly reduced in the small intestine and colon tissue. Reverse transcription‑quantitative polymerase chain reaction analysis of interleukin (IL)‑1α, IL1‑β, IL‑8, granulocyte‑colony stimulating factor and granulocyte macrophage colony‑stimulating factor in the gut tissue showed that these inflammatory cytokines were reduced significantly following treatment (all P<0.01); serum cytokine levels were also decreased. Data suggests that palmatine has a clinical value in colorectal cancer therapeutics, and

  14. Abdominal obesity, independent from caloric intake, accounts for the development of intestinal tumors in Apc(1638N/+) female mice.

    PubMed

    Huffman, Derek M; Augenlicht, Leonard H; Zhang, Xueying; Lofrese, John J; Atzmon, Gil; Chamberland, John P; Mantzoros, Christos S

    2013-03-01

    To determine whether visceral fat (VF), independent of other confounders, is causally linked to intestinal tumorigenesis, we surgically removed visceral fat in Apc(1638/N+) mice. At 15 weeks of age, male and female Apc(1638/N+) mice were randomized to one of three groups: ad libitum, visceral fat removal (VF-) and ad libitum fed, or caloric restriction, and were studied for effects on tumorigenesis and survival. As compared with ad libitum, VF- and caloric restriction reduced macroadenomas to a similar extent (P < 0.05), but only caloric restriction significantly improved survival (P < 0.05). Given that a significant group × gender interaction was observed, we next examined males and females separately. In females, macroadenomas were markedly attenuated by VF- (1.33 ± 0.23 mean ± SE; P < 0.05), but not by caloric restriction (2.35 ± 0.25; P = 0.71), as compared with ad libitum (2.50 ± 0.34). In males, however, caloric restriction (1.71 ± 0.26; P < 0.01), but not VF- (2.94 ± 0.42; P = 0.29), reduced macroadenomas, as compared with ad libitum males (3.47 ± 0.30). In females, both VF- (P = 0.05) and caloric restriction (P < 0.01) improved survival, but not in male mice (P = 0.15). The benefits observed with caloric restriction were consistent with favorable metabolic adaptations, but protection conferred in VF- females was despite lower adiponectin levels (P < 0.05), and failure to reduce body mass, total adiposity, glucose, insulin, leptin, and chemokine (C-X-C motif) ligand 1 (CXCL-1) levels. In conclusion, these data provide the first causal evidence linking visceral fat to intestinal cancer risk, and suggest that factors, other than known metabolic mediators, may impact tumor development. Furthermore, these data emphasize that strategies designed to deplete visceral fat stores in humans should be considered in the prevention of intestinal cancer. Cancer Prev Res; 6(3); 177-87. ©2012 AACR. PMID:23466815

  15. Methylation of multiple genes in hepatitis C virus associated hepatocellular carcinoma.

    PubMed

    Zekri, Abdel-Rahman N; Bahnasy, Abeer A; Shoeab, Fatma Elzahraa M; Mohamed, Waleed S; El-Dahshan, Dina H; Ali, Fahmey T; Sabry, Gilane M; Dasgupta, Nairajana; Daoud, Sayed S

    2014-01-01

    We studied promoter methylation (PM) of 11 genes in Peripheral Blood Lymphocytes (PBLs) and tissues of hepatitis C virus (HCV) associated hepatocellular carcinoma (HCC) and chronic hepatitis (CH) Egyptian patients. The present study included 31 HCC with their ANT, 38 CH and 13 normal hepatic tissue (NHT) samples. In all groups, PM of APC, FHIT, p15, p73, p14, p16, DAPK1, CDH1, RARβ, RASSF1A, O(6)MGMT was assessed by methylation-specific PCR (MSP). APC and O6-MGMT protein expression was assessed by immunohistochemistry (IHC) in the studied HCC and CH (20 samples each) as well as in a different HCC and CH set for confirmation of MSP results. PM was associated with progression from CH to HCC. Most genes showed high methylation frequency (MF) and the methylation index (MI) increased with disease progression. MF of p14, p73, RASSF1A, CDH1 and O(6)MGMT was significantly higher in HCC and their ANT. MF of APC was higher in CH. We reported high concordance between MF in HCC and their ANT, MF in PBL and CH tissues as well as between PM and protein expression of APC and O(6)MGMT. A panel of 4 genes (APC, p73, p14, O(6)MGMT) classifies the cases independently into HCC and CH with high accuracy (89.9%), sensitivity (83.9%) and specificity (94.7%). HCV infection may contribute to hepatocarcinogenesis through enhancing PM of multiple genes. PM of APC occurs early in the cascade while PM of p14, p73, RASSF1A, RARB, CDH1 and O(6)MGMT are late changes. A panel of APC, p73, p14, O6-MGMT could be used in monitoring CH patients for early detection of HCC. Also, we found that, the methylation status is not significantly affected by whether the tissue was from the liver or PBL, indicating the possibility of use PBL as indicator to genetic profile instead of liver tissue regardless the stage of disease. PMID:25685469

  16. Distinguishing Lung Adenocarcinoma from Lung Squamous Cell Carcinoma by Two Hypomethylated and Three Hypermethylated Genes: A Meta-Analysis.

    PubMed

    Huang, Tao; Li, Jinyun; Zhang, Cheng; Hong, Qingxiao; Jiang, Danjie; Ye, Meng; Duan, Shiwei

    2016-01-01

    Significant differences in the aberrant methylation of genes exist among various histological types of non-small cell lung cancer (NSCLC), which includes adenocarcinoma (AC) and squamous cell carcinoma (SCC). Different chemotherapeutic regimens should be administered to the two NSCLC subtypes due to their unique genetic and epigenetic profiles. The purpose of this meta-analysis was to generate a list of differentially methylated genes between AC and SCC. Our meta-analysis encompassed 151 studies on 108 genes among 12946 AC and 10243 SCC patients. Our results showed two hypomethylated genes (CDKN2A and MGMT) and three hypermethylated genes (CDH13, RUNX3 and APC) in ACs compared with SCCs. In addition, our results showed that the pooled specificity and sensitivity values of CDH13 and APC were higher than those of CDKN2A, MGMT and RUNX3. Our findings might provide an alternative method to distinguish between the two NSCLC subtypes. PMID:26862903

  17. Water quality assessment and apportionment of pollution sources using APCS-MLR and PMF receptor modeling techniques in three major rivers of South Florida.

    PubMed

    Haji Gholizadeh, Mohammad; Melesse, Assefa M; Reddi, Lakshmi

    2016-10-01

    In this study, principal component analysis (PCA), factor analysis (FA), and the absolute principal component score-multiple linear regression (APCS-MLR) receptor modeling technique were used to assess the water quality and identify and quantify the potential pollution sources affecting the water quality of three major rivers of South Florida. For this purpose, 15years (2000-2014) dataset of 12 water quality variables covering 16 monitoring stations, and approximately 35,000 observations was used. The PCA/FA method identified five and four potential pollution sources in wet and dry seasons, respectively, and the effective mechanisms, rules and causes were explained. The APCS-MLR apportioned their contributions to each water quality variable. Results showed that the point source pollution discharges from anthropogenic factors due to the discharge of agriculture waste and domestic and industrial wastewater were the major sources of river water contamination. Also, the studied variables were categorized into three groups of nutrients (total kjeldahl nitrogen, total phosphorus, total phosphate, and ammonia-N), water murkiness conducive parameters (total suspended solids, turbidity, and chlorophyll-a), and salt ions (magnesium, chloride, and sodium), and average contributions of different potential pollution sources to these categories were considered separately. The data matrix was also subjected to PMF receptor model using the EPA PMF-5.0 program and the two-way model described was performed for the PMF analyses. Comparison of the obtained results of PMF and APCS-MLR models showed that there were some significant differences in estimated contribution for each potential pollution source, especially in the wet season. Eventually, it was concluded that the APCS-MLR receptor modeling approach appears to be more physically plausible for the current study. It is believed that the results of apportionment could be very useful to the local authorities for the control and

  18. HCV RNA Activates APCs via TLR7/TLR8 While Virus Selectively Stimulates Macrophages Without Inducing Antiviral Responses.

    PubMed

    Zhang, Yuwei; El-Far, Mohamed; Dupuy, Franck P; Abdel-Hakeem, Mohamed S; He, Zhong; Procopio, Francesco Andrea; Shi, Yu; Haddad, Elias K; Ancuta, Petronela; Sekaly, Rafick-Pierre; Said, Elias A

    2016-01-01

    The innate and adaptive immune systems fail to control HCV infection in the majority of infected individuals. HCV is an ssRNA virus, which suggests a role for Toll-like receptors (TLRs) 7 and 8 in initiating the anti-viral response. Here we demonstrate that HCV genomic RNA harbours specific sequences that initiate an anti-HCV immune response through TLR7 and TLR8 in various antigen presenting cells. Conversely, HCV particles are detected by macrophages, but not by monocytes and DCs, through a TLR7/8 dependent mechanism; this leads to chloroquine sensitive production of pro-inflammatory cytokines including IL-1β, while the antiviral type I Interferon response is not triggered in these cells. Antibodies to DC-SIGN, a c-type lectin selectively expressed by macrophages but not pDCs or mDCs, block the production of cytokines. Novel anti-HCV vaccination strategies should target the induction of TLR7/8 stimulation in APCs in order to establish potent immune responses against HCV. PMID:27385120

  19. Reai-time imaging of Myeioid Ceiis Dynamics in ApcMin/+ intestinai Tumors by Spinning Disk Confocai Microscopy

    PubMed Central

    Werb, Zena

    2015-01-01

    Myeloid cells are the most abundant immune cells within tumors and have been shown to promote tumor progression. Modern intravital imaging techniques enable the observation of live cellular behavior inside the organ but can be challenging in some types of cancer due to organ and tumor accessibility such as intestine. Direct observation of intestinal tumors has not been previously reported. A surgical procedure described here allows direct observation of myeloid cell dynamics within the intestinal tumors in live mice by using transgenic fluorescent reporter mice and injectable tracers or antibodies. For this purpose, a four-color, multi-region, micro-lensed spinning disk confocai microscope that allows long-term continuous imaging with rapid image acquisition has been used. ApcMin/+ Spc mice that develop multiple adenomas in the small intestine are crossed with c-fms-EGFP mice to visualize myeloid cells and with ACTB-ECFP mice to visualize intestinal epithelial cells of the crypts. Procedures for labeling different tumor components, such as blood vessels and neutrophils, and the procedure for positioning the tumor for imaging through the serosal surface are also described. Time-lapse movies compiled from several hours of imaging allow the analysis of myeloid cell behavior in situ in the intestinal microenvironment. PMID:25350573

  20. HCV RNA Activates APCs via TLR7/TLR8 While Virus Selectively Stimulates Macrophages Without Inducing Antiviral Responses

    PubMed Central

    Zhang, Yuwei; El-Far, Mohamed; Dupuy, Franck P.; Abdel-Hakeem, Mohamed S.; He, Zhong; Procopio, Francesco Andrea; Shi, Yu; Haddad, Elias K.; Ancuta, Petronela; Sekaly, Rafick-Pierre; Said, Elias A.

    2016-01-01

    The innate and adaptive immune systems fail to control HCV infection in the majority of infected individuals. HCV is an ssRNA virus, which suggests a role for Toll-like receptors (TLRs) 7 and 8 in initiating the anti-viral response. Here we demonstrate that HCV genomic RNA harbours specific sequences that initiate an anti-HCV immune response through TLR7 and TLR8 in various antigen presenting cells. Conversely, HCV particles are detected by macrophages, but not by monocytes and DCs, through a TLR7/8 dependent mechanism; this leads to chloroquine sensitive production of pro-inflammatory cytokines including IL-1β, while the antiviral type I Interferon response is not triggered in these cells. Antibodies to DC-SIGN, a c-type lectin selectively expressed by macrophages but not pDCs or mDCs, block the production of cytokines. Novel anti-HCV vaccination strategies should target the induction of TLR7/8 stimulation in APCs in order to establish potent immune responses against HCV. PMID:27385120

  1. Maternal Antibiotic Treatment Protects Offspring from Diabetes Development in Nonobese Diabetic Mice by Generation of Tolerogenic APCs.

    PubMed

    Hu, Youjia; Peng, Jian; Tai, Ningwen; Hu, Changyun; Zhang, Xiaojun; Wong, F Susan; Wen, Li

    2015-11-01

    Type 1 diabetes (T1D) is a T cell-mediated autoimmune disease that involves the slow, progressive destruction of islet β cells and loss of insulin production, as a result of interaction with environmental factors, in genetically susceptible individuals. The gut microbiome is established very early in life. Commensal microbiota establish mutualism with the host and form an important part of the environment to which individuals are exposed in the gut, providing nutrients and shaping immune responses. In this study, we studied the impact of targeting most Gram-negative bacteria in the gut of NOD mice at different time points in their life, using a combination of three antibiotics--neomycin, polymyxin B, and streptomycin--on diabetes development. We found that the prenatal period is a critical time for shaping the immune tolerance in the progeny, influencing development of autoimmune diabetes. Prenatal neomycin, polymyxin B, and streptomycin treatment protected NOD mice from diabetes development through alterations in the gut microbiota, as well as induction of tolerogenic APCs, which led to reduced activation of diabetogenic CD8 T cells. Most importantly, we found that the protective effect was age dependent, and the most profound protection was found when the mice were treated before birth. This indicates the importance of the prenatal environment and early exposure to commensal bacteria in shaping the host immune system and health. PMID:26401004

  2. Irsogladine maleate, a gastric mucosal protectant, suppresses intestinal polyp development in Apc-mutant mice

    PubMed Central

    Onuma, Wakana; Tomono, Susumu; Miyamoto, Shinngo; Fujii, Gen; Hamoya, Takahiro; Fujimoto, Kyoko; Miyoshi, Noriyuki; Fukai, Fumio; Wakabayashi, Keiji; Mutoh, Michihiro

    2016-01-01

    This study aimed to identify gastric mucosal protectants that suppress intestinal tumorigenesis in a mouse model. We chose six gastric mucosal protectants (ecabet sodium hydrate, irsogladine maleate, rebamipide, sofalcone, teprenone and troxipide) and examined their effects on the activity of oxidative stress-related transcriptional factors, including AP-1, NF-jB, NRF2, p53 and STAT3, in Caco-2 cells using a luciferase reporter gene assay. Among the six protectants, irsogladine maleate clearly inhibited NF-jB and AP-1 transcriptional activity. Furthermore, the chemopreventive property of irsogladine maleate was examined in a Min mouse model of familial adenomatous polyposis. Treatment with irsogladine maleate at doses of 5 and 50 ppm significantly reduced the number of intestinal polyps to 69% and 66% of the untreated control value, respectively. In these polyps, mRNA levels of the downstream targets of NF-jB, such as IL-1β and IL-6, were decreased by irsogladine maleate treatment. Moreover, the levels of oxidative stress-related markers, reactive carbonyl species, in the livers of Min mice were clearly decreased following the administration of irsogladine maleate. This study demonstrated that irsogladine maleate suppresses intestinal polyp formation in Min mice partly through the NF-jB signaling pathway, thus reducing oxidative stress. PMID:26840084

  3. T Cell and APC Dynamics In Situ Control the Outcome of Vaccination

    PubMed Central

    Khanna, Kamal M.; Blair, David A.; Vella, Anthony T.; McSorley, Stephen J.; Datta, Sandip K.; Lefrançois, Leo

    2010-01-01

    The factors controlling the progression of an immune response to generation of protective memory are poorly understood. We compared the in situ and ex vivo characteristics of CD8 T cells responding to different forms of the same immunogen. Immunization with live Listeria monocytogenes, irradiated L. monocytogenes (IRL), or heat-killed L. monocytogenes (HKL) induced rapid activation of CD8 T cells. However, only IRL and live L. monocytogenes inoculation induced sustained proliferation and supported memory development. Gene and protein expression analysis revealed that the three forms of immunization led to three distinct transcriptional and translational programs. Prior to cell division, CD8 T cell–dendritic cell clusters formed in the spleen after live L. monocytogenes and IRL but not after HKL immunization. Furthermore, HKL immunization induced rapid remodeling of splenic architecture, including loss of marginal zone macrophages, which resulted in impaired bacterial clearance. These results identify initial characteristics of a protective T cell response that have implications for the development of more effective vaccination strategies. PMID:20530268

  4. Structural and functional characterization of the Wnt inhibitor APC membrane recruitment 1 (Amer1).

    PubMed

    Tanneberger, Kristina; Pfister, Astrid S; Kriz, Vitezslav; Bryja, Vitezslav; Schambony, Alexandra; Behrens, Jürgen

    2011-06-01

    Amer1/WTX binds to the tumor suppressor adenomatous polyposis coli and acts as an inhibitor of Wnt signaling by inducing β-catenin degradation. We show here that Amer1 directly interacts with the armadillo repeats of β-catenin via a domain consisting of repeated arginine-glutamic acid-alanine (REA) motifs, and that Amer1 assembles the β-catenin destruction complex at the plasma membrane by recruiting β-catenin, adenomatous polyposis coli, and Axin/Conductin. Deletion or specific mutations of the membrane binding domain of Amer1 abolish its membrane localization and abrogate negative control of Wnt signaling, which can be restored by artificial targeting of Amer1 to the plasma membrane. In line, a natural splice variant of Amer1 lacking the plasma membrane localization domain is deficient for Wnt inhibition. Knockdown of Amer1 leads to the activation of Wnt target genes, preferentially in dense compared with sparse cell cultures, suggesting that Amer1 function is regulated by cell contacts. Amer1 stabilizes Axin and counteracts Wnt-induced degradation of Axin, which requires membrane localization of Amer1. The data suggest that Amer1 exerts its negative regulatory role in Wnt signaling by acting as a scaffold protein for the β-catenin destruction complex and promoting stabilization of Axin at the plasma membrane. PMID:21498506

  5. The E3 ligase APC/C-Cdh1 is required for associative fear memory and long-term potentiation in the amygdala of adult mice.

    PubMed

    Pick, Joseph E; Malumbres, Marcos; Klann, Eric

    2013-01-01

    The anaphase promoting complex/cyclosome (APC/C) is an E3 ligase regulated by Cdh1. Beyond its role in controlling cell cycle progression, APC/C-Cdh1 has been detected in neurons and plays a role in long-lasting synaptic plasticity and long-term memory. Herein, we further examined the role of Cdh1 in synaptic plasticity and memory by generating knockout mice where Cdh1 was conditionally eliminated from the forebrain post-developmentally. Although spatial learning and memory in the Morris water maze (MWM) was normal, the Cdh1 conditional knockout (cKO) mice displayed enhanced reversal learning in the MWM and in a water-based Y maze. In addition, we found that the Cdh1 cKO mice had impaired associative fear memory and exhibited impaired long-term potentiation (LTP) in amygdala slices. Finally, we observed increased expression of Shank1 and NR2A expression in amygdalar slices from the Cdh1 cKO mice following the induction of LTP, suggesting a possible molecular mechanism underlying the behavioral and synaptic plasticity impairments displayed in these mice. Our findings are consistent with a role for the APC/C-Cdh1 in fear memory and synaptic plasticity in the amygdala. PMID:23242419

  6. Pilot-scale road subbase made with granular material formulated with MSWI bottom ash and stabilized APC fly ash: environmental impact assessment.

    PubMed

    del Valle-Zermeño, R; Formosa, J; Prieto, M; Nadal, R; Niubó, M; Chimenos, J M

    2014-02-15

    A granular material (GM) to be used as road sub-base was formulated using 80% of weathered bottom ash (WBA) and 20% of mortar. The mortar was prepared separately and consisted in 50% APC and 50% of Portland cement. A pilot-scale study was carried on by constructing three roads in order to environmentally evaluate the performance of GM in a real scenario. By comparing the field results with those of the column experiments, the overestimations observed at laboratory scale can be explained by the potential mechanisms in which water enters into the road body and the pH of the media. An exception was observed in the case of Cu, whose concentration release at the test road was higher. The long-time of exposure at atmospheric conditions might have favoured oxidation of organic matter and therefore the leaching of this element. The results obtained showed that immobilization of all heavy metals and metalloids from APC is achieved by the pozzolanic effect of the cement mortar. This is, to the knowledge of the authors, the only pilot scale study that is considering reutilization of APC as a safe way to disposal. PMID:24394668

  7. Wnt/Wingless Pathway Activation Is Promoted by a Critical Threshold of Axin Maintained by the Tumor Suppressor APC and the ADP-Ribose Polymerase Tankyrase.

    PubMed

    Wang, Zhenghan; Tacchelly-Benites, Ofelia; Yang, Eungi; Thorne, Curtis A; Nojima, Hisashi; Lee, Ethan; Ahmed, Yashi

    2016-05-01

    Wnt/β-catenin signal transduction directs metazoan development and is deregulated in numerous human congenital disorders and cancers. In the absence of Wnt stimulation, a multiprotein "destruction complex," assembled by the scaffold protein Axin, targets the key transcriptional activator β-catenin for proteolysis. Axin is maintained at very low levels that limit destruction complex activity, a property that is currently being exploited in the development of novel therapeutics for Wnt-driven cancers. Here, we use an in vivo approach in Drosophila to determine how tightly basal Axin levels must be controlled for Wnt/Wingless pathway activation, and how Axin stability is regulated. We find that for nearly all Wingless-driven developmental processes, a three- to fourfold increase in Axin is insufficient to inhibit signaling, setting a lower-limit for the threshold level of Axin in the majority of in vivo contexts. Further, we find that both the tumor suppressor adenomatous polyposis coli (APC) and the ADP-ribose polymerase Tankyrase (Tnks) have evolutionarily conserved roles in maintaining basal Axin levels below this in vivo threshold, and we define separable domains in Axin that are important for APC- or Tnks-dependent destabilization. Together, these findings reveal that both APC and Tnks maintain basal Axin levels below a critical in vivo threshold to promote robust pathway activation following Wnt stimulation. PMID:26975665

  8. Anillin is a substrate of anaphase-promoting complex/cyclosome (APC/C) that controls spatial contractility of myosin during late cytokinesis.

    PubMed

    Zhao, Wei-Meng; Fang, Guowei

    2005-09-30

    Anillin, an actin-binding protein localized at the cleavage furrow, is required for cytokinesis. Through an in vitro expression screen, we identified anillin as a substrate of the anaphase-promoting complex/cyclosome (APC/C), a ubiquitin ligase that controls mitotic progression. We found that the levels of anillin fluctuate in the cell cycle, peaking in mitosis and dropping drastically during mitotic exit. Ubiquitination of anillin required a destruction-box and was mediated by Cdh1, an activator of APC/C. Overexpression of Cdh1 reduced the levels of anillin, whereas inactivation of APC/C(Cdh1) increased the half-life of anillin. Functionally, anillin was required for the completion of cytokinesis. In anillin knockdown cells, the cleavage furrow ingressed but failed to complete the ingression. At late cytokinesis, the cytosol and DNA in knockdown cells underwent rapid myosin-based oscillatory movement across the furrow. During this movement, RhoA and active myosin were absent from the cleavage furrow, and myosin was redistributed to cortical patches, which powers the random oscillatory movement. We concluded that anillin functions to maintain the localization of active myosin, thereby ensuring the spatial control of concerted contraction during cytokinesis. PMID:16040610

  9. Systems Biology Modeling of Five Pathways for Regulation and Potent Inhibition of the Anaphase-Promoting Complex (APC/C): Pivotal Roles for MCC and BubR1

    PubMed Central

    2015-01-01

    Abstract Correct DNA segregation is a fundamental process that ensures the precise and reliable inheritance of genomic information for the propagation of cell life. Eukaryotic cells have evolved a conserved surveillance control mechanism for DNA segregation named the Spindle Assembly Checkpoint (SAC).The SAC ensures that the sister chromatids of the duplicated genome are not separated and distributed to the spindle poles before all chromosomes have been properly linked to the microtubules of the mitotic spindle. Biochemically, the SAC delays cell cycle progression by preventing activation of the anaphase-promoting complex (APC/C) or cyclosome whose activation by Cdc20 is required for sister-chromatid separation; this marks the transition into anaphase. In response to activation of the checkpoint, various species control the activity of both APC/C and Cdc20. However, the underlying regulatory pathways remain largely elusive. In this study, five possible model variants of APC/C regulation were constructed, namely BubR1, Mad2, MCC, MCF2, and an all-pathways model variant. These models were validated with experimental data from the literature. A wide range of parameter values has been tested to find the critical values of the APC/C binding rate. The results show that all variants are able to capture the wild-type behavior of the APC/C. However, only one model variant, which included both MCC as well as BubR1 as potent inhibitors of the APC/C, was able to reproduce both wild-type and mutant type behavior of APC/C regulation. In conclusion, the presented work informs the regulation of fundamental processes such as SAC and APC/C in cell biology and has successfully distinguished between five competing dynamical models using a systems biology approach. The results attest that systems-level approaches are vital for molecular and cell biology. PMID:25871779

  10. Methylation of the RARB Gene Increases Prostate Cancer Risk in Black Americans

    PubMed Central

    Tang, Deliang; Kryvenko, Oleksandr N.; Mitrache, Nicoleta; Do, Kieu C.; Jankowski, Michelle; Chitale, Dhananjay A.; Trudeau, Sheri; Rundle, Andrew; Belinsky, Steven A.; Rybicki, Benjamin A.

    2013-01-01

    Purpose Gene promoter hypermethylation may be useful as a biomarker for cancer risk in histopathologically benign prostate specimens. Materials and Methods We performed a nested case-control study of gene promoter methylation status for 5 genes (APC, RARB, CCND2, RASSF1 and MGMT) measured in benign biopsy specimens from 511 prostate cancer case-control pairs. We estimated the overall and race stratified risk of subsequent prostate cancer associated with methylation status. Results On race stratified analysis RARB methylation was associated with a higher cancer risk in black American men (OR 2.18, 95% CI 1.39–3.44). APC methylation was associated with an increased risk of high grade tumors (OR 2.43, 95% CI 1.20–4.90), which was higher in black than in white men (OR 3.21 vs 2.04). In cases RARB and APC gene methylation in benign prostate samples persisted in matched malignant specimens. In black cases the combined risk associated with RARB and APC methylation (OR 3.04, 95% CI 1.44–6.42) was greater than the individual risk of each gene and significantly different from that in white cases (OR 1.14, 95% CI 0.56–2.30). Conclusions RARB gene methylation in histopathologically benign prostate samples was associated with a statistically significant increased risk of subsequent prostate cancer in black men. Methylation data on additional genes may improve risk stratification and clinical decision making algorithms for cancer screening and diagnosis. PMID:23376149

  11. Cdk5-mediated inhibition of APC/C-Cdh1 switches on the cyclin D1-Cdk4-pRb pathway causing aberrant S-phase entry of postmitotic neurons

    PubMed Central

    Veas-Pérez de Tudela, Miguel; Maestre, Carolina; Delgado-Esteban, María; Bolaños, Juan P.; Almeida, Angeles

    2015-01-01

    The anaphase-promoting complex/cyclosome (APC/C) is an E3 ubiquitin ligase that regulates cell cycle progression in proliferating cells. To enter the S-phase, APC/C must be inactivated by phosphorylation of its cofactor, Cdh1. In post-mitotic cells such as neurons APC/C-Cdh1 complex is highly active and responsible for the continuous degradation of mitotic cyclins. However, the specific molecular pathway that determines neuronal cell cycle blockade in post-mitotic neurons is unknown. Here, we show that activation of glutamatergic receptors in rat cortical primary neurons endogenously triggers cyclin-dependent kinase-5 (Cdk5)-mediated phosphorylation of Cdh1 leading to its cytoplasmic accumulation and disassembly from the APC3 core protein, causing APC/C inactivation. Conversely, pharmacological or genetic inhibition of Cdk5 promotes Cdh1 ubiquitination and proteasomal degradation. Furthermore, we show that Cdk5-mediated phosphorylation and inactivation of Cdh1 leads to p27 depletion, which switches on the cyclin D1-cyclin-dependent kinase-4 (Cdk4)-retinoblastoma protein (pRb) pathway to allow the S-phase entry of neurons. However, neurons do not proceed through the cell cycle and die by apoptosis. These results indicate that APC/C-Cdh1 actively suppresses an aberrant cell cycle entry and death of neurons, highlighting its critical function in neuroprotection. PMID:26658992

  12. Influence of Cationic Lipid Composition on Gene Silencing Properties of Lipid Nanoparticle Formulations of siRNA in Antigen-Presenting Cells

    PubMed Central

    Basha, Genc; Novobrantseva, Tatiana I; Rosin, Nicole; Tam, Yuen Yi C; Hafez, Ismail M; Wong, Matthew K; Sugo, Tsukasa; Ruda, Vera M; Qin, June; Klebanov, Boris; Ciufolini, Marco; Akinc, Akin; Tam, Ying K; Hope, Michael J; Cullis, Pieter R

    2011-01-01

    Lipid nanoparticles (LNPs) are currently the most effective in vivo delivery systems for silencing target genes in hepatocytes employing small interfering RNA. Antigen-presenting cells (APCs) are also potential targets for LNP siRNA. We examined the uptake, intracellular trafficking, and gene silencing potency in primary bone marrow macrophages (bmMΦ) and dendritic cells of siRNA formulated in LNPs containing four different ionizable cationic lipids namely DLinDAP, DLinDMA, DLinK-DMA, and DLinKC2-DMA. LNPs containing DLinKC2-DMA were the most potent formulations as determined by their ability to inhibit the production of GAPDH target protein. Also, LNPs containing DLinKC2-DMA were the most potent intracellular delivery agents as indicated by confocal studies of endosomal versus cytoplamic siRNA location using fluorescently labeled siRNA. DLinK-DMA and DLinKC2-DMA formulations exhibited improved gene silencing potencies relative to DLinDMA but were less toxic. In vivo results showed that LNP siRNA systems containing DLinKC2-DMA are effective agents for silencing GAPDH in APCs in the spleen and peritoneal cavity following systemic administration. Gene silencing in APCs was RNAi mediated and the use of larger LNPs resulted in substantially reduced hepatocyte silencing, while similar efficacy was maintained in APCs. These results are discussed with regard to the potential of LNP siRNA formulations to treat immunologically mediated diseases. PMID:21971424

  13. A rapid and scalable system for studying gene function in mice using conditional RNA interference

    PubMed Central

    Premsrirut, Prem K.; Dow, Lukas E.; Kim, Sang Yong; Camiolo, Matthew; Malone, Colin D.; Miething, Cornelius; Scuoppo, Claudio; Zuber, Johannes; Dickins, Ross A.; Kogan, Scott C.; Shroyer, Kenneth R.; Sordella, Raffaella; Hannon, Gregory J.; Lowe, Scott W.

    2011-01-01

    Summary RNA interference is a powerful tool for studying gene function, however, the reproducible generation of RNAi transgenic mice remains a significant limitation. By combining optimized fluorescence-coupled miR30-based shRNAs with high efficiency ES cell targeting, we developed a fast, scalable pipeline for the production of shRNA transgenic mice. Using this system, we generated eight tet-regulated shRNA transgenic lines targeting Firefly and Renilla luciferases, Oct4 and tumor suppressors p53, p16INK4a, p19ARF and APC and demonstrate potent gene silencing and GFP-tracked knockdown in a broad range of tissues in vivo. Further, using an shRNA targeting APC, we illustrate how this approach can identify predicted phenotypes and also unknown functions for a well-studied gene. In addition, through regulated gene silencing we validate APC/Wnt and p19ARF as potential therapeutic targets in T cell acute lymphoblastic leukemia/lymphoma and lung adenocarcinoma, respectively. This system provides a cost-effective and scalable platform for the production of RNAi transgenic mice targeting any mammalian gene. PMID:21458673

  14. Diet- and Genetically-Induced Obesity Differentially Affect the Fecal Microbiome and Metabolome in Apc1638N Mice

    PubMed Central

    Parnell, Laurence D.; Iyer, Lakshmanan K.; Liu, Zhenhua; Kane, Anne V.; Chen, C-Y. Oliver; Tai, Albert K.; Bowman, Thomas A.; Obin, Martin S.; Mason, Joel B.; Greenberg, Andrew S.; Choi, Sang-Woon; Selhub, Jacob; Paul, Ligi; Crott, Jimmy W.

    2015-01-01

    Obesity is a risk factor for colorectal cancer (CRC), and alterations in the colonic microbiome and metabolome may be mechanistically involved in this relationship. The relative contribution of diet and obesity per se are unclear. We compared the effect of diet- and genetically-induced obesity on the intestinal microbiome and metabolome in a mouse model of CRC. Apc1638N mice were made obese by either high fat (HF) feeding or the presence of the Leprdb/db (DbDb) mutation. Intestinal tumors were quantified and stool microbiome and metabolome were profiled. Genetic obesity, and to a lesser extent HF feeding, promoted intestinal tumorigenesis. Each induced distinct microbial patterns: taxa enriched in HF were mostly Firmicutes (6 of 8) while those enriched in DbDb were split between Firmicutes (7 of 12) and Proteobacteria (5 of 12). Parabecteroides distasonis was lower in tumor-bearing mice and its abundance was inversely associated with colonic Il1b production (p<0.05). HF and genetic obesity altered the abundance of 49 and 40 fecal metabolites respectively, with 5 in common. Of these 5, adenosine was also lower in obese and in tumor-bearing mice (p<0.05) and its concentration was inversely associated with colonic Il1b and Tnf production (p<0.05). HF and genetic obesity differentially alter the intestinal microbiome and metabolome. A depletion of adenosine and P.distasonis in tumor-bearing mice could play a mechanistic role in tumor formation. Adenosine and P. distasonis have previously been shown to be anti-inflammatory in the colon and we postulate their reduction could promote tumorigenesis by de-repressing inflammation. PMID:26284788

  15. Gene promoter methylation in colorectal cancer and healthy adjacent mucosa specimens

    PubMed Central

    Coppedè, Fabio; Migheli, Francesca; Lopomo, Angela; Failli, Alessandra; Legitimo, Annalisa; Consolini, Rita; Fontanini, Gabriella; Sensi, Elisa; Servadio, Adele; Seccia, Massimo; Zocco, Giuseppe; Chiarugi, Massimo; Spisni, Roberto; Migliore, Lucia

    2014-01-01

    We evaluated the promoter methylation levels of the APC, MGMT, hMLH1, RASSF1A and CDKN2A genes in 107 colorectal cancer (CRC) samples and 80 healthy adjacent tissues. We searched for correlation with both physical and pathological features, polymorphisms of folate metabolism pathway genes (MTHFR, MTRR, MTR, RFC1, TYMS, and DNMT3B), and data on circulating folate, vitamin B12 and homocysteine, which were available in a subgroup of the CRC patients. An increased number of methylated samples were found in CRC respect to adjacent healthy tissues, with the exception of APC, which was also frequently methylated in healthy colonic mucosa. Statistically significant associations were found between RASSF1A promoter methylation and tumor stage, and between hMLH1 promoter methylation and tumor location. Increasing age positively correlated with both hMLH1 and MGMT methylation levels in CRC tissues, and with APC methylation levels in the adjacent healthy mucosa. Concerning gender, females showed higher hMLH1 promoter methylation levels with respect to males. In CRC samples, the MTR 2756AG genotype correlated with higher methylation levels of RASSF1A, and the TYMS 1494 6bp ins/del polymorphism correlated with the methylation levels of both APC and hMLH1. In adjacent healthy tissues, MTR 2756AG and TYMS 1494 6bp del/del genotypes correlated with APC and MGMT promoter methylation, respectively. Low folate levels were associated with hMLH1 hypermethylation. Present results support the hypothesis that DNA methylation in CRC depends from both physiological and environmental factors, with one-carbon metabolism largely involved in this process. PMID:24500500

  16. Modified irinotecan and infusional 5-fluorouracil (mFOLFIRI) in patients with refractory advanced pancreas cancer (APC): a single-institution experience

    PubMed Central

    Bupathi, M.; Ahn, D. H.; Wu, C.; Ciombor, K. K.; Stephens, J. A.; Reardon, J.; Goldstein, D. A.; Bekaii-Saab, T.

    2016-01-01

    Pancreatic adenocarcinoma is the fourth leading cause of cancer death. Recently, MM-398 (nanoliposomal irinotecan) was shown to be associated with significant improvement in outcome measures with acceptable toxicities when combined with 5-fluorouracil (5-FU)/leucovorin (LV) compared to 5-FU/LV alone in patients failing one line of gemcitabine-based therapy. There is a paucity of data evaluating the role of irinotecan in combination with 5FU in advanced pancreas cancer (APC). We performed a retrospective analysis of all patients who received mFOLFIRI (minus bolus 5FU and LV). All patients with metastatic disease who had failed at least one line of gemcitabine-based therapy prior to receiving mFOLFIRI were included in this study. Descriptive statistics were used to assess the continuous variables and adverse events (AEs), and Kaplan–Meier methods were used to calculate the median progression-free survival (PFS) and overall survival (OS). Forty patients were included in this analysis. Patients received 1–5 lines of prior therapy (25 % with more than 3 lines of prior therapy). The mean age at diagnosis was 60, and 98 % had ECOG of 1. The mean CA 19-9 at the start of therapy was 33,169 U/ml. The median PFS was 2.59 months [95 % confidence interval (CI) (1.90, 3.54)], and OS was 4.75 months [95 % CI (3.14, 8.98)]. The most common AEs included fatigue (98 %), neuropathy (83 %), anorexia (68 %), nausea (60 %) and constipation (55 %). Grade 3 toxicities included fatigue (13 %) and rash (3 %). There were no observed grade 4 toxicities. In this single-institution retrospective analysis, mFOLFIRI was found to be both tolerable and relatively effective in a heavily pretreated patient population with APC. Future prospective studies should consider evaluating the role of mFOLFIRI in refractory APC. PMID:26995224

  17. Spermine synthase overexpression in vivo does not increase susceptibility to DMBA/TPA skin carcinogenesis or Min-Apc intestinal tumorigenesis.

    PubMed

    Welsh, Patricia A; Sass-Kuhn, Suzanne; Prakashagowda, Chethana; McCloskey, Diane; Feith, David

    2012-04-01

    Numerous studies have demonstrated a link between elevated polyamine biosynthesis and neoplastic growth, but the specific contribution of spermine synthase to epithelial tumor development has never been explored in vivo. Mice with widespread overexpression of spermine synthase (CAG-SpmS) exhibit decreased spermidine levels, increased spermine and a significant rise in tissue spermine:spermidine ratio. We characterized the response of CAG-SpmS mice to two-stage skin chemical carcinogenesis as well as spontaneous intestinal carcinogenesis induced by loss of the Apc tumor suppressor in Apc (Min) (/+) (Min) mice. CAG-SpmS mice maintained the canonical increases in ornithine decarboxylase (ODC) activity, polyamine content and epidermal thickness in response to tumor promoter treatment of the skin. The induction of S-adenosylmethionine decarboxylase (AdoMetDC) activity and its product decarboxylated AdoMet were impaired in CAG-SpmS mice, and the spermine:spermidine ratio was increased 3-fold in both untreated and 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated skin. The susceptibility to 7,12-dimethylbenz[a]anthracene (DMBA)/TPA skin carcinogenesis was not altered in CAG-SpmS mice, and SpmS overexpression did not modify the previously described tumor resistance of mice with targeted antizyme expression or the enhanced tumor response in mice with targeted spermidine/spermine-N ( 1) -acetyltransferase expression. CAG-SpmS/Min mice also exhibited elevated spermine:spermidine ratios in the small intestine and colon, yet their tumor multiplicity and size was similar to Min mice. Therefore, studies in two of the most widely used tumorigenesis models demonstrate that increased spermine synthase activity and the resulting elevation of the spermine:spermidine ratio does not alter susceptibility to tumor development initiated by c-Ha-Ras mutation or Apc loss. PMID:22258329

  18. Integration and automation of DoseMapper in a logic fab APC system: application for 45/40/28nm node

    NASA Astrophysics Data System (ADS)

    Le Gratiet, Bertrand; Salagnon, Christophe; de Caunes, Jean; Mikolajczak, Marc; Morin, Vincent; Chojnowski, Nicolas; Sundermann, Frank; Massin, Jean; Pelletier, Alice; Metz, Joel; Blancquaert, Yoann; Bouyssou, Regis; Pelissier, Arthur; Belmont, Olivier; Strapazzon, Anne; Phillips, Anna; Devoivre, Thierry; Bernard, Emilie; Batail, Estelle; Thevenon, Lionel; Bry, Benedicte; Bernard-Granger, Fabrice; Oumina, Ahmed; Baron, Marie-Pierre; Gueze, Didier

    2012-03-01

    The main difficulty related to DoseMapper correction is to generate an appropriate CD datacollection to feed DoseMapper and to generate DoseRecipe in a user friendly way, especially with a complex process mix. We could heavily measure the silicon and create, in feedback mode, the corresponding DoseRecipe. However, such approach in a logic fab becomes a heavy duty due to the number of different masks / product / processes. We have observed that process CD variability is significantly depending on systematic intrawafer and intrafield CD footprints that can be measured and applied has generic pre-correction for any new product/mask process in-line. The applied CD correction is based on a CD (intrafield: Mask + Straylight & intrawafer: Etch Bias) variability "model" handled by the FAB APC (Advanced Process Control). - Individual CD profile correction component are generated "off-line" (1) for Intrafield Mask via automatic CD extraction from a Reticle CD database (2) for Intrafield Straylight via a CD "model" (3) for Intrawafer Etch Bias via engineering input based on process monitoring. - These CD files are handled via the FAB APC/automation system which is remotely taking control of DoseMapper server via WEB services, so that CD profiles are generated "off-line" (before the lot is being processed) and stored in a profile database while DoseRecipes are created "real-time" on demand via the automation when the lot comes to the scanner to be processed. DoseRecipe and CD correction profiles management is done via the APC system. The automated DoseRecipe creation is now running since the beginning of 2011 contributing to bring both intrafield and intrawafer GATE CDu below 1nm 3sigma, for 45/40 & 28nm nodes.

  19. Source apportionment of ambient non-methane hydrocarbons in Hong Kong: application of a principal component analysis/absolute principal component scores (PCA/APCS) receptor model.

    PubMed

    Guo, H; Wang, T; Louie, P K K

    2004-06-01

    Receptor-oriented source apportionment models are often used to identify sources of ambient air pollutants and to estimate source contributions to air pollutant concentrations. In this study, a PCA/APCS model was applied to the data on non-methane hydrocarbons (NMHCs) measured from January to December 2001 at two sampling sites: Tsuen Wan (TW) and Central & Western (CW) Toxic Air Pollutants Monitoring Stations in Hong Kong. This multivariate method enables the identification of major air pollution sources along with the quantitative apportionment of each source to pollutant species. The PCA analysis identified four major pollution sources at TW site and five major sources at CW site. The extracted pollution sources included vehicular internal engine combustion with unburned fuel emissions, use of solvent particularly paints, liquefied petroleum gas (LPG) or natural gas leakage, and industrial, commercial and domestic sources such as solvents, decoration, fuel combustion, chemical factories and power plants. The results of APCS receptor model indicated that 39% and 48% of the total NMHCs mass concentrations measured at CW and TW were originated from vehicle emissions, respectively. 32% and 36.4% of the total NMHCs were emitted from the use of solvent and 11% and 19.4% were apportioned to the LPG or natural gas leakage, respectively. 5.2% and 9% of the total NMHCs mass concentrations were attributed to other industrial, commercial and domestic sources, respectively. It was also found that vehicle emissions and LPG or natural gas leakage were the main sources of C(3)-C(5) alkanes and C(3)-C(5) alkenes while aromatics were predominantly released from paints. Comparison of source contributions to ambient NMHCs at the two sites indicated that the contribution of LPG or natural gas at CW site was almost twice that at TW site. High correlation coefficients (R(2) > 0.8) between the measured and predicted values suggested that the PCA/APCS model was applicable for estimation

  20. Cell cycle-dependent nuclear export of Cdh1p may contribute to the inactivation of APC/C(Cdh1).

    PubMed

    Jaquenoud, Malika; van Drogen, Frank; Peter, Matthias

    2002-12-01

    Cdh1p is a substrate-specific subunit of the anaphase-promoting complex (APC/C), which functions as an E3 ubiquitin ligase to degrade the mitotic cyclin Clb2p and other substrates during the G(1) phase of the cell cycle. Cdh1p is phosphorylated and thereby inactivated at the G(1)/S transition predominantly by Cdc28p-Clb5p. Here we show that Cdh1p is nuclear during the G(1) phase of the cell cycle, but redistributes to the cytoplasm between S phase and the end of mitosis. Nuclear export of Cdh1p is regulated by phosphorylation and requires active Cdc28p kinase. Cdh1p binds to the importin Pse1p and the exportin Msn5p, which is necessary and sufficient to promote efficient export of Cdh1p in vivo. Although msn5delta cells are viable, they are sensitive to Cdh1p overexpression. Likewise, a mutant form of Cdh1p, which is constitutively nuclear, prevents accumulation of Clb2p and leads to cell cycle arrest when overexpressed in wild-type cells. Taken together, these results suggest that phosphorylation-dependent nuclear export of Cdh1p by Msn5p contributes to efficient inactivation of APC/C(Cdh1). PMID:12456658

  1. A novel tankyrase inhibitor decreases canonical Wnt signaling in colon carcinoma cells and reduces tumor growth in conditional APC mutant mice.

    PubMed

    Waaler, Jo; Machon, Ondrej; Tumova, Lucie; Dinh, Huyen; Korinek, Vladimir; Wilson, Steven Ray; Paulsen, Jan Erik; Pedersen, Nina Marie; Eide, Tor J; Machonova, Olga; Gradl, Dietmar; Voronkov, Andrey; von Kries, Jens Peter; Krauss, Stefan

    2012-06-01

    Increased nuclear accumulation of β-catenin, a mediator of canonical Wnt signaling, is found in numerous tumors and is frequently associated with tumor progression and metastasis. Inhibition of Wnt/β-catenin signaling therefore is an attractive strategy for anticancer drugs. In this study, we have identified a novel small molecule inhibitor of the β-catenin signaling pathway, JW55, that functions via inhibition of the PARP domain of tankyrase 1 and tankyrase 2 (TNKS1/2), regulators of the β-catenin destruction complex. Inhibition of TNKS1/2 poly(ADP-ribosyl)ation activity by JW55 led to stabilization of AXIN2, a member of the β-catenin destruction complex, followed by increased degradation of β-catenin. In a dose-dependent manner, JW55 inhibited canonical Wnt signaling in colon carcinoma cells that contained mutations in either the APC (adenomatous polyposis coli) locus or in an allele of β-catenin. In addition, JW55 reduced XWnt8-induced axis duplication in Xenopus embryos and tamoxifen-induced polyposis formation in conditional APC mutant mice. Together, our findings provide a novel chemotype for targeting canonical Wnt/β-catenin signaling through inhibiting the PARP domain of TNKS1/2. PMID:22440753

  2. Modulation of the ligand-independent traffic of Notch by Axin and Apc contributes to the activation of Armadillo in Drosophila.

    PubMed

    Muñoz-Descalzo, Silvia; Tkocz, Katarzyna; Balayo, Tina; Arias, Alfonso Martinez

    2011-04-01

    There is increasing evidence for close functional interactions between Wnt and Notch signalling. In many instances, these are mediated by convergence of the signalling events on common transcriptional targets, but there are other instances that cannot be accounted for in this manner. Studies in Drosophila have revealed that an activated form of Armadillo, the effector of Wnt signalling, interacts with, and is modulated by, the Notch receptor. Specifically, the ligand-independent traffic of Notch serves to set up a threshold for the amount of this form of Armadillo and therefore for Wnt signalling. In the current model of Wnt signalling, a complex assembled around Axin and Apc allows GSK3 (Shaggy) to phosphorylate Armadillo and target it for degradation. However, genetic experiments suggest that the loss of function of any of these three elements does not have the same effect as elevating the activity of β-catenin. Here, we show that Axin and Apc, but not GSK3, modulate the ligand-independent traffic of Notch. This finding helps to explain unexpected differences in the phenotypes obtained by different ways of activating Armadillo function and provides further support for the notion that Wnt and Notch signalling form a single functional module. PMID:21389052

  3. Myostatin gene inactivation prevents skeletal muscle wasting in cancer.

    PubMed

    Gallot, Yann S; Durieux, Anne-Cécile; Castells, Josiane; Desgeorges, Marine M; Vernus, Barbara; Plantureux, Léa; Rémond, Didier; Jahnke, Vanessa E; Lefai, Etienne; Dardevet, Dominique; Nemoz, Georges; Schaeffer, Laurent; Bonnieu, Anne; Freyssenet, Damien G

    2014-12-15

    Cachexia is a muscle-wasting syndrome that contributes significantly to morbidity and mortality of many patients with advanced cancers. However, little is understood about how the severe loss of skeletal muscle characterizing this condition occurs. In the current study, we tested the hypothesis that the muscle protein myostatin is involved in mediating the pathogenesis of cachexia-induced muscle wasting in tumor-bearing mice. Myostatin gene inactivation prevented the severe loss of skeletal muscle mass induced in mice engrafted with Lewis lung carcinoma (LLC) cells or in Apc(Min) (/+) mice, an established model of colorectal cancer and cachexia. Mechanistically, myostatin loss attenuated the activation of muscle fiber proteolytic pathways by inhibiting the expression of atrophy-related genes, MuRF1 and MAFbx/Atrogin-1, along with autophagy-related genes. Notably, myostatin loss also impeded the growth of LLC tumors, the number and the size of intestinal polyps in Apc(Min) (/+) mice, thus strongly increasing survival in both models. Gene expression analysis in the LLC model showed this phenotype to be associated with reduced expression of genes involved in tumor metabolism, activin signaling, and apoptosis. Taken together, our results reveal an essential role for myostatin in the pathogenesis of cancer cachexia and link this condition to tumor growth, with implications for furthering understanding of cancer as a systemic disease. PMID:25336187

  4. MGMT-B gene promoter hypermethylation in patients with inflammatory bowel disease - a novel finding.

    PubMed

    Mokarram, Pooneh; Kavousipour, Soudabeh; Sarabi, Mostafa Moradi; Mehrabani, Golnosh; Fahmidehkar, Mohammad Ali; Shamsdin, Seyedeh Azra; Alipour, Abbas; Naini, Mahvash Alizade

    2015-01-01

    Inflammatory bowel disease (IBD) is a disease strongly associated with colorectal cancer (CRC) as a well-known precancerous condition. Alterations in DNA methylation and mutation in K-ras are believed to play an early etiopathogenic role in CRC and may also an initiating event through deregulation of molecular signaling. Epigenetic silencing of APC and SFRP2 in the WNT signaling pathway may also be involved in IBD-CRC. The role of aberrant DNA methylation in precancerous state of colorectal cancer (CRC) is under intensive investigation worldwide. The aim of this study was to investigate the status of promoter methylation of MGMT-B, APC1A and SFRP2 genes, in inflamed and normal colon tissues of patients with IBD compared with control normal tissues. A total of 52 IBD tissues as well as corresponding normal tissues and 30 samples from healthy participants were obtained. We determined promoter methylation status of MGMT-B, SFRP2 and APC1A genes by chemical treatment with sodium bisulfite and subsequent MSP. The most frequently methylated locus was MGMT-B (71%; 34 of 48), followed by SFRP2 (66.6 %; 32 of 48), and APC1A (43.7%; 21 of 48). Our study demonstrated for the first time that hypermethylation of the MGMT-B and the SFRP2 gene promoter regions might be involved in IBD development. Methylation of MGMT-B and SFRP2 in IBD patients may provide a method for early detection of IBD-associated neoplasia. PMID:25773792

  5. CFTR is a tumor suppressor gene in murine and human intestinal cancer

    PubMed Central

    Than, BLN; Linnekamp, JF; Starr, TK; Largaespada, DA; Rod, A; Zhang, Y; Bruner, V; Abrahante, J; Schumann, A; Luczak, T; Niemczyk, A; O’Sullivan, MG; Medema, JP; Fijneman, RJA; Meijer, GA; Van den Broek, E; Hodges, CA; Scott, PM; Vermeulen, L; Cormier, RT

    2016-01-01

    CFTR, the cystic fibrosis (CF) gene, encodes for the CFTR protein that plays an essential role in anion regulation and tissue homeostasis of various epithelia. In the gastrointestinal (GI) tract CFTR promotes chloride and bicarbonate secretion, playing an essential role in ion and acid–base homeostasis. Cftr has been identified as a candidate driver gene for colorectal cancer (CRC) in several Sleeping Beauty DNA transposon-based forward genetic screens in mice. Further, recent epidemiological and clinical studies indicate that CF patients are at high risk for developing tumors in the colon. To investigate the effects of CFTR dysregulation on GI cancer, we generated ApcMin mice that carried an intestinal-specific knockout of Cftr. Our results indicate that Cftr is a tumor suppressor gene in the intestinal tract as Cftr mutant mice developed significantly more tumors in the colon and the entire small intestine. In Apc+/+ mice aged to ~ 1 year, Cftr deficiency alone caused the development of intestinal tumors in >60% of mice. Colon organoid formation was significantly increased in organoids created from Cftr mutant mice compared with wild-type controls, suggesting a potential role of Cftr in regulating the intestinal stem cell compartment. Microarray data from the Cftr-deficient colon and the small intestine identified dysregulated genes that belong to groups of immune response, ion channel, intestinal stem cell and other growth signaling regulators. These associated clusters of genes were confirmed by pathway analysis using Ingenuity Pathway Analysis and gene set enrichment analysis (GSEA). We also conducted RNA Seq analysis of tumors from Apc+/+ Cftr knockout mice and identified sets of genes dysregulated in tumors including altered Wnt β-catenin target genes. Finally we analyzed expression of CFTR in early stage human CRC patients stratified by risk of recurrence and found that loss of expression of CFTR was significantly associated with poor disease

  6. CFTR is a tumor suppressor gene in murine and human intestinal cancer.

    PubMed

    Than, B L N; Linnekamp, J F; Starr, T K; Largaespada, D A; Rod, A; Zhang, Y; Bruner, V; Abrahante, J; Schumann, A; Luczak, T; Niemczyk, A; O'Sullivan, M G; Medema, J P; Fijneman, R J A; Meijer, G A; Van den Broek, E; Hodges, C A; Scott, P M; Vermeulen, L; Cormier, R T

    2016-08-11

    CFTR, the cystic fibrosis (CF) gene, encodes for the CFTR protein that plays an essential role in anion regulation and tissue homeostasis of various epithelia. In the gastrointestinal (GI) tract CFTR promotes chloride and bicarbonate secretion, playing an essential role in ion and acid-base homeostasis. Cftr has been identified as a candidate driver gene for colorectal cancer (CRC) in several Sleeping Beauty DNA transposon-based forward genetic screens in mice. Further, recent epidemiological and clinical studies indicate that CF patients are at high risk for developing tumors in the colon. To investigate the effects of CFTR dysregulation on GI cancer, we generated Apc(Min) mice that carried an intestinal-specific knockout of Cftr. Our results indicate that Cftr is a tumor suppressor gene in the intestinal tract as Cftr mutant mice developed significantly more tumors in the colon and the entire small intestine. In Apc(+/+) mice aged to ~1 year, Cftr deficiency alone caused the development of intestinal tumors in >60% of mice. Colon organoid formation was significantly increased in organoids created from Cftr mutant mice compared with wild-type controls, suggesting a potential role of Cftr in regulating the intestinal stem cell compartment. Microarray data from the Cftr-deficient colon and the small intestine identified dysregulated genes that belong to groups of immune response, ion channel, intestinal stem cell and other growth signaling regulators. These associated clusters of genes were confirmed by pathway analysis using Ingenuity Pathway Analysis and gene set enrichment analysis (GSEA). We also conducted RNA Seq analysis of tumors from Apc(+/+) Cftr knockout mice and identified sets of genes dysregulated in tumors including altered Wnt β-catenin target genes. Finally we analyzed expression of CFTR in early stage human CRC patients stratified by risk of recurrence and found that loss of expression of CFTR was significantly associated with poor disease

  7. Modified irinotecan and infusional 5-fluorouracil (mFOLFIRI) in patients with refractory advanced pancreas cancer (APC): a single-institution experience.

    PubMed

    Bupathi, M; Ahn, D H; Wu, C; Ciombor, K K; Stephens, J A; Reardon, J; Goldstein, D A; Bekaii-Saab, T

    2016-04-01

    Pancreatic adenocarcinoma is the fourth leading cause of cancer death. Recently, MM-398 (nanoliposomal irinotecan) was shown to be associated with significant improvement in outcome measures with acceptable toxicities when combined with 5-fluorouracil (5-FU)/leucovorin (LV) compared to 5-FU/LV alone in patients failing one line of gemcitabine-based therapy. There is a paucity of data evaluating the role of irinotecan in combination with 5FU in advanced pancreas cancer (APC). We performed a retrospective analysis of all patients who received mFOLFIRI (minus bolus 5FU and LV). All patients with metastatic disease who had failed at least one line of gemcitabine-based therapy prior to receiving mFOLFIRI were included in this study. Descriptive statistics were used to assess the continuous variables and adverse events (AEs), and Kaplan-Meier methods were used to calculate the median progression-free survival (PFS) and overall survival (OS). Forty patients were included in this analysis. Patients received 1-5 lines of prior therapy (25 % with more than 3 lines of prior therapy). The mean age at diagnosis was 60, and 98 % had ECOG of 1. The mean CA 19-9 at the start of therapy was 33,169 U/ml. The median PFS was 2.59 months [95 % confidence interval (CI) (1.90, 3.54)], and OS was 4.75 months [95 % CI (3.14, 8.98)]. The most common AEs included fatigue (98 %), neuropathy (83 %), anorexia (68 %), nausea (60 %) and constipation (55 %). Grade 3 toxicities included fatigue (13 %) and rash (3 %). There were no observed grade 4 toxicities. In this single-institution retrospective analysis, mFOLFIRI was found to be both tolerable and relatively effective in a heavily pretreated patient population with APC. Future prospective studies should consider evaluating the role of mFOLFIRI in refractory APC. PMID:26995224

  8. COGENT (COlorectal cancer GENeTics) revisited

    PubMed Central

    Houlston, Richard S.

    2012-01-01

    Many colorectal cancers (CRCs) develop in genetically susceptible individuals most of whom are not carriers of germ line mismatch repair or APC gene mutations and much of the heritable risk of CRC appears to be attributable to the co-inheritance of multiple low-risk variants. The accumulated experience to date in identifying this class of susceptibility allele has highlighted the need to conduct statistically and methodologically rigorous studies and the need for the multi-centre collaboration. This has been the motivation for establishing the COGENT (COlorectal cancer GENeTics) consortium which now includes over 20 research groups in Europe, Australia, the Americas, China and Japan actively working on CRC genetics. Here, we review the rationale for identifying low-penetrance variants for CRC and the current and future challenges for COGENT. PMID:22294761

  9. Genes and Gene Therapy

    MedlinePlus

    ... a child can have a genetic disorder. Gene therapy is an experimental technique that uses genes to ... prevent disease. The most common form of gene therapy involves inserting a normal gene to replace an ...

  10. Genes and Gene Therapy

    MedlinePlus

    ... correctly, a child can have a genetic disorder. Gene therapy is an experimental technique that uses genes to ... or prevent disease. The most common form of gene therapy involves inserting a normal gene to replace an ...

  11. Evaluation and Analysis of SEASAT-A Scanning Multichannel Microwave Radiometer (SSMR) Antenna Pattern Correction (APC) Algorithm. Sub-task 4: Interim Mode T Sub B Versus Cross and Nominal Mode T Sub B

    NASA Technical Reports Server (NTRS)

    Kitzis, J. L.; Kitzis, S. N.

    1979-01-01

    The brightness temperature data produced by the SMMR Antenna Pattern Correction algorithm are evaluated. The evaluation consists of: (1) a direct comparison of the outputs of the interim, cross, and nominal APC modes; (2) a refinement of the previously determined cos beta estimates; and (3) a comparison of the world brightness temperature (T sub B) map with actual SMMR measurements.

  12. Los Alamos National Laboratory.

    ERIC Educational Resources Information Center

    Hammel, Edward F., Jr.

    1982-01-01

    Current and post World War II scientific research at the Los Alamos National Laboratory (New Mexico) is discussed. The operation of the laboratory, the Los Alamos consultant program, and continuation education, and continuing education activities at the laboratory are also discussed. (JN)

  13. A multicenter phase 1/2a dose-escalation study of the antioxidant moiety of vitamin E 2,2,5,7,8-pentamethyl-6-chromanol (APC-100) in men with advanced prostate cancer.

    PubMed

    Kyriakopoulos, Christos E; Heath, Elisabeth I; Eickhoff, Jens C; Kolesar, Jill; Yayehyirad, Mulusew; Moll, Thomas; Wilding, George; Liu, Glenn

    2016-04-01

    Background A phase 1/2a dose escalation study of APC-100 (2,2,5,7,8-Pentamethyl-6-chromanol) was conducted to determine maximum tolerated dose (MTD), recommended phase 2 dose, toxicities and efficacy in men with castrate-resistant prostate cancer (CRPC). Methods This open label phase 1/2a study utilizes a time-to-event reassessment method (TITE-CRM) design. Patients in cohorts of 3 were treated with escalating doses of APC-100 (900 mg-2400 mg) orally once daily continuously. Cycles were 28 days. Results Twenty patients with CRPC were enrolled in the dose escalation cohort. One possible DLT (elevated ALT) was seen at dose level 1. No other DLTs were seen and no dose reductions were required. Most frequent AEs included nausea (grade 1 in 6 patients) and elevated transaminases (grade 1-3 in 5 patients). After enrolment of 20 patients the MTD was not reached, however the maximal feasible dose was exceeded due to the number of capsules ingested. Five of the 20 patients had stable disease as their best response. The median progression free survival (PFS) for the cohort was 2.8 months (range 1-8). Conclusions APC-100 is a novel agent with dual mechanism of action functioning both as potent antioxidant as well as antiandrogen. No detectable APC-100 was found in the plasma at dose level 5 (2100 mg) and it was felt that maximal feasibility was nearly reached. APC-100 is being reformulated as a tablet to allow further dose escalation. Once a recommended phase 2 dose is established, future studies in prostate cancer chemoprevention should be conducted. PMID:26924129

  14. A recellularized human colon model identifies cancer driver genes.

    PubMed

    Chen, Huanhuan Joyce; Wei, Zhubo; Sun, Jian; Bhattacharya, Asmita; Savage, David J; Serda, Rita; Mackeyev, Yuri; Curley, Steven A; Bu, Pengcheng; Wang, Lihua; Chen, Shuibing; Cohen-Gould, Leona; Huang, Emina; Shen, Xiling; Lipkin, Steven M; Copeland, Neal G; Jenkins, Nancy A; Shuler, Michael L

    2016-08-01

    Refined cancer models are needed to bridge the gaps between cell line, animal and clinical research. Here we describe the engineering of an organotypic colon cancer model by recellularization of a native human matrix that contains cell-populated mucosa and an intact muscularis mucosa layer. This ex vivo system recapitulates the pathophysiological progression from APC-mutant neoplasia to submucosal invasive tumor. We used it to perform a Sleeping Beauty transposon mutagenesis screen to identify genes that cooperate with mutant APC in driving invasive neoplasia. We identified 38 candidate invasion-driver genes, 17 of which, including TCF7L2, TWIST2, MSH2, DCC, EPHB1 and EPHB2 have been previously implicated in colorectal cancer progression. Six invasion-driver genes that have not, to our knowledge, been previously described were validated in vitro using cell proliferation, migration and invasion assays and ex vivo using recellularized human colon. These results demonstrate the utility of our organoid model for studying cancer biology. PMID:27398792

  15. Stockpile Stewardship: Los Alamos

    SciTech Connect

    McMillan, Charlie; Morgan, Nathanial; Goorley, Tom; Merrill, Frank; Funk, Dave; Korzekwa, Deniece; Laintz, Ken

    2012-01-26

    "Heritage of Science" is a short video that highlights the Stockpile Stewardship program at Los Alamos National Laboratory. Stockpile Stewardship was conceived in the early 1990s as a national science-based program that could assure the safety, security, and effectiveness of the U.S. nuclear deterrent without the need for full-scale underground nuclear testing. This video was produced by Los Alamos National Laboratory for screening at the Lab's Bradbury Science Museum in Los Alamos, NM and is narrated by science correspondent Miles O'Brien.

  16. Stockpile Stewardship: Los Alamos

    ScienceCinema

    McMillan, Charlie; Morgan, Nathanial; Goorley, Tom; Merrill, Frank; Funk, Dave; Korzekwa, Deniece; Laintz, Ken

    2014-08-12

    "Heritage of Science" is a short video that highlights the Stockpile Stewardship program at Los Alamos National Laboratory. Stockpile Stewardship was conceived in the early 1990s as a national science-based program that could assure the safety, security, and effectiveness of the U.S. nuclear deterrent without the need for full-scale underground nuclear testing. This video was produced by Los Alamos National Laboratory for screening at the Lab's Bradbury Science Museum in Los Alamos, NM and is narrated by science correspondent Miles O'Brien.

  17. Gene Therapy Induces Antigen-Specific Tolerance in Experimental Collagen-Induced Arthritis

    PubMed Central

    Jirholt, Pernilla; Turesson, Olof; Wing, Kajsa; Holmdahl, Rikard; Kihlberg, Jan; Stern, Anna; Mårtensson, Inga-Lill; Henningsson, Louise; Gustafsson, Kenth; Gjertsson, Inger

    2016-01-01

    Here, we investigate induction of immunological tolerance by lentiviral based gene therapy in a mouse model of rheumatoid arthritis, collagen II-induced arthritis (CIA). Targeting the expression of the collagen type II (CII) to antigen presenting cells (APCs) induced antigen-specific tolerance, where only 5% of the mice developed arthritis as compared with 95% of the control mice. In the CII-tolerized mice, the proportion of Tregs as well as mRNA expression of SOCS1 (suppressors of cytokine signaling 1) increased at day 3 after CII immunization. Transfer of B cells or non-B cell APC, as well as T cells, from tolerized to naïve mice all mediated a certain degree of tolerance. Thus, sustainable tolerance is established very early during the course of arthritis and is mediated by both B and non-B cells as APCs. This novel approach for inducing tolerance to disease specific antigens can be used for studying tolerance mechanisms, not only in CIA but also in other autoimmune diseases. PMID:27159398

  18. Detection of a Tumor Suppressor Gene Variant Predisposing to Colorectal Cancer in an 18th Century Hungarian Mummy

    PubMed Central

    Feldman, Michal; Hershkovitz, Israel; Sklan, Ella H.; Kahila Bar-Gal, Gila; Pap, Ildikó; Szikossy, Ildikó; Rosin-Arbesfeld, Rina

    2016-01-01

    Mutations of the Adenomatous polyposis coli (APC) gene are common and strongly associated with the development of colorectal adenomas and carcinomas. While extensively studied in modern populations, reports on visceral tumors in ancient populations are scarce. To the best of our knowledge, genetic characterization of mutations associated with colorectal cancer in ancient specimens has not yet been described. In this study we have sequenced hotspots for mutations in the APC gene isolated from 18th century naturally preserved human Hungarian mummies. While wild type APC sequences were found in two mummies, we discovered the E1317Q missense mutation, known to be a colorectal cancer predisposing mutation, in a large intestine tissue of an 18th century mummy. Our data suggests that this genetic predisposition to cancer already existed in the pre-industrialization era. This study calls for similar investigations of ancient specimens from different periods and geographical locations to be conducted and shared for the purpose of obtaining a larger scale analysis that will shed light on past cancer epidemiology and on cancer evolution. PMID:26863316

  19. HLA-DQ2.5 genes associated with celiac disease risk are preferentially expressed with respect to non-predisposing HLA genes: Implication for anti-gluten T cell response.

    PubMed

    Pisapia, Laura; Camarca, Alessandra; Picascia, Stefania; Bassi, Virginia; Barba, Pasquale; Del Pozzo, Giovanna; Gianfrani, Carmen

    2016-06-01

    HLA genes represent the main risk factor in autoimmune disorders. In celiac disease (CD), the great majority of patients carry the HLA DQA1*05 and DQB1*02 alleles, both of which encode the DQ2.5 molecule. The formation of complexes between DQ2.5 and gluten peptides on antigen-presenting cells (APCs) is necessary to activate pathogenic CD4(+) T lymphocytes. It is widely accepted that the DQ2.5 genes establish the different intensities of anti-gluten immunity, depending whether they are in a homozygous or a heterozygous configuration. Here, we demonstrated that HLA DQA1*05 and DQB1*02 gene expression is much higher than expression of non-CD-associated genes. This influences the protein levels and causes a comparable cell surface exposure of DQ2.5 heterodimers between DQ2.5 homozygous and heterozygous celiac patients. As a consequence, the magnitude of the anti-gluten CD4(+) T cell response is strictly dependent on the antigen dose and not on the DQ2.5 gene configuration of APCs. Furthermore, our findings support the concept that the expression of DQ2.5 genes is an important risk factor in celiac disease. The preferential expression of DQ2.5 alleles provides a new functional explanation of why these genes are so frequently associated with celiac disease and with other autoimmune disorders. PMID:27083396

  20. Evaluation and analysis of Seasat-A Scanning multichannel Microwave radiometer (SMMR) Antenna Pattern Correction (APC) algorithm. Sub-task 2: T sub B measured vs. T sub B calculated comparison results

    NASA Technical Reports Server (NTRS)

    Kitzis, J. L.; Kitzis, S. N.

    1979-01-01

    Interim Antenna Pattern Correction (APC) brightness temperature measurements for all ten SMMR channels are compared with calculated values generated from surface truth data. Plots and associated statistics are presented for the available points of coincidence between SMMR and surface truth measurements acquired for the Gulf of Alaska SEASAT Experiment. The most important conclusions of the study deal with the apparent existence of different instrument biases for each SMMR channel, and their variation across the scan.

  1. Diverse HLA-I Peptide Repertoires of the APC Lines MUTZ3-Derived Immature and Mature Dendritic Cells and THP1-Derived Macrophages.

    PubMed

    Nyambura, Lydon Wainaina; Jarmalavicius, Saulius; Baleeiro, Renato Brito; Walden, Peter

    2016-09-15

    Dendritic cells (DCs) and macrophages are specialized APCs that process and present self-Ags for induction of tolerance and foreign Ags to initiate T cell-mediated immunity. Related to differentiation states they have specific phenotypes and functions. However, the impact of these differentiations on Ag processing and presentation remains poorly defined. To gain insight into this, we analyzed and compared the HLA-I peptidomes of MUTZ3-derived human immature and mature DC lines and THP1-derived macrophages by liquid chromatography tandem mass spectrometry. We found that the HLA-I peptidomes were heterogeneous and individualized and were dominated by nonapeptides with similar HLA-I binding affinities and anchor residues. MUTZ3-derived DCs and THP1-derived macrophages were able to sample peptides from source proteins of almost all subcellular locations and were involved in various cellular functions in similar proportion, with preference to proteins involved in cell communication, signal transduction, protein metabolism, and transcription factor/regulator activity. PMID:27543614

  2. Distinct APC Subtypes Drive Spatially Segregated CD4+ and CD8+ T-Cell Effector Activity during Skin Infection with HSV-1

    PubMed Central

    Macleod, Bethany L.; Bedoui, Sammy; Hor, Jyh Liang; Mueller, Scott N.; Russell, Tiffany A.; Hollett, Natasha A.; Heath, William R.; Tscharke, David C.

    2014-01-01

    Efficient infection control requires potent T-cell responses at sites of pathogen replication. However, the regulation of T-cell effector function in situ remains poorly understood. Here, we show key differences in the regulation of effector activity between CD4+ and CD8+ T-cells during skin infection with HSV-1. IFN-γ-producing CD4+ T cells disseminated widely throughout the skin and draining lymph nodes (LN), clearly exceeding the epithelial distribution of infectious virus. By contrast, IFN-γ-producing CD8+ T cells were only found within the infected epidermal layer of the skin and associated hair follicles. Mechanistically, while various subsets of lymphoid- and skin-derived dendritic cells (DC) elicited IFN-γ production by CD4+ T cells, CD8+ T cells responded exclusively to infected epidermal cells directly presenting viral antigen. Notably, uninfected cross-presenting DCs from both skin and LNs failed to trigger IFN-γ production by CD8+ T-cells. Thus, we describe a previously unappreciated complexity in the regulation of CD4+ and CD8+ T-cell effector activity that is subset-specific, microanatomically distinct and involves largely non-overlapping types of antigen-presenting cells (APC). PMID:25121482

  3. CD24 knockout prevents colorectal cancer in chemically induced colon carcinogenesis and in APC(Min)/CD24 double knockout transgenic mice.

    PubMed

    Naumov, Inna; Zilberberg, Alona; Shapira, Shiran; Avivi, Doran; Kazanov, Dina; Rosin-Arbesfeld, Rina; Arber, Nadir; Kraus, Sarah

    2014-09-01

    Increased expression of CD24 is seen in a large variety of solid tumors, including up to 90% of gastrointestinal (GI) tumors. Stable derivatives of SW480 colorectal cancer (CRC) cells that overexpress CD24 proliferate faster, and increase cell motility, saturation density, plating efficiency, and growth in soft agar. They also produce larger tumors in nude mice as compared to the parental SW480 cells. Most significantly, even depletion of one copy of the CD24 allele in the APC(Min/+) mice of a transgenic mouse model led to a dramatic reduction in tumor burden in all sections of the small intestine. Homozygous deletion of both CD24 alleles resulted in complete abolishment of tumor formation. Moreover, CD24 knockout mice exhibited resistance to chemically induced inflammation-associated CRC. Finally, a new signal transduction pathway is suggested: namely, CD24 expression downstream to COX2 and PGE2 synthesis, which is directly regulated by β-catenin. CD24 is shown in vitro and in vivo as being an important oncogene in the gut, and one that plays a critical role in the initiation and progression of carcinogenesis. PMID:24500912

  4. Los Alamos offers Fellowships

    NASA Astrophysics Data System (ADS)

    Los Alamos National Laboratory in New Mexico is calling for applications for postdoctoral appointments and research fellowships. The positions are available in geoscience as well as other scientific disciplines.The laboratory, which is operated by the University of California for the Department of Energy, awards J. Robert Oppenheimer Research Fellowships to scientists that either have or will soon complete doctoral degrees. The appointments are for two years, are renewable for a third year, and carry a stipend of $51,865 per year. Potential applicants should send a resume or employment application and a statement of research goals to Carol M. Rich, Div. 89, Human Resources Development Division, MS P290, Los Alamos National Laboratory, Los Alamos, New Mexico 87545 by mid-November.

  5. Human adipose-derived mesenchymal stem cells engineered to secrete IL-10 inhibit APC function and limit CNS autoimmunity.

    PubMed

    Payne, Natalie L; Sun, Guizhi; McDonald, Courtney; Moussa, Leon; Emerson-Webber, Ashley; Loisel-Meyer, Séverine; Medin, Jeffrey A; Siatskas, Christopher; Bernard, Claude C A

    2013-05-01

    Interleukin (IL)-10 is an important immunoregulatory cytokine shown to impact inflammatory processes as manifested in patients with multiple sclerosis (MS) and in its animal model, experimental autoimmune encephalomyelitis (EAE). Several lines of evidence indicate that the effectiveness of IL-10-based therapies may be dependent on the timing and mode of delivery. In the present study we engineered the expression of IL-10 in human adipose-derived mesenchymal stem cells (Adi-IL-10-MSCs) and transplanted these cells early in the disease course to mice with EAE. Adi-IL-10-MSCs transplanted via the intraperitoneal route prevented or delayed the development of EAE. This protective effect was associated with several anti-inflammatory response mechanisms, including a reduction in peripheral T-cell proliferative responses, a decrease in pro-inflammatory cytokine secretion as well as a preferential inhibition of Th17-mediated neuroinflammation. In vitro analyses revealed that Adi-IL-10-MSCs inhibited the phenotypic maturation, cytokine production and antigen presenting capacity of bone marrow-derived myeloid dendritic cells, suggesting that the mechanism of action may involve an indirect effect on pathogenic T-cells via the modulation of antigen presenting cell function. Collectively, these results suggest that early intervention with gene modified Adi-MSCs may be beneficial for the treatment of autoimmune diseases such as MS. PMID:23369732

  6. Synthesis, characterization, molecular modeling and antioxidant activity of (1E,5E)-1,5-bis(1-(pyridin-2-yl)ethylidene)carbonohydrazide (H2APC) and its zinc(II), cadmium(II) and mercury(II) complexes

    NASA Astrophysics Data System (ADS)

    El-Gammal, O. A.; Abu El-Reash, G. M.; Ghazy, S. E.; Radwan, A. H.

    2012-08-01

    A new series of Zn(II), Cd(II) and Hg(II) complexes of (1E,5E)-1,5-bis(1-(pyridin-2-yl)ethylidene)carbonohydrazide (H2APC) have been prepared and characterized by elemental analyses, spectral (IR, UV-visible, mass and 1H NMR) as well as magnetic and thermal measurements. The data revealed that the ligand acts a monobasic hexadentate, neutral tri- and monodentate in Zn(II), Cd(II) and Hg(II) complexes, respectively. An octahedral geometry is proposed for Zn(II) complex, a trigonal bi-pyramid for Cd(II) complex and a tetrahedral one for Hg(II) complex. The bond length, bond angle, HOMO, LUMO and charges on the atoms have been calculated to confirm the geometry of the ligand and the investigated complexes using material studio program. Kinetic parameters were determined for each thermal degradation stage of some complexes using Coats-Redfern and Horowitz-Metzger methods. The antioxidant, anti-hemolytic, and cytotoxic activities of the compounds have been screened. H2APC showed moderate antioxidant activity using ABTS and DPPH methods. With respect to erythrocyte hemolysis and in vitro Ehrlich ascites assay, H2APC exhibited the potent antioxidative activity followed by Cd(II) and Zn(II) complexes while Hg(II) complex showed very weak activity.

  7. Malignancy of Cancers and Synthetic Lethal Interactions Associated With Mutations of Cancer Driver Genes

    PubMed Central

    Wang, Xiaosheng; Zhang, Yue; Han, Ze-Guang; He, Kun-Yan

    2016-01-01

    Abstract The mutation status of cancer driver genes may correlate with different degrees of malignancy of cancers. The doubling time and multidrug resistance are 2 phenotypes that reflect the degree of malignancy of cancer cells. Because most of cancer driver genes are hard to target, identification of their synthetic lethal partners might be a viable approach to treatment of the cancers with the relevant mutations. The genome-wide screening for synthetic lethal partners is costly and labor intensive. Thus, a computational approach facilitating identification of candidate genes for a focus synthetic lethal RNAi screening will accelerate novel anticancer drug discovery. We used several publicly available cancer cell lines and tumor tissue genomic data in this study. We compared the doubling time and multidrug resistance between the NCI-60 cell lines with mutations in some cancer driver genes and those without the mutations. We identified some candidate synthetic lethal genes to the cancer driver genes APC, KRAS, BRAF, PIK3CA, and TP53 by comparison of their gene phenotype values in cancer cell lines with the relevant mutations and wild-type background. Further, we experimentally validated some of the synthetic lethal relationships we predicted. We reported that mutations in some cancer driver genes mutations in some cancer driver genes such as APC, KRAS, or PIK3CA might correlate with cancer proliferation or drug resistance. We identified 40, 21, 5, 43, and 18 potential synthetic lethal genes to APC, KRAS, BRAF, PIK3CA, and TP53, respectively. We found that some of the potential synthetic lethal genes show significantly higher expression in the cancers with mutations of their synthetic lethal partners and the wild-type counterparts. Further, our experiments confirmed several synthetic lethal relationships that are novel findings by our methods. We experimentally validated a part of the synthetic lethal relationships we predicted. We plan to perform further

  8. The Los Alamos primer

    SciTech Connect

    Serber, R.

    1992-01-01

    This book contains the 1943 lecture notes of Robert Serber. Serber was a protege of J. Robert Oppenheimer and member of the team that built the first atomic bomb - reveal what the Los Alamos scientists knew, and did not know, about the terrifying weapon they were building.

  9. Fasting protects against the side effects of irinotecan but preserves its anti-tumor effect in Apc15lox mutant mice

    PubMed Central

    Huisman, Sander A; Bijman-Lagcher, Wendy; IJzermans, Jan NM; Smits, Ron; de Bruin, Ron WF

    2015-01-01

    Irinotecan is a widely used topoisomerase-I-inhibitor with a very narrow therapeutic window because of its severe toxicity. In the current study we have examined the effects of fasting prior to irinotecan treatment on toxicity and anti-tumor activity. FabplCre;Apc15lox/+ mice, which spontaneously develop intestinal tumors, of 27 weeks of age were randomized into 3-day fasted and ad libitum fed groups, followed by treatment with a flat-fixed high dose of irinotecan or vehicle. Side-effects were recorded until 11 days after the start of the experiment. Tumor size, and markers for cell-cycle activity, proliferation, angiogenesis, and senescence were measured. Fasted mice were protected against the side-effects of irinotecan treatment. Ad libitum fed mice developed visible signs of discomfort including weight loss, lower activity, ruffled coat, hunched-back posture, diarrhea, and leukopenia. Irinotecan reduced tumor size in fasted and ad libitum fed groups similarly compared to untreated controls (2.4 ± 0.67 mm and 2.4 ± 0.82 mm versus 3.0 ± 1.05 mm and 2.8 ± 1.08 mm respectively, P < 0.001). Immunohistochemical analysis showed reduced proliferation, a reduced number of vascular endothelial cells, and increased levels of senescence in tumors of both irinotecan treated groups. In conclusion, 3 days of fasting protects against the toxic side-effects of irinotecan in a clinically relevant mouse model of spontaneously developing colorectal cancer without affecting its anti-tumor activity. These results support fasting as a powerful way to improve treatment of colorectal carcinoma patients. PMID:25955194

  10. TGF-β activates APC through Cdh1 binding for Cks1 and Skp2 proteasomal destruction stabilizing p27kip1 for normal endometrial growth.

    PubMed

    Pavlides, Savvas C; Lecanda, Jon; Daubriac, Julien; Pandya, Unnati M; Gama, Patricia; Blank, Stephanie; Mittal, Khushbakhat; Shukla, Pratibha; Gold, Leslie I

    2016-04-01

    We previously reported that aberrant TGF-β/Smad2/3 signaling in endometrial cancer (ECA) leads to continuous ubiquitylation of p27(kip1)(p27) by the E3 ligase SCF-Skp2/Cks1 causing its degradation, as a putative mechanism involved in the pathogenesis of this cancer. In contrast, normal intact TGF-β signaling prevents degradation of nuclear p27 by SCF-Skp2/Cks1 thereby accumulating p27 to block Cdk2 for growth arrest. Here we show that in ECA cell lines and normal primary endometrial epithelial cells, TGF-β increases Cdh1 and its binding to APC/C to form the E3 ligase complex that ubiquitylates Cks1 and Skp2 prompting their proteasomal degradation and thus, leaving p27 intact. Knocking-down Cdh1 in ECA cell lines increased Skp2/Cks1 E3 ligase activity, completely diminished nuclear and cytoplasmic p27, and obviated TGF-β-mediated inhibition of proliferation. Protein synthesis was not required for TGF-β-induced increase in nuclear p27 and decrease in Cks1 and Skp2. Moreover, half-lives of Cks1 and Skp2 were extended in the Cdh1-depleted cells. These results suggest that the levels of p27, Skp2 and Cks1 are strongly or solely regulated by proteasomal degradation. Finally, an inverse relationship of low p27 and high Cks1 in the nucleus was shown in patients in normal proliferative endometrium and grade I-III ECAs whereas differentiated secretory endometrium showed the reverse. These studies implicate Cdh1 as the master regulator of TGF-β-induced preservation of p27 tumor suppressor activity. Thus, Cdh1 is a potential therapeutic target for ECA and other human cancers showing an inverse relationship between Cks1/Skp2 and p27 and/or dysregulated TGF-β signaling. PMID:26963853

  11. Mouse models for the discovery of colorectal cancer driver genes

    PubMed Central

    Clark, Christopher R; Starr, Timothy K

    2016-01-01

    Colorectal cancer (CRC) constitutes a major public health problem as the third most commonly diagnosed and third most lethal malignancy worldwide. The prevalence and the physical accessibility to colorectal tumors have made CRC an ideal model for the study of tumor genetics. Early research efforts using patient derived CRC samples led to the discovery of several highly penetrant mutations (e.g., APC, KRAS, MMR genes) in both hereditary and sporadic CRC tumors. This knowledge has enabled researchers to develop genetically engineered and chemically induced tumor models of CRC, both of which have had a substantial impact on our understanding of the molecular basis of CRC. Despite these advances, the morbidity and mortality of CRC remains a cause for concern and highlight the need to uncover novel genetic drivers of CRC. This review focuses on mouse models of CRC with particular emphasis on a newly developed cancer gene discovery tool, the Sleeping Beauty transposon-based mutagenesis model of CRC. PMID:26811627

  12. Genes and gene regulation

    SciTech Connect

    MacLean, N.

    1988-01-01

    Genetics has long been a central topic for biologists, and recent progress has captured the public imagination as well. This book addresses questions that are at the leading edge of this continually advancing discipline. In tune with the increasing emphasis on molecular biology and genetic engineering, this text emphasizes the molecular aspects of gene expression, and the evolution of gene sequence organization and control. It reviews the genetic material of viruses, bacteria, and of higher organisms. Cells and organisms are compared in terms of gene numbers, their arrangements within a cell, and the control mechanisms which regulate the activity of genes.

  13. Aberrant DNA methylation of WNT pathway genes in the development and progression of CIMP-negative colorectal cancer.

    PubMed

    Galamb, Orsolya; Kalmár, Alexandra; Péterfia, Bálint; Csabai, István; Bodor, András; Ribli, Dezső; Krenács, Tibor; Patai, Árpád V; Wichmann, Barnabás; Barták, Barbara Kinga; Tóth, Kinga; Valcz, Gábor; Spisák, Sándor; Tulassay, Zsolt; Molnár, Béla

    2016-08-01

    The WNT signaling pathway has an essential role in colorectal carcinogenesis and progression, which involves a cascade of genetic and epigenetic changes. We aimed to analyze DNA methylation affecting the WNT pathway genes in colorectal carcinogenesis in promoter and gene body regions using whole methylome analysis in 9 colorectal cancer, 15 adenoma, and 6 normal tumor adjacent tissue (NAT) samples by methyl capture sequencing. Functional methylation was confirmed on 5-aza-2'-deoxycytidine-treated colorectal cancer cell line datasets. In parallel with the DNA methylation analysis, mutations of WNT pathway genes (APC, β-catenin/CTNNB1) were analyzed by 454 sequencing on GS Junior platform. Most differentially methylated CpG sites were localized in gene body regions (95% of WNT pathway genes). In the promoter regions, 33 of the 160 analyzed WNT pathway genes were differentially methylated in colorectal cancer vs. normal, including hypermethylated AXIN2, CHP1, PRICKLE1, SFRP1, SFRP2, SOX17, and hypomethylated CACYBP, CTNNB1, MYC; 44 genes in adenoma vs. NAT; and 41 genes in colorectal cancer vs. adenoma comparisons. Hypermethylation of AXIN2, DKK1, VANGL1, and WNT5A gene promoters was higher, while those of SOX17, PRICKLE1, DAAM2, and MYC was lower in colon carcinoma compared to adenoma. Inverse correlation between expression and methylation was confirmed in 23 genes, including APC, CHP1, PRICKLE1, PSEN1, and SFRP1. Differential methylation affected both canonical and noncanonical WNT pathway genes in colorectal normal-adenoma-carcinoma sequence. Aberrant DNA methylation appears already in adenomas as an early event of colorectal carcinogenesis. PMID:27245242

  14. Preparation and characterization of nonpolar fluorinated carbosilane dendrimers by APcI mass spectrometry and small-angle X-ray scattering

    SciTech Connect

    Omotowa, B.A.; Keefer, K.D.; Kirchmeier, R.L.; Shreeve, J.M.

    1999-12-08

    The following highly fluorinated nonpolar dendrimers were obtained in high yields from multiple hydrosilylation reactions between core hydride terminated carbosilane dendrimers and allyl-1,1-dihydrotrifluoroethyl ether or allyl-1,1-dihydroheptadecafluorononyl ether through divergent synthetic routes: Si[CH{sub 2}CH{sub 2}SiMe{sub 2}(CH{sub 2}CH{sub 2}CH{sub 2}OCH{sub 2}CF{sub 3})]{sub 4} (7), Si{l{underscore}brace}CH{sub 2}CH{sub 2}SiMe[CH{sub 2}{l{underscore}brace}CH{sub 2}SiMe(CH{sub 2}CH{sub 2}CH{sub 2}OCH{sub 2}CF{sub 3}){sub 2}]{sub 2}{r{underscore}brace}{sub 4} (8), Si[CH{sub 2}CH{sub 2}Si(CH{sub 2}CH{sub 2}CH{sub 2}OCH{sub 2}C{sub 8}F{sub 17}){sub 3}]{sub 4} (9), Si[CH{sub 2}CH{sub 2}SiMe{sub 2}(CH{sub 2}CH{sub 2}OCH{sub 2}C{sub 8}F{sub 17})]{sub 4} (10), and Si{l{underscore}brace}CH{sub 2}CH{sub 2}Si[CH{sub 2}CH{sub 2}Si(CH{sub 2}CH{sub 2}OCH{sub 2}C{sub 8}F{sub 17}){sub 3}]{sub 3}{r{underscore}brace}{sub 4} (11). These compounds were characterized by elemental and spectroscopic analyses. Valuable mass spectral data were obtained by using atmospheric pressure chemical ionization (APcI). The fluorinated dendrimer molecule and the nonfluorinated core scatter X-ray light differently and present unique slopes on the Guinier Plot of data from small-angle X-ray light scattering (SAXS) in hexafluorobenzene. Glass transition temperatures (T{sub g}) and thermogravimetric analyses (TGA) of the dendrimers were determined.

  15. Studying Genes

    MedlinePlus

    ... Area What are genes? Genes are sections of DNA that contain instructions for making the molecules—many ... material in an organism. This includes genes and DNA elements that control the activity of genes. Does ...

  16. Promoter methylation status of tumor suppressor genes and inhibition of expression of DNA methyltransferase 1 in non-small cell lung cancer.

    PubMed

    Liu, Bangqing; Song, Jianfei; Luan, Jiaqiang; Sun, Xiaolin; Bai, Jian; Wang, Haiyong; Li, Angui; Zhang, Lifei; Feng, Xiaoyan; Du, Zhenzong

    2016-08-01

    DNA methylation is an epigenetic DNA modification catalyzed by DNA methyltransferase 1 (DNMT1). The purpose of this study was to investigate DNMT1 gene and protein expression and the effects of methylation status on tumor suppressor genes in human non-small cell lung cancer (NSCLC) cell lines grown in vitro and in vivo Human lung adenocarcinoma cell lines, A549 and H838, were grown in vitro and inoculated subcutaneously into nude mice to form tumors and were then treated with the DNA methylation inhibitor, 5-aza-2'-deoxycytidine, with and without treatment with the benzamide histone deacetylase inhibitor, entinostat (MS-275). DNMT1 protein expression was quantified by Western blot. Promoter methylation status of tumor suppressor genes (RASSF1A, ASC, APC, MGMT, CDH13, DAPK, ECAD, P16, and GATA4) was evaluated by methylation-specific polymerase chain reaction. Methylation status of the tumor suppressor genes was regulated by the DNMT1 gene, with the decrease of DNMT1 expression following DNA methylation treatment. Demethylation of tumor suppressor genes (APC, ASC, and RASSF1A) restored tumor growth in nude mice. The results of this study support a role for methylation of DNA as a potential epigenetic clinical biomarker of prognosis or response to therapy and for DNMT1 as a potential therapeutic target in NSCLC. PMID:27190263

  17. Los Alamos National Laboratory

    SciTech Connect

    Dogliani, Harold O

    2011-01-19

    The purpose of the briefing is to describe general laboratory technical capabilities to be used for various groups such as military cadets or university faculty/students and post docs to recruit into a variety of Los Alamos programs. Discussed are: (1) development and application of high leverage science to enable effeictive, predictable and reliability outcomes; (2) deter, detect, characterize, reverse and prevent the proliferation of weapons of mass destruction and their use by adversaries and terrorists; (3) modeling and simulation to define complex processes, predict outcomes, and develop effective prevention, response, and remediation strategies; (4) energetic materials and hydrodynamic testing to develop materials for precise delivery of focused energy; (5) materials cience focused on fundamental understanding of materials behaviors, their quantum-molecular properties, and their dynamic responses, and (6) bio-science to rapidly detect and characterize pathogens, to develop vaccines and prophylactic remedies, and to develop attribution forensics.

  18. Los biocombustibles y el futuro

    NASA Video Gallery

    ¿Cómo podremos utilizar los biocombustibles en el futuro? La ingeniera aeroespacial de la NASA, Diana Centeno Gómez nos explica el futuro de los biocombustibles y cómo un día podrías trabajar con d...

  19. Quantitative DNA methylation analysis of candidate genes in cervical cancer.

    PubMed

    Siegel, Erin M; Riggs, Bridget M; Delmas, Amber L; Koch, Abby; Hakam, Ardeshir; Brown, Kevin D

    2015-01-01

    Aberrant DNA methylation has been observed in cervical cancer; however, most studies have used non-quantitative approaches to measure DNA methylation. The objective of this study was to quantify methylation within a select panel of genes previously identified as targets for epigenetic silencing in cervical cancer and to identify genes with elevated methylation that can distinguish cancer from normal cervical tissues. We identified 49 women with invasive squamous cell cancer of the cervix and 22 women with normal cytology specimens. Bisulfite-modified genomic DNA was amplified and quantitative pyrosequencing completed for 10 genes (APC, CCNA, CDH1, CDH13, WIF1, TIMP3, DAPK1, RARB, FHIT, and SLIT2). A Methylation Index was calculated as the mean percent methylation across all CpG sites analyzed per gene (~4-9 CpG site) per sequence. A binary cut-point was defined at >15% methylation. Sensitivity, specificity and area under ROC curve (AUC) of methylation in individual genes or a panel was examined. The median methylation index was significantly higher in cases compared to controls in 8 genes, whereas there was no difference in median methylation for 2 genes. Compared to HPV and age, the combination of DNA methylation level of DAPK1, SLIT2, WIF1 and RARB with HPV and age significantly improved the AUC from 0.79 to 0.99 (95% CI: 0.97-1.00, p-value = 0.003). Pyrosequencing analysis confirmed that several genes are common targets for aberrant methylation in cervical cancer and DNA methylation level of four genes appears to increase specificity to identify cancer compared to HPV detection alone. Alterations in DNA methylation of specific genes in cervical cancers, such as DAPK1, RARB, WIF1, and SLIT2, may also occur early in cervical carcinogenesis and should be evaluated. PMID:25826459

  20. Quantitative DNA Methylation Analysis of Candidate Genes in Cervical Cancer

    PubMed Central

    Siegel, Erin M.; Riggs, Bridget M.; Delmas, Amber L.; Koch, Abby; Hakam, Ardeshir; Brown, Kevin D.

    2015-01-01

    Aberrant DNA methylation has been observed in cervical cancer; however, most studies have used non-quantitative approaches to measure DNA methylation. The objective of this study was to quantify methylation within a select panel of genes previously identified as targets for epigenetic silencing in cervical cancer and to identify genes with elevated methylation that can distinguish cancer from normal cervical tissues. We identified 49 women with invasive squamous cell cancer of the cervix and 22 women with normal cytology specimens. Bisulfite-modified genomic DNA was amplified and quantitative pyrosequencing completed for 10 genes (APC, CCNA, CDH1, CDH13, WIF1, TIMP3, DAPK1, RARB, FHIT, and SLIT2). A Methylation Index was calculated as the mean percent methylation across all CpG sites analyzed per gene (~4-9 CpG site) per sequence. A binary cut-point was defined at >15% methylation. Sensitivity, specificity and area under ROC curve (AUC) of methylation in individual genes or a panel was examined. The median methylation index was significantly higher in cases compared to controls in 8 genes, whereas there was no difference in median methylation for 2 genes. Compared to HPV and age, the combination of DNA methylation level of DAPK1, SLIT2, WIF1 and RARB with HPV and age significantly improved the AUC from 0.79 to 0.99 (95% CI: 0.97–1.00, p-value = 0.003). Pyrosequencing analysis confirmed that several genes are common targets for aberrant methylation in cervical cancer and DNA methylation level of four genes appears to increase specificity to identify cancer compared to HPV detection alone. Alterations in DNA methylation of specific genes in cervical cancers, such as DAPK1, RARB, WIF1, and SLIT2, may also occur early in cervical carcinogenesis and should be evaluated. PMID:25826459

  1. La inserción en el mercado laboral de los inmigrantes latinos en España y en los Estados Unidos: Diferencias por país de origen y estatus legal

    PubMed Central

    Connor, Phillip; Massey, Douglas

    2013-01-01

    Resumen Este artículo compara los resultados económicos entre los inmigrantes latinoamericanos en España y Estados Unidos. Detectamos un efecto de selección por el que la mayoría de los inmigrantes latinoamericanos en España proceden de Sudamérica de un entorno de clases medias, mientras la mayoría de los inmigrantes que van a los Estados Unidos son centroamericanos de clase baja. Este efecto de selección explica las diferencias transnacionales en la probabilidad de empleo, logro ocupacional y salarios obtenidos. A pesar de las diferencias en los orígenes y las características de los latinoamericanos en ambos países, los factores demográficos, humanos y de capital social parecen operar de forma similar en ambos países; y cuando los modelos se estiman separadamente por estatus legal, descubrimos que los efectos se acentúan más entre los inmigrantes irregulares cuando se los compara con los regulares, especialmente en Estados Unidos. PMID:24532857

  2. Los Alamos National Laboratory Overview

    SciTech Connect

    Neu, Mary

    2010-06-02

    Mary Neu, Associate Director for Chemistry, Life and Earth Sciences at Los Alamos National Laboratory, delivers opening remarks at the "Sequencing, Finishing, Analysis in the Future" meeting in Santa Fe, NM

  3. Distribution of the hallucinogens N,N-dimethyltryptamine and 5-methoxy-N,N-dimethyltryptamine in rat brain following intraperitoneal injection: application of a new solid-phase extraction LC-APcI-MS-MS-isotope dilution method.

    PubMed

    Barker, S A; Littlefield-Chabaud, M A; David, C

    2001-02-10

    A method for the solid-phase extraction (SPE) and liquid chromatographic-atmospheric pressure chemical ionization-mass spectrometric-mass spectrometric-isotope dilution (LC-APcI-MS-MS-ID) analysis of the indole hallucinogens N,N-dimethyltryptamine (DMT) and 5-methoxy DMT (or O-methyl bufotenin, OMB) from rat brain tissue is reported. Rats were administered DMT or OMB by the intraperitoneal route at a dose of 5 mg/kg and sacrificed 15 min post treatment. Brains were dissected into discrete areas and analyzed by the methods described as a demonstration of the procedure's applicability. The synthesis and use of two new deuterated internal standards for these purposes are also reported. PMID:11232854

  4. Gene-specific promoter methylation is associated with micronuclei frequency in urothelial cells from individuals exposed to organic solvents and paints.

    PubMed

    Hoyos-Giraldo, L S; Escobar-Hoyos, L F; Saavedra-Trujillo, D; Reyes-Carvajal, I; Muñoz, A; Londoño-Velasco, E; Tello, A; Cajas-Salazar, N; Ruíz, M; Carvajal, S; Santella, R M

    2016-05-01

    Sufficient epidemiologic evidence has established an etiologic link between bladder cancer risk and occupational exposure as a painter to organic solvents. Currently, it remains to be established whether gene-specific promoter methylation contributes to bladder cancer development, including by enhancing chromosome breakage or loss. We investigated the effect of chronic exposure to organic solvents and paints on DNA methylation profiles in the promoter regions of four genes (GSTP1, p16(INK4a), APC and CDH1) and micronucleus (MN) frequency in exfoliated urothelial cells from voided urine from Colombian male non-smoking car painters and age-matched unexposed individuals. The exposed group had a higher percentage of individuals with >2 MNs/2000 cells compared with the unexposed group (P=0.04). Gene-specific analysis showed a significantly higher percentage of individuals with methylated GSTP1, p16(INK4a) and APC in the exposed group. Poisson regression analysis indicated that exposed individuals with methylated GSTP1 and p16(INK4a) promoters were more than twofold more likely to have an increase in MN frequency as compared with the reference. Finally, among exposed individuals with GSTP1 and p16(INK4a) methylated promoters, those with a greater age had a higher RR of increased MN frequency compared with younger exposed individuals with methylated promoters. These results support the conclusion that gene-specific promoter methylation may increase MN frequency in a dependent or independent interaction with occupational exposure to organic solvents. PMID:25993025

  5. Initial formal toxicity evaluation of APC-2, a novel fluorescent tracer agent for real-time measurement of glomerular filtration rate in preparation for a first-in-man clinical trial

    NASA Astrophysics Data System (ADS)

    Bugaj, Joseph E.; Dorshow, Richard B.

    2014-03-01

    The fluorescent tracer agent 2,5-bis[N-(1-carboxy-2-hydroxy)]carbamoyl-3,6-diaminopyrazine, designated APC-2, has been developed with properties and attributes necessary for use as a direct measure of glomerular filtration rate (GFR). Comparison to known standard exogenous GFR agents in animal models has demonstrated an excellent correlation. A clinical trial to demonstrate this same correlation in humans is in preparation. A battery of formal toxicity tests necessary for regulatory clearance to proceed with a clinical trial has been recently completed on this new fluorescent tracer agent. These include single dose toxicity studies in rats and dogs to determine overall toxicity and toxicokinetics of the compound. Blood compatibility, mutation assay, chromosomal aberration assay, and several other assays were also completed. Toxicity assessments were based on mortality, clinical signs, body weight, food consumption and anatomical pathology. Blood samples were collected to assess pharmacokinetic parameters including half-life, area under the curve, and clearance. Urine samples were collected to assess distribution. Doses of up to 200-300 times the estimated human dose were administered. No test-article related effects were noted on body weight, food consumption, ophthalmic observations and no abnormal pathology was seen in either macroscopic or microscopic evaluations of any organs or tissues. All animals survived to scheduled sacrifice. Transient discoloration of skin and urine was noted at the higher dose levels in both species as expected from a highly fluorescent compound and was not considered pathological. Thus initial toxicology studies of this new fluorescent tracer agent APC-2 have resulted in no demonstrable pathological test article concerns.

  6. Global and gene-specific DNA methylation pattern discriminates cholecystitis from gallbladder cancer patients in Chile

    PubMed Central

    Kagohara, Luciane Tsukamoto; Schussel, Juliana L; Subbannayya, Tejaswini; Sahasrabuddhe, Nandini; Lebron, Cynthia; Brait, Mariana; Maldonado, Leonel; Valle, Blanca L; Pirini, Francesca; Jahuira, Martha; Lopez, Jaime; Letelier, Pablo; Brebi-Mieville, Priscilla; Ili, Carmen; Pandey, Akhilesh; Chatterjee, Aditi; Sidransky, David; Guerrero-Preston, Rafael

    2015-01-01

    Aim The aim of the study was to evaluate the use of global and gene-specific DNA methylation changes as potential biomarkers for gallbladder cancer (GBC) in a cohort from Chile. Material & methods DNA methylation was analyzed through an ELISA-based technique and quantitative methylation-specific PCR. Results Global DNA Methylation Index (p = 0.02) and promoter methylation of SSBP2 (p = 0.01) and ESR1 (p = 0.05) were significantly different in GBC when compared with cholecystitis. Receiver curve operator analysis revealed promoter methylation of APC, CDKN2A, ESR1, PGP9.5 and SSBP2, together with the Global DNA Methylation Index, had 71% sensitivity, 95% specificity, a 0.97 area under the curve and a positive predictive value of 90%. Conclusion Global and gene-specific DNA methylation may be useful biomarkers for GBC clinical assessment. PMID:25066711

  7. Mayo de Los Capomos, Sinaloa (Mayo of Los Capomos, Sinaloa).

    ERIC Educational Resources Information Center

    Freeze, Ray A.

    This document is one of 17 volumes on indigenous Mexican languages and is the result of a project undertaken by the Archivo de Lenguas Indigenas de Mexico. This volume contains information on Mayo, an indigenous language of Mexico spoken in Los Capomos, in the state of Sinaloa. The objective of collecting such a representative sampling of the…

  8. Los NIH anuncian el lanzamiento de los estudios ALCHEMIST

    Cancer.gov

    Los Estudios sobre la Secuenciación e Identificación de Marcadores para el Mejoramiento de la Terapia Adjuvante para el Cáncer de Pulmón, ALCHEMIST, identificarán a pacientes con cáncer de pulmón en estadio inicial cuyos tumores tienen cambios genéticos.

  9. High Prevalence and Clinical Relevance of Genes Affected by Chromosomal Breaks in Colorectal Cancer

    PubMed Central

    van den Broek, Evert; Dijkstra, Maurits J. J.; Krijgsman, Oscar; Sie, Daoud; Haan, Josien C.; Traets, Joleen J. H.; van de Wiel, Mark A.; Nagtegaal, Iris D.; Punt, Cornelis J. A.; Carvalho, Beatriz; Ylstra, Bauke; Abeln, Sanne; Meijer, Gerrit A.; Fijneman, Remond J. A.

    2015-01-01

    Background Cancer is caused by somatic DNA alterations such as gene point mutations, DNA copy number aberrations (CNA) and structural variants (SVs). Genome-wide analyses of SVs in large sample series with well-documented clinical information are still scarce. Consequently, the impact of SVs on carcinogenesis and patient outcome remains poorly understood. This study aimed to perform a systematic analysis of genes that are affected by CNA-associated chromosomal breaks in colorectal cancer (CRC) and to determine the clinical relevance of recurrent breakpoint genes. Methods Primary CRC samples of patients with metastatic disease from CAIRO and CAIRO2 clinical trials were previously characterized by array-comparative genomic hybridization. These data were now used to determine the prevalence of CNA-associated chromosomal breaks within genes across 352 CRC samples. In addition, mutation status of the commonly affected APC, TP53, KRAS, PIK3CA, FBXW7, SMAD4, BRAF and NRAS genes was determined for 204 CRC samples by targeted massive parallel sequencing. Clinical relevance was assessed upon stratification of patients based on gene mutations and gene breakpoints that were observed in >3% of CRC cases. Results In total, 748 genes were identified that were recurrently affected by chromosomal breaks (FDR <0.1). MACROD2 was affected in 41% of CRC samples and another 169 genes showed breakpoints in >3% of cases, indicating that prevalence of gene breakpoints is comparable to the prevalence of well-known gene point mutations. Patient stratification based on gene breakpoints and point mutations revealed one CRC subtype with very poor prognosis. Conclusions We conclude that CNA-associated chromosomal breaks within genes represent a highly prevalent and clinically relevant subset of SVs in CRC. PMID:26375816

  10. Palace Revolt in Los Angeles?

    ERIC Educational Resources Information Center

    Fuller, Bruce

    2010-01-01

    Antonio Villaraigosa, the mayor of Los Angeles, comes alive when recalling his start in local politics--as a labor organizer agitating for reform inside decrepit and overcrowded schools. In his quest to turn around the schools, the mayor has united working-class Latino parents, civil rights leaders, and big-money Democrats to challenge union…

  11. RFQ development at Los Alamos

    SciTech Connect

    Wangler, T.P.; Crandall, K.R.; Stokes, R.H.

    1982-01-01

    The basic principles of the radio-frequency quadrupole (RFQ) linac are reviewed and a summary of past and present Los Alamos work is presented. Some beam-dynamics effects, important for RFQ design, are discussed. A design example is shown for xenon and a brief discussion of low-frequency RFQ structures is given.

  12. Aberrant gene methylation in non-neoplastic mucosa as a predictive marker of ulcerative colitis-associated CRC

    PubMed Central

    Castagliuolo, Ignazio; Erroi, Francesca; Kotsafti, Andromachi; Basato, Silvia; Brun, Paola; D'Incà, Renata; Rugge, Massimo

    2016-01-01

    Background Promoter hypermethylation plays a major role in cancer through transcriptional silencing of critical genes. The aim of our study is to evaluate the methylation status of these genes in the colonic mucosa without dysplasia or adenocarcinoma at the different steps of sporadic and UC-related carcinogenesis and to investigate the possible role of genomic methylation as a marker of CRC. Results The expression of Dnmts 1 and 3A was significantly increased in UC-related carcinogenesis compared to non inflammatory colorectal carcinogenesis. In non-neoplastic colonic mucosa, the number of methylated genes resulted significantly higher in patients with CRC and in those with UC-related CRC compared to the HC and UC patients and patients with dysplastic lesion of the colon. The number of methylated genes in non-neoplastic colonic mucosa predicted the presence of CRC with good accuracy either in non inflammatory and inflammatory related CRC. Methods Colonic mucosal samples were collected from healthy subjects (HC) (n = 30) and from patients with ulcerative colitis (UC) (n = 29), UC and dysplasia (n = 14), UC and cancer (n = 10), dysplastic adenoma (n = 14), and colon adenocarcinoma (n = 10). DNA methyltransferases-1, -3a, -3b, mRNA expression were quantified by real time qRT-PCR. The methylation status of CDH13, APC, MLH1, MGMT1 and RUNX3 gene promoters was assessed by methylation-specific PCR. Conclusions Methylation status of APC, CDH13, MGMT, MLH1 and RUNX3 in the non-neoplastic mucosa may be used as a marker of CRC: these preliminary results could allow for the adjustment of a patient's surveillance interval and to select UC patients who should undergo intensive surveillance. PMID:26862732

  13. Insertional mutagenesis identifies multiple networks of cooperating genes driving intestinal tumorigenesis.

    PubMed

    March, H Nikki; Rust, Alistair G; Wright, Nicholas A; ten Hoeve, Jelle; de Ridder, Jeroen; Eldridge, Matthew; van der Weyden, Louise; Berns, Anton; Gadiot, Jules; Uren, Anthony; Kemp, Richard; Arends, Mark J; Wessels, Lodewyk F A; Winton, Douglas J; Adams, David J

    2011-12-01

    The evolution of colorectal cancer suggests the involvement of many genes. To identify new drivers of intestinal cancer, we performed insertional mutagenesis using the Sleeping Beauty transposon system in mice carrying germline or somatic Apc mutations. By analyzing common insertion sites (CISs) isolated from 446 tumors, we identified many hundreds of candidate cancer drivers. Comparison to human data sets suggested that 234 CIS-targeted genes are also dysregulated in human colorectal cancers. In addition, we found 183 CIS-containing genes that are candidate Wnt targets and showed that 20 CISs-containing genes are newly discovered modifiers of canonical Wnt signaling. We also identified mutations associated with a subset of tumors containing an expanded number of Paneth cells, a hallmark of deregulated Wnt signaling, and genes associated with more severe dysplasia included those encoding members of the FGF signaling cascade. Some 70 genes had co-occurrence of CIS pairs, clustering into 38 sub-networks that may regulate tumor development. PMID:22057237

  14. Molecular basis of variegate porphyria: a missense mutation in the protoporphyrinogen oxidase gene.

    PubMed Central

    Frank, J; Lam, H; Zaider, E; Poh-Fitzpatrick, M; Christiano, A M

    1998-01-01

    Variegate porphyria (VP) is an autosomal dominant disorder characterised by a partial defect in the activity of protoporphyrinogen oxidase (PPO), and has recently been genetically linked to the PPO gene on chromosome 1q22-23 (Z=6.62). In this study, we identified a mutation in the PPO gene in a patient with VP and two unaffected family members. The mutation consisted of a previously unreported T to C transition in exon 13 of the PPO gene, resulting in the substitution of a polar serine by a non-polar proline (S450P). This serine residue is evolutionarily highly conserved in man, mouse, and Bacillus subtilis, attesting to the importance of this residue. Interestingly, the gene for Gardner's syndrome (FAP) also segregates in this family, independently of the VP mutation. Gardner's syndrome or familial adenomatous polyposis (FAP) is also an autosomal dominantly inherited genodermatosis, and typically presents with colorectal cancer in early adult life secondary to extensive adenomatous polyps of the colon. The specific gene on chromosome 5 that is the site of the mutation in this disorder is known as APC (adenomatous polyposis coli), and the gene has been genetically linked to the region of 5q22. Images PMID:9541112

  15. Los Angeles and Its Mistress Machine

    ERIC Educational Resources Information Center

    Marx, Wesley

    1973-01-01

    Los Angeles city has acute air pollution problems because of lack of an adequate mass transit system and the type of local industries. Air pollution in Los Angeles has affected agricultural production, vegetation, and public health in nearby areas. (PS)

  16. RFQ development at Los Alamos

    SciTech Connect

    Armstrong, D.D.; Cornelius, W.D.; Purser, F.O.; Jameson, R.A.; Wangler, T.P.

    1984-01-01

    We report recent progress on the two radio-frequency quadrupole (RFQ) structures being developed at Los Alamos. First, we report on the second 425-MHz RFQ for H/sup -/ acceleration, which is being built in a research effort to understand and further develop the RFQ. Second, we discuss progress on the 80-MHz cw RFQ for deuterons, which is being built for the Fusion Materials Irradiation Test (FMIT) facility.

  17. Targeted ultra-deep sequencing reveals recurrent and mutually exclusive mutations of cancer genes in blastic plasmacytoid dendritic cell neoplasm

    PubMed Central

    Pfarr, Nicole; Andrulis, Mindaugas; Jöhrens, Korinna; Klauschen, Frederick; Siebolts, Udo; Wolf, Thomas; Koch, Philipp-Sebastian; Schulz, Miriam; Hartschuh, Wolfgang; Goerdt, Sergij; Lennerz, Jochen K.; Wickenhauser, Claudia; Klapper, Wolfram

    2014-01-01

    Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare haematopoietic malignancy characterized by dismal prognosis and overall poor therapeutic response. Since the biology of BPDCN is barely understood, our study aims to shed light on the genetic make-up of these highly malignant tumors. Using targeted high-coverage massive parallel sequencing, we investigated 50 common cancer genes in 33 BPDCN samples. We detected point mutations in NRAS (27.3% of cases), ATM (21.2%), MET, KRAS, IDH2, KIT (9.1% each), APC and RB1 (6.1% each), as well as in VHL, BRAF, MLH1, TP53 and RET (3% each). Moreover, NRAS, KRAS and ATM mutations were found to be mutually exclusive and we observed recurrent mutations in NRAS, IDH2, APC and ATM. CDKN2A deletions were detected in 27.3% of the cases followed by deletions of RB1 (9.1%), PTEN and TP53 (3% each). The mutual exclusive distribution of some mutations may point to different subgroups of BPDCN whose biological significance remains to be explored. PMID:25115387

  18. Targeted ultra-deep sequencing reveals recurrent and mutually exclusive mutations of cancer genes in blastic plasmacytoid dendritic cell neoplasm.

    PubMed

    Stenzinger, Albrecht; Endris, Volker; Pfarr, Nicole; Andrulis, Mindaugas; Jöhrens, Korinna; Klauschen, Frederick; Siebolts, Udo; Wolf, Thomas; Koch, Philipp-Sebastian; Schulz, Miriam; Hartschuh, Wolfgang; Goerdt, Sergij; Lennerz, Jochen K; Wickenhauser, Claudia; Klapper, Wolfram; Anagnostopoulos, Ioannis; Weichert, Wilko

    2014-08-15

    Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare haematopoietic malignancy characterized by dismal prognosis and overall poor therapeutic response. Since the biology of BPDCN is barely understood, our study aims to shed light on the genetic make-up of these highly malignant tumors. Using targeted high-coverage massive parallel sequencing, we investigated 50 common cancer genes in 33 BPDCN samples. We detected point mutations in NRAS (27.3% of cases), ATM (21.2%), MET, KRAS, IDH2, KIT (9.1% each), APC and RB1 (6.1%), as well as in VHL, BRAF, MLH1, TP53 and RET1 (3% each). Moreover, NRAS-, KRAS- and ATM-mutations were found to be mutually exclusive and we observed recurrent mutations in NRAS, IDH2, APC and ATM. CDKN2A deletions were detected in 27.3% of the cases followed by deletions of RB1 (9.1%), PTEN and TP53 (3% each). The mutual exclusive distribution of some mutations may point to different subgroups of BPDCN whose biological significance remains to be explored. PMID:25115387

  19. Gene Therapy

    PubMed Central

    Baum, Bruce J

    2014-01-01

    Applications of gene therapy have been evaluated in virtually every oral tissue, and many of these have proved successful at least in animal models. While gene therapy will not be used routinely in the next decade, practitioners of oral medicine should be aware of the potential of this novel type of treatment that doubtless will benefit many patients with oral diseases. PMID:24372817

  20. Trichoderma genes

    DOEpatents

    Foreman, Pamela; Goedegebuur, Frits; Van Solingen, Pieter; Ward, Michael

    2012-06-19

    Described herein are novel gene sequences isolated from Trichoderma reesei. Two genes encoding proteins comprising a cellulose binding domain, one encoding an arabionfuranosidase and one encoding an acetylxylanesterase are described. The sequences, CIP1 and CIP2, contain a cellulose binding domain. These proteins are especially useful in the textile and detergent industry and in pulp and paper industry.

  1. A new approach to gene therapy using Sleeping Beauty to genetically modify clinical-grade T cells to target CD19

    PubMed Central

    Singh, Harjeet; Huls, Helen; Cooper, Laurence JN

    2014-01-01

    Summary The advent of efficient approaches to the genetic modification of T cells has provided investigators with clinically appealing approaches to improve the potency of tumor-specific clinical grade T cells. For example, gene therapy has been successfully used to enforce expression of chimeric antigen receptors (CAR) that provide T cells with ability to directly recognize tumor-associated antigens without the need for presentation by human leukocyte antigen. Gene transfer of CARs can be undertaken using viral-based and non-viral approaches. We have advanced DNA vectors derived from the Sleeping Beauty (SB) system to avoid the expense and manufacturing difficulty associated with transducing T cells with recombinant viral vectors. After electroporation, the transposon/transposase system improves the efficiency of integration of plasmids used to express CAR and other transgenes in T cells. The SB system combined with artificial antigen-presenting cells (aAPC) can selectively propagate and thus retrieve CAR+ T cells suitable for human application. This review describes the translation of the SB system and aAPC for use in clinical trials and highlights how a nimble and cost-effective approach to developing genetically modified T cells can be used to implement clinical trials infusing next-generation T cells with improved therapeutic potential. PMID:24329797

  2. Selectively Reduced Intracellular Proliferation of Salmonella enterica Serovar Typhimurium within APCs Limits Antigen Presentation and Development of a Rapid CD8 T Cell Response1

    PubMed Central

    Albaghdadi, Homam; Robinson, Nirmal; Finlay, Brett; Krishnan, Lakshmi; Sad, Subash

    2014-01-01

    Ag presentation to CD8+ T cells commences immediately after infection, which facilitates their rapid expansion and control of pathogen. This paradigm is not followed during infection with virulent Salmonella enterica serovar Typhimurium (ST), an intracellular bacterium that causes mortality in susceptible C57BL/6J mice within 7 days and a chronic infection in resistant mice (129 × 1SvJ). Infection of mice with OVA-expressing ST results in the development of a CD8+ T cell response that is detectable only after the second week of infection despite the early detectable bacterial burden. The mechanism behind the delayed CD8+ T cell activation was evaluated, and it was found that dendritic cells/macrophages or mice infected with ST-OVA failed to present Ag to OVA-specific CD8+ T cells. Lack of early Ag presentation was not rescued when mice or dendritic cells/macrophages were infected with an attenuated aroA mutant of ST or with mutants having defective Salmonella pathogenicity island I/II genes. Although extracellular ST proliferated extensively, the replication of ST was highly muted once inside macrophages. This muted intracellular proliferation of ST resulted in the generation of poor levels of intracellular Ag and peptide-MHC complex on the surface of dendritic cells. Additional experiments revealed that ST did not actively inhibit Ag presentation, rather it inhibited the uptake of another intracellular pathogen, Listeria monocytogenes, thereby causing inhibition of Ag presentation against L. monocytogenes. Taken together, this study reveals a dichotomy in the proliferation of ST and indicates that selectively reduced intra-cellular proliferation of virulent pathogens may be an important mechanism of immune evasion. PMID:19692639

  3. Expression analysis of eight amphioxus genes involved in the Wnt/β-catenin signaling pathway

    PubMed Central

    WANG, Jing; LI, Guang; QIAN, Guang-Hui; HUA, Jun-Hao; WANG, Yi-Quan

    2016-01-01

    The Wnt/β-catenin signaling pathway plays a crucial role in the embryonic development of metazoans. Although the pathway has been studied extensively in many model animals, its function in amphioxus, the most primitive chordate, remains largely uncharacterized. To obtain basic data for functional analysis, we identified and isolated seven genes (Lrp5/6, Dvl, APC, CkIα, CkIδ, Gsk3β, and Gro) of the Wnt/β-catenin signaling pathway from the amphioxus (Branchiostoma floridae) genome. Phylogenetic analysis revealed that amphioxus had fewer members of each gene family than that found in vertebrates. Whole-mount in situ hybridization showed that the genes were maternally expressed and broadly distributed throughout the whole embryo at the cleavage and blastula stages. Among them, Dvl was expressed asymmetrically towards the animal pole, while the others were evenly distributed in all blastomeres. At the mid-gastrula stage, the genes were specifically expressed in the primitive endomesoderm, but displayed different patterns. When the embryo developed into the neurula stage, the gene expressions were mainly detected in either paraxial somites or the tail bud. With the development of the embryo, the expression levels further decreased gradually and remained only in some pharyngeal regions or the tail bud at the larva stage. Our results suggest that the Wnt/β-catenin pathway might be involved in amphioxus somite formation and posterior growth, but not in endomesoderm specification. PMID:27265651

  4. Expression analysis of eight amphioxus genes involved in the Wnt/β-catenin signaling pathway.

    PubMed

    Wang, Jing; Li, Guang; Qian, Guang-Hui; Hua, Jun-Hao; Wang, Yi-Quan

    2016-05-18

    The Wnt/β-catenin signaling pathway plays a crucial role in the embryonic development of metazoans. Although the pathway has been studied extensively in many model animals, its function in amphioxus, the most primitive chordate, remains largely uncharacterized. To obtain basic data for functional analysis, we identified and isolated seven genes (Lrp5/6, Dvl, APC, CkIα, CkIδ, Gsk3β, and Gro) of the Wnt/β-catenin signaling pathway from the amphioxus (Branchiostoma floridae) genome. Phylogenetic analysis revealed that amphioxus had fewer members of each gene family than that found in vertebrates. Whole-mount in situ hybridization showed that the genes were maternally expressed and broadly distributed throughout the whole embryo at the cleavage and blastula stages. Among them, Dvl was expressed asymmetrically towards the animal pole, while the others were evenly distributed in all blastomeres. At the mid-gastrula stage, the genes were specifically expressed in the primitive endomesoderm, but displayed different patterns. When the embryo developed into the neurula stage, the gene expressions were mainly detected in either paraxial somites or the tail bud. With the development of the embryo, the expression levels further decreased gradually and remained only in some pharyngeal regions or the tail bud at the larva stage. Our results suggest that the Wnt/β-catenin pathway might be involved in amphioxus somite formation and posterior growth, but not in endomesoderm specification. PMID:27265651

  5. Nested Patch PCR enables highly multiplexed mutation discovery in candidate genes

    PubMed Central

    Varley, Katherine Elena; Mitra, Robi David

    2008-01-01

    Medical resequencing of candidate genes in individual patient samples is becoming increasingly important in the clinic and in clinical research. Medical resequencing requires the amplification and sequencing of many candidate genes in many patient samples. Here we introduce Nested Patch PCR, a novel method for highly multiplexed PCR that is very specific, can sensitively detect SNPs and mutations, and is easy to implement. This is the first method that couples multiplex PCR with sample-specific DNA barcodes and next-generation sequencing to enable highly multiplex mutation discovery in candidate genes for multiple samples in parallel. In our pilot study, we amplified exons from colon cancer and matched normal human genomic DNA. From each sample, we successfully amplified 96% (90 of 94) targeted exons from across the genome, totaling 21.6 kbp of sequence. Ninety percent of all sequencing reads were from targeted exons, demonstrating that Nested Patch PCR is highly specific. We found that the abundance of reads per exon was reproducible across samples. We reliably detected germline SNPs and discovered a colon tumor specific nonsense mutation in APC, a gene causally implicated in colorectal cancer. With Nested Patch PCR, candidate gene mutation discovery across multiple individual patient samples can now utilize the power of second-generation sequencing. PMID:18849522

  6. John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL WEST ELEVATION FROM PARKING LOT ACROSS SPRING STREET, FACING EAST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  7. John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL TENTH FLOOR SOUTH WING CAFETERIA FOOD LINE, FACING NORTH - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  8. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FOURTEENTH FLOOR Y-CORRIDOR, FACING NORTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  9. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FOURTEENTH FLOOR Y-CORRIDOR, FACING SOUTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  10. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FOURTEENTH FLOOR Y-CORRIDOR NEAR ROOM 1403, FACING SOUTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  11. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR Y-CORRIDOR SOUTH SIDE OF ELEVATOR LOBBY, FACING WEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  12. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR Y-CORRIDOR SOUTH SIDE OF ELEVATOR LOBBY, FACING NORTHEAST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  13. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR Y-CORRIDOR NORTH SIDE OF ELEVATOR LOBBY, FACING EAST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  14. John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL NINTH FLOOR NORTH OFFICE WING SHOWING PARTITIONS, WINDOWS AND RADIATOR, FACING EAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  15. John Ash, ALA., Photographer August 1997. VIEW OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, ALA., Photographer August 1997. VIEW OF LOS ANGELES CITY HALL NINTH FLOOR NORTH OFFICE WING SHOWING PARTITIONS, WINDOWS AND RADIATOR, FACING SOUTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  16. John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL TENTH FLOOR NORTH OFFICE WING SHOWING RADIATOR AND WINDOW, FACING EAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  17. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL WEST ENTRANCE COURTYARD SHOWING FLOORING, COLUMNS AND BRONZE DOORS, FACING EAST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  18. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL WEST ENTRANCE COURTYARD SHOWING BRONZE DOORS AND HANDRAILS, FACING EAST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  19. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL WEST ENTRANCE COURTYARD SHOWING BRONZE DOORS, LIGHT FIXTURES AND GRILLS, FACING EAST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  20. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL WEST ENTRANCE COURTYARD SOUTH ARCADE SHOWING FLOORING AND WALL MOSAICS, FACING WEST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  1. Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL ELEVENTH FLOOR ELEVATOR LOBBY, FACING NORTH - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  2. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL WEST ENTRANCE COURTYARD NORTHWEST CORNER SHOWING DAMAGED STONE, FACING NORTHWEST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  3. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL FOURTEENTH FLOOR SOUTH OFFICE WING SHOWING TYPICAL DOOR HARDWARE, FACING SOUTH - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  4. John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL NINTH FLOOR NORTH CORRIDOR STAIR NUMBER NINE DOOR HARDWARE, FACING WEST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  5. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL THIRD FLOOR SHOWING ROTUNDA COLUMN CAPITALS AND LIGHT FIXTURES, FACING SOUTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  6. Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL SIXTH FLOOR ZONING ADMINISTRATORS OFFICE SHOWING WOOD AND GLASS PARTITIONS, FACING SOUTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  7. John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL THIRD FLOOR CITY COUNCIL CHAMBERS SHOWING COLUMN, ARCADE AND SPECTATOR SEATING, FACING SOUTHWEST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  8. John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL THIRD FLOOR CITY COUNCIL CHAMBERS SHOWING PODIUM AND SEATING, FACING SOUTH. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  9. John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL THIRD FLOOR CITY COUNCIL CHAMBERS SHOWING COLUMN, ARCADE AND SPECTATOR SEATING, FACING SOUTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  10. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL NINETEENTH FLOOR MAIN OFFICE AREA, FACING NORTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  11. John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR NORTH OFFICE AREA, FACING NORTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  12. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL NINETEENTH FLOOR MAIN OFFICE AREA SHOWING DEMOLITION OF WEST WALL, FACING WEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  13. John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL SEVENTH FLOOR SOUTH OFFICE AREA SHOWING STRUCTURAL PIERS AND FLORESCENT LIGHTS, FACING NORTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  14. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FOURTEENTH FLOOR, SERVICE AREA DOOR NEAR ELEVATOR LOBBY, FACING SOUTH - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  15. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR NORTH OFFICE AREA, FACING NORTH - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  16. Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL SIXTH FLOOR NORTH OFFICE AREA, FACING NORTH - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  17. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL NINETEENTH FLOOR MAIN OFFICE AREA SHOWING BEAM AND COLUMN CONNECTION NEAR SOUTHEAST CORNER, FACING SOUTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  18. John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL SEVENTH FLOOR SOUTH OFFICE AREA SHOWING RADIATOR AND WINDOWS, FACING EAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  19. John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL SEVENTH FLOOR SOUTH OFFICE AREA SHOWING WOOD AND GLASS PARTITIONS, FACING SOUTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  20. Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Bruce D. Judd, FAIA, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL ELEVENTH FLOOR KITCHEN OF EXECUTIVE DINING AREA SHOWING ARCHED STRUCTURE, FACING NORTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  1. Bruce D. Judd, FAIA, Photographer August 1997. DETAIL OF LOS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Bruce D. Judd, FAIA, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL SIXTH FLOOR NORTH OFFICE AREA WINDOW, FACING NORTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  2. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL NINETEENTH FLOOR MAIN OFFICE AREA SHOWING DEMOLITION OF SOUTH WALL, FACING SOUTH - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  3. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL NINETEENTH FLOOR MAIN OFFICE AREA SHOWING BEAM AND COLUMN CONNECTION NEAR NORTHWEST CORNER, FACING NORTHWEST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  4. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL NINETEENTH FLOOR MAIN OFFICE AREA, FACING NORTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  5. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR WEST OFFICE AREA THAT WAS ORIGINAL ART COMMISSION ROOMS, FACING NORTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  6. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR BREAK ROOM OFF OF ORIGINAL ART COMMISSION ROOMS, FACING SOUTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  7. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR BREAK ROOM OFF OF ORIGINAL ART COMMISSION ROOMS, FACING NORTHEAST - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  8. Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. VIEW OF LOS ANGELES CITY HALL FIFTH FLOOR WEST OFFICE AREA THAT WAS ORIGINAL ART COMMISSION ROOMS, FACING NORTHWEST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  9. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL WEST ENTRANCE COURTYARD NORTH ARCADE SHOWING TILE WORK, FACING NORTH. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  10. Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Monica Griesbach, Photographer August 1997. DETAIL OF LOS ANGELES CITY HALL SECOND FLOOR SHOWING SOUTH LOBBY TILE WORK, FACING NORTHWEST. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  11. Los Alamos Science: Number 16

    SciTech Connect

    Cooper, N.G.

    1988-01-01

    It was an unusually stimulating day and a half at Los Alamos when two Nobel Laureates in physiology, a leading paleontologist, and a leading bio-astrophysicist came together to discuss ''Unsolved Problems in the Science of Life,'' the topic of the second in a series of special meetings sponsored by the Fellows of the Laboratory. Just like the first one on ''Creativity in Science,'' this colloquium took us into a broader arena of ideas and viewpoints than is our usual daily fare. To contemplate the evolution and mysteries of intelligent life from the speakers' diverse points of view at one time, in one place was indeed a rare experience.

  12. [Language gene].

    PubMed

    Takahashi, Hiroshi

    2006-11-01

    The human capacity for acquiring speech and language must derive, at least in part, from the genome. Recent advance in the field of molecular genetics finally discovered 'Language Gene'. Disruption of FOXP2 gene, the firstly identified 'language gene' causes severe speech and language disorder. To elucidate the anatomical basis of language processing in the brain, we examined the expression pattern of FOXP2/Foxp2 genes in the monkey and rat brains through development. We found the preferential expression of FOXP2/Foxp2 in the striosomal compartment of the developing striatum. Thus, we suggest the striatum, particularly striosomal system may participate in neural information processing for language and speech. Our suggestion is consistent with the declarative/ procedural model of language proposed by Ullman (1997, 2001), which the procedural memory-dependent mental grammar is rooted in the basal ganglia and the frontal cortex, and the declarative memory-dependent mental lexicon is rooted in the temporal lobe. PMID:17432197

  13. Genes V.

    SciTech Connect

    Lewin, B.

    1994-12-31

    This fifth edition book encompasses a wide range of topics covering 1,272 pages. The book is arranged into nine parts with a total of 36 chapters. These nine parts include Introduction; DNA as a Store of Information; Translation; Constructing Cells; Control of Prokaryotypic Gene Expression; Perpetuation of DNA; Organization of the Eukaryotypic Genome; Eukaryotypic Transcription and RNA Processing; The Dynamic Genome; and Genes in Development.

  14. Single Gene Prognostic Biomarkers in Ovarian Cancer: A Meta-Analysis

    PubMed Central

    Willis, Scooter; Villalobos, Victor M.; Gevaert, Olivier; Abramovitz, Mark; Williams, Casey; Sikic, Branimir I.; Leyland-Jones, Brian

    2016-01-01

    Purpose To discover novel prognostic biomarkers in ovarian serous carcinomas. Methods A meta-analysis of all single genes probes in the TCGA and HAS ovarian cohorts was performed to identify possible biomarkers using Cox regression as a continuous variable for overall survival. Genes were ranked by p-value using Stouffer’s method and selected for statistical significance with a false discovery rate (FDR) <.05 using the Benjamini-Hochberg method. Results Twelve genes with high mRNA expression were prognostic of poor outcome with an FDR <.05 (AXL, APC, RAB11FIP5, C19orf2, CYBRD1, PINK1, LRRN3, AQP1, DES, XRCC4, BCHE, and ASAP3). Twenty genes with low mRNA expression were prognostic of poor outcome with an FDR <.05 (LRIG1, SLC33A1, NUCB2, POLD3, ESR2, GOLPH3, XBP1, PAXIP1, CYB561, POLA2, CDH1, GMNN, SLC37A4, FAM174B, AGR2, SDR39U1, MAGT1, GJB1, SDF2L1, and C9orf82). Conclusion A meta-analysis of all single genes identified thirty-two candidate biomarkers for their possible role in ovarian serous carcinoma. These genes can provide insight into the drivers or regulators of ovarian cancer and should be evaluated in future studies. Genes with high expression indicating poor outcome are possible therapeutic targets with known antagonists or inhibitors. Additionally, the genes could be combined into a prognostic multi-gene signature and tested in future ovarian cohorts. PMID:26886260

  15. The biology of novel animal genes: Mouse APEX gene knockout

    SciTech Connect

    MacInnes, M.; Altherr, M.R.; Ludwig, D.; Pedersen, R.; Mold, C.

    1997-07-01

    This is the final report of a one-year, Laboratory Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). The controlled breeding of novel genes into mice, including the gene knockout (KO), or conversely by adding back transgenes provide powerful genetic technologies that together suffice to determine in large part the biological role(s) of novel genes. Inbred mouse remains the best understood and most useful mammalian experimental system available for tackling the biology of novel genes. The major mammalian apurinic/apyrimidinic (AP) endonuclease (APE), is involved in a key step in the repair of spontaneous and induced AP sites in DNA. Efficient repair of these lesions is imperative to prevent the stable incorporation of mutations into the cellular genome which may lead to cell death or transformation. Loss or modulation of base excison repair activity in vivo may elevate the spontaneous mutation rate in cells, and may lead to a substantial increase in the incidence of cancer. Despite extensive biochemical analysis, however, the significance of these individual APE functions in vivo has not been elucidated. Mouse embryonic stem (ES) cells heterozygous for a deletion mutation in APE have been generated and whole animals containing the APE mutation have been derived from these ES cells. Animals homozygous for the APE null mutation die early in gestation, underscoring the biological significance of this DNA repair gene.

  16. Identification of tandem genes involved in lipooligosaccharide expression by Haemophilus ducreyi.

    PubMed Central

    Stevens, M K; Klesney-Tait, J; Lumbley, S; Walters, K A; Joffe, A M; Radolf, J D; Hansen, E J

    1997-01-01

    A transposon insertion mutant of Haemophilus ducreyi 35000 possessing a truncated lipooligosaccharide (LOS) failed to bind the LOS-specific monoclonal antibody 3E6 (M. K. Stevens, L. D. Cope, J. D. Radolf, and E. J. Hansen, Infect. Immun. 63:2976-2982, 1995). This transposon was found to have inserted into the first of two tandem genes and also caused a deletion of chromosomal DNA upstream of this gene. These two genes, designated lbgA and lbgB, encoded predicted proteins with molecular masses of 25,788 and 40,236 Da which showed homology with proteins which function in lipopolysaccharide biosynthetic in other gram-negative bacteria. The tandem arrangement of the lbgA and lbgB genes was found to be conserved among H. ducreyi strains. Isogenic LOS mutants, constructed by the insertion of a cat cartridge into either the lbgA or the lbgB gene, expressed an LOS phenotype indistinguishable from that of the original transposon-derived LOS mutant. The wild-type LOS phenotype could be restored by complementation with the appropriate wild-type allele. These two LOS mutants proved to be as virulent as the wild-type parent strain in an animal model. A double mutant with a deletion of the lbgA and lbgB genes yielded equivocal results when its virulence was tested in an animal model. PMID:9009327

  17. Los Alamos PC estimating system

    SciTech Connect

    Stutz, R.A.; Lemon, G.D.

    1987-01-01

    The Los Alamos Cost Estimating System (QUEST) is being converted to run on IBM personal computers. This very extensive estimating system is capable of supporting cost estimators from many different and varied fields. QUEST does not dictate any fixed method for estimating. QUEST supports many styles and levels of detail estimating. QUEST can be used with or without data bases. This system allows the estimator to provide reports based on levels of detail defined by combining work breakdown structures. QUEST provides a set of tools for doing any type of estimate without forcing the estimator to use any given method. The level of detail in the estimate can be mixed based on the amount of information known about different parts of the project. The system can support many different data bases simultaneously. Estimators can modify any cost in any data base.

  18. Analysis of Candidate Genes in Occurrence and Growth of Colorectal Adenomas

    PubMed Central

    Olschwang, Sylviane; Vernerey, Déwi; Cottet, Vanessa; Pariente, Alexandre; Nalet, Bernard; Lafon, Jacques; Faivre, Jean; Laurent-Puig, Pierre; Bonithon-Kopp, Claire; Bonaiti-Pellié, Catherine

    2009-01-01

    Predisposition to sporadic colorectal tumours is influenced by genes with minor phenotypic effects. A case-control study was set up on 295 patients treated for a large adenoma matched with polyp-free individuals on gender, age, and geographic origin in a 1 : 2 proportion. A second group of 302 patients treated for a small adenoma was also characterized to distinguish effects on adenoma occurrence and growth. We focussed the study on 38 single nucleotide polymorphisms (SNPs) encompassing 14 genes involved in colorectal carcinogenesis. Effect of SNPs was tested using unconditional logistic regression. Comparisons were made for haplotypes within a given gene and for biologically relevant genes combinations using the combination test. The APC p.Glu1317Gly variant appeared to influence the adenoma growth (P = .04, exact test) but not its occurrence. This result needs to be replicated and genome-wide association studies may be necessary to fully identify low-penetrance alleles involved in early stages of colorectal tumorigenesis. PMID:19888426

  19. A phase I study of hydralazine to demethylate and reactivate the expression of tumor suppressor genes

    PubMed Central

    Zambrano, Pilar; Segura-Pacheco, Blanca; Perez-Cardenas, Enrique; Cetina, Lucely; Revilla-Vazquez, Alma; Taja-Chayeb, Lucía; Chavez-Blanco, Alma; Angeles, Enrique; Cabrera, Gustavo; Sandoval, Karina; Trejo-Becerril, Catalina; Chanona-Vilchis, Jose; Duenas-González, Alfonso

    2005-01-01

    Background The antihypertensive compound hydralazine is a known demethylating agent. This phase I study evaluated the tolerability and its effects upon DNA methylation and gene reactivation in patients with untreated cervical cancer. Methods Hydralazine was administered to cohorts of 4 patients at the following dose levels: I) 50 mg/day, II) 75 mg/day, III) 100 mg/day and IV) 150 mg/day. Tumor biopsies and peripheral blood samples were taken the day before and after treatment. The genes APC, MGMT; ER, GSTP1, DAPK, RARβ, FHIT and p16 were evaluated pre and post-treatment for DNA promoter methylation and gene expression by MSP (Methylation-Specific PCR) and RT-PCR respectively in each of the tumor samples. Methylation of the imprinted H19 gene and the "normally methylated" sequence clone 1.2 was also analyzed. Global DNA methylation was analyzed by capillary electrophoresis and cytosine extension assay. Toxicity was evaluated using the NCI Common Toxicity Criteria. Results Hydralazine was well tolerated. Toxicities were mild being the most common nausea, dizziness, fatigue, headache and palpitations. Overall, 70% of the pretreatment samples and all the patients had at least one methylated gene. Rates of demethylation at the different dose levels were as follows: 50 mg/day, 40%; 75 mg/day, 52%, 100 mg/day, 43%, and 150 mg/day, 32%. Gene expression analysis showed only 12 informative cases, of these 9 (75%) re-expressed the gene. There was neither change in the methylation status of H19 and clone 1.2 nor changes in global DNA methylation. Conclusion Hydralazine at doses between 50 and 150 mg/day is well tolerated and effective to demethylate and reactivate the expression of tumor suppressor genes without affecting global DNA methylation PMID:15862127

  20. Investigadores simulan los beneficios reales de los exámenes de detección

    Cancer.gov

    Artículo sobre los esfuerzos de la Red de Modelado de Intervención y Vigilancia del Cáncer (CISNET) del NCI para determinar los beneficios reales de los exámenes de detección del cáncer en la población general.

  1. Nutritionally driven differential gene expression leads to heterochronic brain development in honeybee castes.

    PubMed

    Moda, Lívia Maria; Vieira, Joseana; Guimarães Freire, Anna Cláudia; Bonatti, Vanessa; Bomtorin, Ana Durvalina; Barchuk, Angel Roberto; Simões, Zilá Luz Paulino

    2013-01-01

    The differential feeding regimes experienced by the queen and worker larvae of the honeybee Apis mellifera shape a complex endocrine response cascade that ultimately gives rise to differences in brain morphologies. Brain development analyzed at the morphological level from the third (L3) through fifth (L5) larval instars revealed an asynchrony between queens and workers. In the feeding phase of the last larval instar (L5F), two well-formed structures, pedunculi and calyces, are identifiable in the mushroom bodies of queens, both of which are not present in workers until a later phase (spinning phase, L5S). Genome-wide expression analyses and normalized transcript expression experiments monitoring specific genes revealed that this differential brain development starts earlier, during L3. Analyzing brains from L3 through L5S1 larvae, we identified 21 genes with caste-specific transcription patterns (e.g., APC-4, GlcAT-P, fax, kr-h1 and shot), which encode proteins that are potentially involved in the development of brain tissues through controlling the cell proliferation rate (APC4, kr-h1) and fasciculation (GlcAT-P, fax, and shot). Shot, whose expression is known to be required for axon extension and cell proliferation, was found to be transcribed at significantly higher levels in L4 queens compared with worker larvae. Moreover, the protein encoded by this gene was immunolocalized to the cytoplasm of cells near the antennal lobe neuropiles and proximal to the Kenyon cells in the brains of L4 queens. In conclusion, during the larval period, the brains of queens are larger and develop more rapidly than workers' brains, which represents a developmental heterochrony reflecting the effect of the differential feeding regime of the two castes on nervous system development. Furthermore, this differential development is characterized by caste-specific transcriptional profiles of a set of genes, thus pointing to a link between differential nutrition and differential

  2. Trouble Brewing in Los Angeles. Policy Brief

    ERIC Educational Resources Information Center

    Buck, Stuart

    2010-01-01

    The city of Los Angeles will face enormous budgetary pressures from the growing deficits in public pensions, both at a state and local level. In this policy brief, the author estimates that Los Angeles faces a total $152.6 billion liability for pensions that are underfunded--including $49.1 billion for the city pension systems, $2.4 billion for…

  3. Vacunas contra los virus del papiloma humano

    Cancer.gov

    Una hoja informativa acerca de las vacunas contra los virus del papiloma humano (VPH) para prevenir infecciones con ciertos tipos de VPH, los cuales son la causa principal del cáncer de cuello del útero o cérvix.

  4. Significant Silence in Elena Garro's "Los Perros"

    ERIC Educational Resources Information Center

    Burke, Jessica

    2010-01-01

    Elena Garro's one-act play "Los perros" (1958) confronts the difficult issue of sexual violence in rural Mexico, a problem that persists today. The characters struggle with the social reality of rape, alluding to the threat of sexual violence while avoiding addressing it directly. While words are granted an almost magical power in "Los perros",…

  5. Attention Genes

    ERIC Educational Resources Information Center

    Posner, Michael I.; Rothbart, Mary K.; Sheese, Brad E.

    2007-01-01

    A major problem for developmental science is understanding how the cognitive and emotional networks important in carrying out mental processes can be related to individual differences. The last five years have seen major advances in establishing links between alleles of specific genes and the neural networks underlying aspects of attention. These…

  6. Designer Genes.

    ERIC Educational Resources Information Center

    Miller, Judith; Miller, Mark

    1983-01-01

    Genetic technologies may soon help fill some of the most important needs of humanity from food to energy to health care. The research of major designer genes companies and reasons why the initial mad rush for biotechnology has slowed are reviewed. (SR)

  7. Molecular and cellular effects of cis-9, trans-11-conjugated linoleic acid in enterocytes: effects on proliferation, differentiation, and gene expression.

    PubMed

    Lampen, A; Leifheit, M; Voss, J; Nau, H

    2005-06-15

    It has been hypothesized that dietary conjugated linoleic acids (CLA) may inhibit colon tumorigenesis. The aim of our study was to investigate the cellular and molecular effects of cis-9 (9Z), trans-11 (11E)-CLA on the proliferation, differentiation, interaction with peroxisome proliferator-activated receptors (PPARs), and expression of genes relevant in the APC-beta-catenin-TCF4 signalling pathway in human HT-29 and Caco-2 colon cells. We found that 9Z,11E-CLA inhibited the proliferation of HT-29 and Caco-2 cells. Trans-vaccenic acid (VA) showed no antiproliferative effects at all. We determined that 9Z,11E-CLA induced cell differentiation as measured by intestinal alkaline phosphatase (IAP) enzyme activity in Caco-2 cells, mRNA expression of IAP, and activation of a 5' flanking region of IAP. The 9Z,11E-CLA activated human PPARdelta as measured in a reporter gene assay. Treatment of HT29 cells in the poliferation phase with 9Z,11E-CLA repressed mRNA-expression of proliferation genes such as c-myc, cyclin D1 and c-jun in a concentration dependent manner. The promoter activities of c-myc and AP1 were also inhibited after incubation with 9Z,11E-CLA. beta-Catenin mRNA and protein expression was also repressed by the treatment with 9Z,11E-CLA. In addition, the mRNA expression of PPARdelta was repressed by treatment of the HT-29 cells with 9Z,11E-CLA. We conclude that 9Z,11E-CLA has an antiproliferative effect at the cellular and molecular levels in human colon cells. The results indicate that the preventive effects of CLA in the development of colon cancer may be due to their downregulation of some target genes of the APC-beta-catenin-TCF-4- and PPARdelta signalling pathway. PMID:15935729

  8. Clofarabine, a novel adenosine analogue, reactivates DNA methylation-silenced tumour suppressor genes and inhibits cell growth in breast cancer cells.

    PubMed

    Lubecka-Pietruszewska, Katarzyna; Kaufman-Szymczyk, Agnieszka; Stefanska, Barbara; Cebula-Obrzut, Barbara; Smolewski, Piotr; Fabianowska-Majewska, Krystyna

    2014-01-15

    Clofarabine (2-chloro-2'-fluoro-2'-deoxyarabinosyladenine, ClF) is a second-generation 2'-deoxyadenosine analogue that is structurally related to cladribine (2-chloro-2'-deoxyadenosine, 2CdA) and fludarabine (9-beta-d-arabinosyl-2-fluoroadenine, F-ara-A). It demonstrates potent antitumour activity at much lower doses than parent compounds with high therapeutic efficacy in paediatric blood cancers. Our previous studies in breast cancer cells indicate that 2CdA and F-ara-A are involved in epigenetic regulation of gene transcription. We therefore investigated whether ClF influences methylation and expression of selected tumour suppressor genes, such as adenomatous polyposis coli (APC), phosphatase and tensin homologue (PTEN), and retinoic acid receptor beta 2 (RARbeta2), as well as expression of p53, p21 and DNA methyltransferase 1 (DNMT1) in MCF-7 and MDA-MB-231 breast cancer cell lines with different invasive potential. Promoter methylation and gene expression were estimated using methylation-sensitive restriction analysis (MSRA) and real-time PCR, respectively. ClF demonstrated potent growth inhibitory activity in MCF-7 and MDA-MB-231 cells after 96h treatment with IC50 determined as equal to 640nM and 50nM, respectively. In both breast cancer cell lines, ClF led to hypomethylation and up-regulation of APC, PTEN and RARbeta2 as well as increase in p21 expression. Only in non-invasive MCF-7 cells, these changes were associated with down-regulation of DNMT1. Our results provide first evidence of ClF implications in epigenetic regulation of transcriptional activity of selected tumour suppressor genes in breast cancer. It seems to be a new important element of ClF anticancer activity and may indicate its potential efficacy in epigenetic therapy of solid tumours, especially at early stages of carcinogenesis. PMID:24296317

  9. Comprehensive BEOL control using scatterometry and APC

    NASA Astrophysics Data System (ADS)

    Timoney, Padraig; Tsai, Jamie; Baral, Sudhir; Economikos, Laertis; Vaid, Alok; Lu, Haibo; Kang, Byungcheol; Isbester, Paul; Dasari, Prasad; Kort, Roy; Yellai, Naren

    2015-03-01

    Copper interconnects have been adopted in advanced semiconductor manufacturing due to benefits of reduced RC delay, cross talk and power consumption. With each technology node, interconnects reduce in size resulting in increased line resistivity, a critical metric in determining the device performance. Reactive Ion Etching (RIE) and Copper Chemical Mechanical Polishing (Cu CMP) are two of the key back end of the line (BEOL) processes that affect the interconnect performance. Due to variations from incoming processes and the inherent variability induced by these processes, dielectric trench depth and resulting copper line height variations that can potentially result from these processes have direct impact to RC delay. Traditional inline metrology methods used are time consuming and do not provide the needed wafer level metrics. In addition, measurement of remaining dielectric thickness on solid pads is not a good representative of the actual device structures and has been inaccurate for process due to dishing of the copper pads. Efficient control of BEOL processes requires measurement of metal line thickness and other critical profile parameters from which resistance can be extracted. In order to relate BEOL process steps and understand their interactions, it is necessary to have a directly comparable measurement methodology on a similar measurement structure. Over the past several years, scatterometry has been proven as the only metrology method to provide the full profile information of the Cu lines. Scatterometry is a diffraction based optical measurement technique using Rigorous Coupled Wave Analysis (RCWA), where light diffracted from a periodic structure is used to characterize the details of profile. Unique algorithms, such as Holistic Metrology can be used to make the scatterometry development process faster. In this paper, we will present how scatterometry can be used to measure copper line height on 3D structures and how feed forward from RIE can be applied for control of Cu CMP process for 20nm technology node. The importance of incoming trench depth variations is demonstrated for CMP polish time control in order to stabilize the copper line height. Validation data is presented for different scatterometry models including accuracy, repeatability and DoE tracking. Electrical resistance is shown to correlate to the copper trench profile measured by scatterometry. The paper will demonstrate the capability for reducing copper line height variation and the correlation of the reducing trench height variation to improved stabilization of electrical resistance.

  10. Los Alamos Laser Eye Investigation.

    SciTech Connect

    Odom, C. R.

    2005-01-01

    A student working in a laser laboratory at Los Alamos National Laboratory sustained a serious retinal injury to her left eye when she attempted to view suspended particles in a partially evacuated target chamber. The principle investigator was using the white light from the flash lamp of a Class 4 Nd:YAG laser to illuminate the particles. Since the Q-switch was thought to be disabled at the time of the accident, the principal investigator assumed it would be safe to view the particles without wearing laser eye protection. The Laboratory Director appointed a team to investigate the accident and to report back to him the events and conditions leading up to the accident, equipment malfunctions, safety management causal factors, supervisory and management action/inaction, adequacy of institutional processes and procedures, emergency and notification response, effectiveness of corrective actions and lessons learned from previous similar events, and recommendations for human and institutional safety improvements. The team interviewed personnel, reviewed documents, and characterized systems and conditions in the laser laboratory during an intense six week investigation. The team determined that the direct and primary failures leading to this accident were, respectively, the principle investigator's unsafe work practices and the institution's inadequate monitoring of worker performance. This paper describes the details of the investigation, the human and institutional failures, and the recommendations for improving the laser safety program.

  11. DNA methylation and gene deletion analysis of brain metastases in melanoma patients identifies mutually exclusive molecular alterations

    PubMed Central

    Marzese, Diego M.; Scolyer, Richard A.; Roqué, Maria; Vargas-Roig, Laura M.; Huynh, Jamie L.; Wilmott, James S.; Murali, Rajmohan; Buckland, Michael E.; Barkhoudarian, Garni; Thompson, John F.; Morton, Donald L.; Kelly, Daniel F.; Hoon, Dave S. B.

    2014-01-01

    Background The brain is a common target of metastases for melanoma patients. Little is known about the genetic and epigenetic alterations in melanoma brain metastases (MBMs). Unraveling these molecular alterations is a key step in understanding their aggressive nature and identifying novel therapeutic targets. Methods Genome-wide DNA methylation analyses of MBMs (n = 15) and normal brain tissues (n = 91) and simultaneous multigene DNA methylation and gene deletion analyses of metastatic melanoma tissues (99 MBMs and 43 extracranial metastases) were performed. BRAF and NRAS mutations were evaluated in MBMs by targeted sequencing. Results MBMs showed significant epigenetic heterogeneity. RARB, RASSF1, ESR1, APC, PTEN, and CDH13 genes were frequently hypermethylated. Deletions were frequently detected in the CDKN2A/B locus. Of MBMs, 46.1% and 28.8% had BRAF and NRAS missense mutations, respectively. Compared with lung and liver metastases, MBMs exhibited higher frequency of CDH13 hypermethylation and CDKN2A/B locus deletion. Mutual exclusivity between hypermethylated genes and CDKN2A/B locus deletion identified 2 clinically relevant molecular subtypes of MBMs. CDKN2A/B deletions were associated with multiple MBMs and frequently hypermethylated genes with shorter time to brain metastasis. Conclusions Melanoma cells that colonize the brain harbor numerous genetically and epigenetically altered genes. This study presents an integrated genomic and epigenomic analysis that reveals MBM-specific molecular alterations and mutually exclusive molecular subtypes. PMID:24968695

  12. Umbilical cord gene expression reveals the molecular architecture of the fetal inflammatory response in extremely preterm newborns

    PubMed Central

    Costa, Daniel; Castelo, Robert

    2016-01-01

    Background: The fetal inflammatory response (FIR) in placental membranes to an intrauterine infection often precedes premature birth raising neonatal mortality and morbidity. However, the precise molecular events behind FIR still remain largely unknown, and little has been investigated at gene expression level. Methods: We collected publicly available microarray expression data profiling umbilical cord (UC) tissue derived from the cohort of extremely low gestational age newborns (ELGANs) and interrogate them for differentially expressed (DE) genes between FIR and non–FIR-affected ELGANs. Results: We found a broad and complex FIR UC gene expression signature, changing up to 19% (3,896/20,155) of all human genes at 1% false discovery rate. Significant changes of a minimum 50% magnitude (1,097/3,896) affect the upregulation of many inflammatory pathways and molecules, such as cytokines, toll-like receptors, and calgranulins. Remarkably, they also include the downregulation of neurodevelopmental pathways and genes, such as Fragile-X mental retardation 1 (FMR1), contactin 1 (CNTN1), and adenomatous polyposis coli (APC). Conclusion: The FIR expression signature in UC tissue contains molecular clues about signaling pathways that trigger FIR, and it is consistent with an acute inflammatory response by fetal innate and adaptive immune systems, which participate in the pathogenesis of neonatal brain damage. PMID:26539667

  13. 40. PLEASANT VALLEY RESERVOIR DAM LOOKING NORTHWEST Los Angeles ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    40. PLEASANT VALLEY RESERVOIR DAM LOOKING NORTHWEST - Los Angeles Aqueduct, From Lee Vining Intake (Mammoth Lakes) to Van Norman Reservoir Complex (San Fernando Valley), Los Angeles, Los Angeles County, CA

  14. Genes and Hearing Loss

    MedlinePlus

    ... Meeting Calendar Find an ENT Doctor Near You Genes and Hearing Loss Genes and Hearing Loss Patient ... mutation may only have dystopia canthorum. How Do Genes Work? Genes are a road map for the ...

  15. Environmental surveillance at Los Alamos during 1994

    SciTech Connect

    1996-07-01

    This report describes environmental monitoring activities at Los Alamos National Laboratory for 1994. Data were collected to assess external penetrating radiation, airborne emissions, liquid effluents, radioactivity of environmental materials and food stuffs, and environmental compliance.

  16. New Rad Lab for Los Alamos

    ScienceCinema

    None

    2010-01-08

    The topping out ceremony for a key construction stage in the Los Alamos National Laboratory's newest facility, the Radiological Laboratory Utility & Office Building. This is part of the National Nu...  

  17. Shuttle Endeavour Flyover of Los Angeles Landmarks

    NASA Video Gallery

    Space shuttle Endeavour atop NASA's Shuttle Carrier Aircraft flew over many Los Angeles area landmarks on its final ferry flight Sept. 21, 2012, including the Coliseum, the Hollywood Sign, Griffith...

  18. Publications of Los Alamos research 1988

    SciTech Connect

    Varjabedian, K.; Dussart, S.A.; McClary, W.J.; Rich, J.A.

    1989-12-01

    This bibliography lists unclassified publications of work done at the Los Alamos National Laboratory for 1988. The entries, which are subdivided by broad subject categories, are cross-referenced with an author index and a numeric index.

  19. New Rad Lab for Los Alamos

    SciTech Connect

    2008-08-06

    The topping out ceremony for a key construction stage in the Los Alamos National Laboratory's newest facility, the Radiological Laboratory Utility & Office Building. This is part of the National Nu...  

  20. Edward Teller Returns to LOS Alamos

    NASA Astrophysics Data System (ADS)

    Hecker, Siegfried S.

    2010-01-01

    I was asked to share some reflections of Edward Teller's return to Los Alamos during my directorship. I met Teller late in his life. My comments focus on that time and they will be mostly in the form of stories of my interactions and those of my colleagues with Teller. Although the focus of this symposium is on Teller's contributions to science, at Los Alamos it was never possible to separate Teller's science from policy and controversy ...

  1. Internship at Los Alamos National Laboratory

    SciTech Connect

    Dunham, Ryan Q.

    2012-07-11

    Los Alamos National Laboratory (LANL) is located in Los Alamos, New Mexico. It provides support for our country's nuclear weapon stockpile as well as many other scientific research projects. I am an Undergraduate Student Intern in the Systems Design and Analysis group within the Nuclear Nonproliferation division of the Global Security directorate at LANL. I have been tasked with data analysis and modeling of particles in a fluidized bed system for the capture of carbon dioxide from power plant flue gas.

  2. SNP discovery in candidate adaptive genes using exon capture in a free-ranging alpine ungulate.

    PubMed

    Roffler, Gretchen H; Amish, Stephen J; Smith, Seth; Cosart, Ted; Kardos, Marty; Schwartz, Michael K; Luikart, Gordon

    2016-09-01

    Identification of genes underlying genomic signatures of natural selection is key to understanding adaptation to local conditions. We used targeted resequencing to identify SNP markers in 5321 candidate adaptive genes associated with known immunological, metabolic and growth functions in ovids and other ungulates. We selectively targeted 8161 exons in protein-coding and nearby 5' and 3' untranslated regions of chosen candidate genes. Targeted sequences were taken from bighorn sheep (Ovis canadensis) exon capture data and directly from the domestic sheep genome (Ovis aries v. 3; oviAri3). The bighorn sheep sequences used in the Dall's sheep (Ovis dalli dalli) exon capture aligned to 2350 genes on the oviAri3 genome with an average of 2 exons each. We developed a microfluidic qPCR-based SNP chip to genotype 476 Dall's sheep from locations across their range and test for patterns of selection. Using multiple corroborating approaches (lositan and bayescan), we detected 28 SNP loci potentially under selection. We additionally identified candidate loci significantly associated with latitude, longitude, precipitation and temperature, suggesting local environmental adaptation. The three methods demonstrated consistent support for natural selection on nine genes with immune and disease-regulating functions (e.g. Ovar-DRA, APC, BATF2, MAGEB18), cell regulation signalling pathways (e.g. KRIT1, PI3K, ORRC3), and respiratory health (CYSLTR1). Characterizing adaptive allele distributions from novel genetic techniques will facilitate investigation of the influence of environmental variation on local adaptation of a northern alpine ungulate throughout its range. This research demonstrated the utility of exon capture for gene-targeted SNP discovery and subsequent SNP chip genotyping using low-quality samples in a nonmodel species. PMID:27327375

  3. Exome sequencing identifies potential novel candidate genes in patients with unexplained colorectal adenomatous polyposis.

    PubMed

    Spier, Isabel; Kerick, Martin; Drichel, Dmitriy; Horpaopan, Sukanya; Altmüller, Janine; Laner, Andreas; Holzapfel, Stefanie; Peters, Sophia; Adam, Ronja; Zhao, Bixiao; Becker, Tim; Lifton, Richard P; Holinski-Feder, Elke; Perner, Sven; Thiele, Holger; Nöthen, Markus M; Hoffmann, Per; Timmermann, Bernd; Schweiger, Michal R; Aretz, Stefan

    2016-04-01

    In up to 30% of patients with colorectal adenomatous polyposis, no germline mutation in the known genes APC, causing familial adenomatous polyposis, MUTYH, causing MUTYH-associated polyposis, and POLE or POLD1, causing Polymerase-Proofreading-associated polyposis can be identified, although a hereditary etiology is likely. To uncover new causative genes, exome sequencing was performed using DNA from leukocytes and a total of 12 colorectal adenomas from seven unrelated patients with unexplained sporadic adenomatous polyposis. For data analysis and variant filtering, an established bioinformatics pipeline including in-house tools was applied. Variants were filtered for rare truncating point mutations and copy-number variants assuming a dominant, recessive, or tumor suppressor model of inheritance. Subsequently, targeted sequence analysis of the most promising candidate genes was performed in a validation cohort of 191 unrelated patients. All relevant variants were validated by Sanger sequencing. The analysis of exome sequencing data resulted in the identification of rare loss-of-function germline mutations in three promising candidate genes (DSC2, PIEZO1, ZSWIM7). In the validation cohort, further variants predicted to be pathogenic were identified in DSC2 and PIEZO1. According to the somatic mutation spectra, the adenomas in this patient cohort follow the classical pathways of colorectal tumorigenesis. The present study identified three candidate genes which might represent rare causes for a predisposition to colorectal adenoma formation. Especially PIEZO1 (FAM38A) and ZSWIM7 (SWS1) warrant further exploration. To evaluate the clinical relevance of these genes, investigation of larger patient cohorts and functional studies are required. PMID:26780541

  4. Asymmetrical distribution of CpG in an 'average' mammalian gene.

    PubMed Central

    McClelland, M; Ivarie, R

    1982-01-01

    The frequency and distribution of the rare dinucleotide CpG was examined in 15 mammalian genes. CpG is highly methylated at cytosine in mammalian DNA (1,2) and 5-methylcytosine (5mC) is thought to undergo a transition mutation via deamination to produce thymine (3). This would result in the accumulation of TpG and CpA and depletion of CpG during evolution (4). Consistent with this hypothesis, the gene sample of 26,541 dinucleotides contained CpG at 40% the frequency expected by base composition and the CpG transition products, TpG+CpA, were significantly elevated at 124% of expected random frequency. However, because CpG occurs at only 25% of expected random frequency in the genome, the sampled genes were considerably enriched in this dinucleotide. CpGs were asymmetrically distributed in sequences flanking the genes. 5'-flanking sequences were enriched in CpG at 135% of the frequency expected assuming a symmetrical distribution of all the CpGs in the sampled genes (p less than 0.01), while 3'-flanking regions were depleted in CpG at 40% of expected values (p less than 0.0001). This asymmetry may reflect the role of 5-methylcytosine in gene expression. In contrast the frequencies of GpC and GpT+ ApC did not differ significantly from that predicted by base composition and these dinucleotides were not asymmetrically distributed. PMID:7155899

  5. Nuclear DNA sequences from the Middle Pleistocene Sima de los Huesos hominins.

    PubMed

    Meyer, Matthias; Arsuaga, Juan-Luis; de Filippo, Cesare; Nagel, Sarah; Aximu-Petri, Ayinuer; Nickel, Birgit; Martínez, Ignacio; Gracia, Ana; Bermúdez de Castro, José María; Carbonell, Eudald; Viola, Bence; Kelso, Janet; Prüfer, Kay; Pääbo, Svante

    2016-03-24

    A unique assemblage of 28 hominin individuals, found in Sima de los Huesos in the Sierra de Atapuerca in Spain, has recently been dated to approximately 430,000 years ago. An interesting question is how these Middle Pleistocene hominins were related to those who lived in the Late Pleistocene epoch, in particular to Neanderthals in western Eurasia and to Denisovans, a sister group of Neanderthals so far known only from southern Siberia. While the Sima de los Huesos hominins share some derived morphological features with Neanderthals, the mitochondrial genome retrieved from one individual from Sima de los Huesos is more closely related to the mitochondrial DNA of Denisovans than to that of Neanderthals. However, since the mitochondrial DNA does not reveal the full picture of relationships among populations, we have investigated DNA preservation in several individuals found at Sima de los Huesos. Here we recover nuclear DNA sequences from two specimens, which show that the Sima de los Huesos hominins were related to Neanderthals rather than to Denisovans, indicating that the population divergence between Neanderthals and Denisovans predates 430,000 years ago. A mitochondrial DNA recovered from one of the specimens shares the previously described relationship to Denisovan mitochondrial DNAs, suggesting, among other possibilities, that the mitochondrial DNA gene pool of Neanderthals turned over later in their history. PMID:26976447

  6. John Ash, AIA, Photographer August 1997. CONTEXT VIEW OF LOS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    John Ash, AIA, Photographer August 1997. CONTEXT VIEW OF LOS ANGELES CITY HALL SHOWING SOUTH ELEVATION WITH FIRST STREET PARK AND PARKING LOT IN FOREGROUND, FACING NORTH. - Los Angeles City Hall, 200 North Spring Street, Los Angeles, Los Angeles County, CA

  7. Compare Gene Profiles

    SciTech Connect

    2014-05-31

    Compare Gene Profiles (CGP) performs pairwise gene content comparisons among a relatively large set of related bacterial genomes. CGP performs pairwise BLAST among gene calls from a set of input genome and associated annotation files, and combines the results to generate lists of common genes, unique genes, homologs, and genes from each genome that differ substantially in length from corresponding genes in the other genomes. CGP is implemented in Python and runs in a Linux environment in serial or parallel mode.

  8. The Effects of Los Angeles Universal Preschool on Quality Preschool Teacher Retention in Los Angeles County

    ERIC Educational Resources Information Center

    Lopez Stevens, Holly Anne

    2010-01-01

    The purpose of this study was to determine if the implementation of Los Angeles Universal Preschool (LAUP) programs has a positive effect on the retention of quality preschool teachers in Los Angeles County. In prior work, preschool teacher retention is associated with wages, program structure, program process, professional development, and…

  9. New Generation of Los Alamos Opacity Tables

    NASA Astrophysics Data System (ADS)

    Colgan, James; Kilcrease, D. P.; Magee, N. H.; Sherrill, M. E.; Abdallah, J.; Hakel, P.; Fontes, C. J.; Guzik, J. A.; Mussack, K. A.

    2016-05-01

    We present a new generation of Los Alamos OPLIB opacity tables that have been computed using the ATOMIC code. Our tables have been calculated for all 30 elements from hydrogen through zinc and are publicly available through our website. In this poster we discuss the details of the calculations that underpin the new opacity tables. We also show several recent applications of the use of our opacity tables to solar modeling and other astrophysical applications. In particular, we demonstrate that use of the new opacities improves the agreement between solar models and helioseismology, but does not fully resolve the long-standing `solar abundance' problem. The Los Alamos National Laboratory is operated by Los Alamos National Security, LLC for the National Nuclear Security Administration of the U.S. Department of Energy under Contract No. DE-AC5206NA25396.

  10. Status of Monte Carlo at Los Alamos

    SciTech Connect

    Thompson, W.L.; Cashwell, E.D.

    1980-01-01

    At Los Alamos the early work of Fermi, von Neumann, and Ulam has been developed and supplemented by many followers, notably Cashwell and Everett, and the main product today is the continuous-energy, general-purpose, generalized-geometry, time-dependent, coupled neutron-photon transport code called MCNP. The Los Alamos Monte Carlo research and development effort is concentrated in Group X-6. MCNP treats an arbitrary three-dimensional configuration of arbitrary materials in geometric cells bounded by first- and second-degree surfaces and some fourth-degree surfaces (elliptical tori). Monte Carlo has evolved into perhaps the main method for radiation transport calculations at Los Alamos. MCNP is used in every technical division at the Laboratory by over 130 users about 600 times a month accounting for nearly 200 hours of CDC-7600 time.

  11. Publications of Los Alamos Research, 1983

    SciTech Connect

    Sheridan, C.J.; McClary, W.J.; Rich, J.A.; Rodriguez, L.L.

    1984-10-01

    This bibliography is a compilation of unclassified publications of work done at the Los Alamos National Laboratory for 1983. Papers published in 1982 are included regardless of when they were actually written. Publications received too late for inclusion in earlier compilations have also been listed. Declassification of previously classified reports is considered to constitute publication. All classified issuances are omitted - even those papers, themselves unclassified, which were published only as part of a classified document. If a paper was published more than once, all places of publication are included. The bibliography includes Los Alamos National Laboratory reports, papers released as non-Laboratory reports, journal articles, books, chapters of books, conference papers either published separately or as part of conference proceedings issued as books or reports, papers publishd in congressional hearings, theses, and US patents. Publications by Los Alamos authors that are not records of Laboratory-sponsored work are included when the Library becomes aware of them.

  12. Publications of Los Alamos research 1980

    SciTech Connect

    Salazar, C.A.; Willis, J.K.

    1981-09-01

    This bibliography is a compilation of unclassified publications of work done at the Los Alamos National Laboratory for 1980. Papers published in 1980 are included regardless of when they were actually written. Publications received too late for inclusion in earlier compilations have also been listed. Declassification of previously classified reports is considered to constitute publication. All classified issuances are omitted-even those papers, themselves unclassified, which were published only as part of a classified document. If a paper was pubished more than once, all places of publication are included. The bibliography includes Los Alamos National Laboratory reports, papers released as non-laboratory reports, journal articles, books, chapters of books, conference papers published either separately or as part of conference proceedings issued as books or reports, papers published in congressional hearings, theses, and US patents. Publications by Los Alamos authors that are not records of Laboratory-sponsored work are included when the Library becomes aware of them.

  13. Hepatic Gene Expression Profile in Mice Perorally Infected with Echinococcus multilocularis Eggs

    PubMed Central

    Gottstein, Bruno; Wittwer, Matthias; Schild, Marc; Merli, Michael; Leib, Stephen L.; Müller, Norbert; Müller, Joachim; Jaggi, Rolf

    2010-01-01

    Background Alveolar echinococcosis (AE) is a severe chronic hepatic parasitic disease currently emerging in central and eastern Europe. Untreated AE presents a high mortality (>90%) due to a severe hepatic destruction as a result of parasitic metacestode proliferation which behaves like a malignant tumor. Despite this severe course and outcome of disease, the genetic program that regulates the host response leading to organ damage as a consequence of hepatic alveolar echinococcosis is largely unknown. Methodology/Principal Findings We used a mouse model of AE to assess gene expression profiles in the liver after establishment of a chronic disease status as a result of a primary peroral infection with eggs of the fox tapeworm Echinococcus multilocularis. Among 38 genes differentially regulated (false discovery rate adjusted p≤0.05), 35 genes were assigned to the functional gene ontology group , while 3 associated with the functional group . Upregulated genes associated with could be clustered into functional subgroups including , <APCs>, , and . Two downregulated genes related to and , respectively. The genes either associated with an or an pathway. From the overexpressed genes, 18 genes were subsequently processed with a Custom Array microfluidic card system in order to assess respective expression status at the mRNA level relative to 5 reference genes (Gapdh, Est1, Rlp3, Mdh-1, Rpl37) selected upon a constitutive and stable expression level. The results generated by the two independent tools used for the assessment of gene expression, i.e., microarray and microfluidic card system, exhibited a high level of congruency (Spearman correlation rho = 0.81, p = 7.87e-5) and thus validated the applied

  14. A Survey of Methylated Candidate Tumor Suppressor Genes in Nasopharyngeal Carcinoma

    PubMed Central

    Loyo, Myriam; Brait, Mariana; Kim, Myoung S; Ostrow, Kimberly L.; Jie, Chunfa C; Chuang, Alice Y; Califano, Joseph A.; Liégeois, Nanette J; Begum, Shahnaz; Westra, William H; Hoque, Mohammad O; Tao, Qian; Sidransky, David

    2010-01-01

    Nasopharyngeal carcinoma (NPC) is a rare malignancy with unique genetic, viral and environmental characteristic that distinguishes it from other head and neck carcinomas. The clinical management of NPC remains challenging largely due to the lack of early detection strategies for this tumor. In the present study we have sought to identify novel genes involved in the pathogenesis of NPC that might provide insight into this tumor's biology and could potentially be used as biomarkers. To identify these genes, we studied the epigenetics of NPC by characterizing a panel of methylation markers. Eighteen genes were evaluated by quantitative methylation-specific PCR in cell lines as well as in tissue samples including 50 NPC tumors and 28 benign nasopharyngeal biopsies. Significance was evaluated using Fisher's exact test and quantitative values were optimized using cut off values derived from receiver-operator characteristic curves. The methylation status of AIM1, APC, CALCA, DCC, DLEC, DLC1, ESR, FHIT, KIF1A, and PGP9.5 was significantly associated with NPC compared to controls. The sensitivity of the individual genes ranged from 26 to 66% and the specificity was above 92% for all genes except FHIT. The combination of PGP9.5, KIF1A, and DLEC had a sensitivity of 84% and a specificity of 92%. Ectopic expression of DCC and DLC1 lead to decrease in colony formation and invasion properties. Our results indicate that methylation of novel biomarkers in NPC could be used to enhance early detection approaches. Additionally, our functional studies reveal previously unknown tumor suppressor roles in NPC. PMID:20473931

  15. DNA methylation status is more reliable than gene expression at detecting cancer in prostate biopsy

    PubMed Central

    Paziewska, A; Dabrowska, M; Goryca, K; Antoniewicz, A; Dobruch, J; Mikula, M; Jarosz, D; Zapala, L; Borowka, A; Ostrowski, J

    2014-01-01

    Background: We analysed critically the potential usefulness of RNA- and DNA-based biomarkers in supporting conventional histological diagnostic tests for prostate carcinoma (PCa) detection. Methods: Microarray profiling of gene expression and DNA methylation was performed on 16 benign prostatic hyperplasia (BPH) and 32 cancerous and non-cancerous prostate samples extracted by radical prostatectomy. The predictive value of the selected biomarkers was validated by qPCR-based methods using tissue samples extracted from the 58 prostates and, separately, using 227 prostate core biopsies. Results: HOXC6, AMACR and PCA3 expression showed the best discrimination between PCa and BPH. All three genes were previously reported as the most promising mRNA-based markers for distinguishing cancerous lesions from benign prostate lesions; however, none were sufficiently sensitive and specific to meet the criteria for a PCa diagnostic biomarker. By contrast, DNA methylation levels of the APC, TACC2, RARB, DGKZ and HES5 promoter regions achieved high discriminating sensitivity and specificity, with area under the curve (AUCs) reaching 0.95−1.0. Only a small overlap was detected between the DNA methylation levels of PCa-positive and PCa-negative needle biopsies, with AUCs ranging between 0.854 and 0.899. Conclusions: DNA methylation-based biomarkers reflect the prostate malignancy and might be useful in supporting clinical decisions for suspected PCa following an initial negative prostate biopsy. PMID:24937670

  16. Gene gymnastics

    PubMed Central

    Vijayachandran, Lakshmi S; Thimiri Govinda Raj, Deepak B; Edelweiss, Evelina; Gupta, Kapil; Maier, Josef; Gordeliy, Valentin; Fitzgerald, Daniel J; Berger, Imre

    2013-01-01

    Most essential activities in eukaryotic cells are catalyzed by large multiprotein assemblies containing up to ten or more interlocking subunits. The vast majority of these protein complexes are not easily accessible for high resolution studies aimed at unlocking their mechanisms, due to their low cellular abundance and high heterogeneity. Recombinant overproduction can resolve this bottleneck and baculovirus expression vector systems (BEVS) have emerged as particularly powerful tools for the provision of eukaryotic multiprotein complexes in high quality and quantity. Recently, synthetic biology approaches have begun to make their mark in improving existing BEVS reagents by de novo design of streamlined transfer plasmids and by engineering the baculovirus genome. Here we present OmniBac, comprising new custom designed reagents that further facilitate the integration of heterologous genes into the baculovirus genome for multiprotein expression. Based on comparative genome analysis and data mining, we herein present a blueprint to custom design and engineer the entire baculovirus genome for optimized production properties using a bottom-up synthetic biology approach. PMID:23328086

  17. SEDs at Los Alamos: A Personal Memoir

    NASA Astrophysics Data System (ADS)

    Bederson, Benjamin

    2001-03-01

    I have written this personal memoir approximately 55 years after the events I describe. It is based almost exclusively on memory, since apart from the diary I kept while on Tinian, I have few documents concerning it. It covers my service in the U.S. Army's Special Engineering Detachment (SED) in Oak Ridge and Los Alamos in 1944-45, on Tinian island, the launching pad for the bombing raids on Japan, in the summer and fall of 1945, and my return to Los Alamos until my discharge in January 1946.

  18. Upper Los Alamos canyon fact sheet

    SciTech Connect

    Berger, Jeffrey H

    2007-01-01

    Los Alamos National Laboratory is planning to make environmental assessments in portions of Upper Los Alamos Canyon. Upper Los Alamos Canyon is one of the areas included in the 2005 Consent Order agreed to by Los Alamos National Laboratory, the National Nuclear Security Administration, and the New Mexico Environment Department. As such, it must be evaluated for potential contamination. The area is located within and south of the Los Alamos townsite in Technical Areas 00, 01, 03, 32, 41, 43, and 61 of Los Alamos National Laboratory and includes a total of 115 solid waste management units and areas of concern. This area was home to some of the earliest operations at Los Alamos, dating from the 1940s. Of the 115 solid-waste management units and areas of concern, 54 have been addressed previously. The remaining 61 are the focus of this project. These include septic tanks and outfalls, sanitary and industrial waste lines, storm drains, soil contamination areas, landfill and surface disposal areas, transformer sites, and incinerators. The Consent Order requires the Laboratory to evaluate historical work sites for the potential presence of residual contamination. It also requires the Laboratory to identify and implement corrective actions should contamination be found. The Laboratory began performing these types of activities in the 1990s. The Upper Los Alamos Canyon project entails: (1) collecting soil and rock samples using the most efficient and least-invasive methods practicable; (2) defining the nature and extent of any residual contamination associated with each solid waste management unit or area of concern; and (3) gathering additional data if needed to evaluate potential remedial alternatives. A variety of methods, including studies of engineering drawings, nonintrusive geophysical surveys, and trenching, may be used to identify the final sampling locations. The field team then determines which collection method to use at each location, based on such site

  19. Plutonium 238 facilities at Los Alamos

    NASA Astrophysics Data System (ADS)

    Rinehart, Gary H.

    1991-01-01

    Plutonium 238 operations at Los Alamos are performed at the Plutonium Facility (TA-55), the Chemistry and Metallurgy Research (CMR) Building, and the Radioisotope Fuels Impact Test Facility. The plutonium 238 facilities at Los Alamos support a wide variety of heat source activities including development of new fuel forms and containment materials, research on the high temperature properties of containment materials, investigation of the high temperature compatibility of fuels with potential container materials, processing plutonium 238 fuel forms, manufacture of heat sources under quality assurance surveillance, and performing safety testing on heat sources and radioisotope thermoelectric generators.

  20. Plutonium-238 facilities at Los Alamos

    NASA Astrophysics Data System (ADS)

    Rinehart, Gary H.

    Plutonium-238 operations at Los Alamos are performed at the Plutonium Facility (TA-55), the Chemistry and Metallurgy Research (CMR) Building, and the Radioisotope Fuels Impact Test Facility. The plutonium-238 facilities at Los Alamos support a wide variety of heat source activities including development of new fuel forms and containment materials, research on the high temperature properties of containment materials, investigation of the high temperature compatibility of fuels with potential container materials, processing plutonium-238 fuel forms, manufacture of heat sources under quality assurance surveillance, and performing safety testing on heat sources and radioisotope thermoelectric generators.

  1. Proceedings of the Los Alamos neutrino workshop

    SciTech Connect

    Boehm, F.; Stephenson, G.J. Jr.

    1982-08-01

    A workshop on neutrino physics was held at Los Alamos from June 8 to 12, 1981. The material presented has been provided in part by the organizers, in part by the chairmen of the working sessions. Closing date for contributions was October 1981.

  2. Small mammals from Sima de los Huesos.

    PubMed

    Cuenca-Bescós, G; Laplana Conesa, C; Canudo, J I; Arsuaga, J L

    1997-01-01

    A small collection of rodents from Sima de los Huesos helps to clarify the stratigraphic position of this famous human locality. The presence of Allocricetus bursae and Pliomys lenki relictus and the size of A. bursae, Apodemus sylvaticus and Eliomys quercinus suggest a Middle Pleistocene age (Saalian) to the Clays where humans have been found. PMID:9300341

  3. Los Compadres: ESL Student Mentor Program.

    ERIC Educational Resources Information Center

    Coryell, Joellen; Gonzales, Edna; Claudia, Cary; Cisneros, Joe

    Los Compadres is a program that pairs advanced high school Spanish students with elementary English-as-a-Second-Language native Spanish speakers. Teachers prepare lessons based upon authentic literature, written in English and Spanish, which include vocabulary review and literature response activities. The high school students prepare for the…

  4. REACTIVE HYDROCARBON CONTROL COSTS FOR LOS ANGELES

    EPA Science Inventory

    This report documents the results of a study to determine the costs associated with controlling reactive organic emissions in the Metropolitan Los Angeles Air Quality Control Region. An inventory of organic emissions from 26 categories of stationary and mobile sources was develop...

  5. Induction inserts at the Los Alamos PSR

    SciTech Connect

    King-Yuen Ng

    2002-09-30

    Ferrite-loaded induction tuners installed in the Los Alamos Proton Storage Ring have been successful in compensating space-charge effects. However, the resistive part of the ferrite introduces unacceptable microwave instability and severe bunch lengthening. An effective cure was found by heating the ferrite cores up to {approx} 130 C. An understanding of the instability and cure is presented.

  6. African American Art: A Los Angeles Legacy.

    ERIC Educational Resources Information Center

    Walker, Harriet

    This curriculum unit focuses on the importance of Los Angeles (California) as a center for African American art and shows how African American artists have developed their own styles and how critics and collectors have encouraged them. The unit consists of four lessons, each of which can stand alone or be used in conjunction with the others. It…

  7. Educational and Demographic Profile: Los Angeles County

    ERIC Educational Resources Information Center

    California Postsecondary Education Commission, 2004

    2004-01-01

    This profile uniquely presents a variety of educational and socioeconomic information for Los Angeles County, nearby counties, and the state. The profile highlights the relationship between various factors that affect the economic well-being of individuals and communities. This presentation of information provides a framework for enhanced…

  8. Minorities in Suburbs: The Los Angeles Experience.

    ERIC Educational Resources Information Center

    Rabinovitz, Francine F.; Siembieda, William J.

    This book focuses on black suburbanization in the Los Angeles area, and questions whether the national increase in black suburbanization should be viewed with optimism or pessimism. The study addresses three questions: (1) Does the presence of substantial black populations in suburban areas represent suburbanization as it is normally thought of,…

  9. Los Alamos waste drum shufflers users manual

    SciTech Connect

    Rinard, P.M.; Adams, E.L.; Painter, J.

    1993-08-24

    This user manual describes the Los Alamos waste drum shufflers. The primary purpose of the instruments is to assay the mass of {sup 235}U (or other fissile materials) in drums of assorted waste. It can perform passive assays for isotopes that spontaneously emit neutrons or active assays using the shuffler technique as described on this manual.

  10. Los Angeles Settles ACLU Suit on Layoffs

    ERIC Educational Resources Information Center

    Sawchuk, Stephen

    2010-01-01

    A settlement crafted last week seeking to curb the use of seniority as a factor in teacher layoffs in the Los Angeles school system could become one of the nation's most far-reaching overhauls of the "last hired, first fired" policies common in school districts. If approved by a judge, the settlement would shield up to 45 low-performing schools in…

  11. Latina Adolescent Childbearing in East Los Angeles.

    ERIC Educational Resources Information Center

    Erickson, Pamela I.

    This book is about teenage pregnancy among Latina teenagers in East Los Angeles (California). It focuses on teenage pregnancy and motherhood among economically disadvantaged Latinas aged 17 and under. The young mothers in this study were participants in a series of intervention efforts to prevent repeat pregnancy at a family planning clinic. This…

  12. Amoxicillin pulsatile - MiddleBrook: APC 111, APC-111, PULSYS-enhanced amoxicillin.

    PubMed

    2007-01-01

    MiddleBrook Pharmaceuticals (formerly Advancis Pharmaceutical) is developing an improved version of amoxicillin using its pulsatile oral drug delivery technology, called PULSYS. Amoxicillin PULSYS is intended to provide a lower treatment dose, once-daily alternative to currently approved amoxicillin and penicillin regimens for the treatment of adolescents/adults with pharyngitis and/or tonsillitis. If amoxicillin PULSYS is approved, it will be the first and only once-daily amoxicillin therapy approved for use in the US. Regulatory submissions for the treatment of pharyngitis/tonsillitis have been made in the US. Amoxicillin PULSYS is in clinical development for the treatment of pharyngitis and/or tonsillitis due to group A streptococcal infections in adolescents/adults as a tablet formulation. MiddleBrook was conducting clinical development of a sprinkle formulation for children. However, this has been put on hold for financial reasons. MiddleBrook is seeking regulatory approval for this product as a 505(b)(2) product, which is one that is not considered to be a completely new product, but is also not a generic product. It is a product with some differences from a previously approved product and clinical data to support such differences are required; however, the basic safety and efficacy studies may have been conducted by other organisations. In June 2007, Advancis Pharmaceutical was renamed as MiddleBrook Pharmaceuticals, Inc. MiddleBrook and Par Pharmaceuticals entered a co-promotion agreement for this product in June 2004. Par was to fund future development in exchange for co-exclusive marketing rights and exclusive rights to sell amoxicillin PULSYS. MiddleBrook retained responsibility for the manufacturing programme and also retained all patents and brand names and was responsible for their enforcement. However, this collaboration was subsequently terminated in August 2005 by Par Pharmaceutical. MiddleBrook received the US $4.75 million R&D reimbursement payment due in the third quarter of 2005 and expects no further payments under the collaboration. Under certain circumstances, the termination clauses of the agreement may entitle Par to receive a share of net profits up to 50% of its total US $23 million investment in the development of certain amoxicillin PULSYS products, should a product covered by the agreement be successfully commercialised. Following the end of the first quarter of 2005, MiddleBrook entered into agreements with Clonmel Healthcare Ltd (STADA Group), which will provide MiddleBrook with commercial supply of its amoxicillin PULSYS products being evaluated in phase III trials. MiddleBrook has closed US $24 million in private placement of common equity; funds will be used to support the regulatory approval process for the once-daily amoxicillin pulsatile product. The company conducted phase III trials of this once-daily pulsatile amoxicillin product for the treatment of pharyngitis/tonsillitis due to group A streptococcal infections (commonly referred to as strep throat). Two trials of a 775 mg tablet formulation were conducted in adolescent/adult populations. In December 2006, the company entered into a definitive private placement agreement, raising US $18 million in gross proceeds. It intends to use the proceeds to prepare for the potential commercial launch of amoxicillin PULSYS and to continue the development of other products. The first of the adolescent/adult phase III trials was initiated in October 2004 in the US. This double-blind, non-inferiority, pivotal trial enrolled 510 such patients who received amoxicillin PULSYS 775 mg administered in tablet form once daily for 7 days or the US FDA standard comparator regimen, penicillin 250 mg administered four times daily for 10 days. However, in June 2005, the company reported that amoxicillin PULSYS failed to achieve the primary endpoint of this trial (i.e. bacterial eradication). The company initiated the second adolescent/adult phase III trial in November 2005. This two-arm, double-blind, double-dummy, non-inferiority trial enrolled a total of 620 patients from the US and Canada. Patients received amoxicillin PULSYS (775 mg tablet once daily for 10 days) or the standard penicillin regimen previously mentioned. Top-line results presented in August 2006 showed that the desired microbiological and clinical endpoints were achieved in this trial. In addition, the trial showed that amoxicillin PULSYS achieved 85% bacterial eradication in the per-protocol population, in accordance with the FDA's guidance for approval as a first-line therapy for pharyngitis. MiddleBrook has completed a phase I dose finding trial of its paediatric 'sprinkle' formulation of amoxicillin PULSYS in healthy volunteers. The company commenced a two-arm, investigator-blinded, non-inferiority, US-based, phase III trial in Janary 2005 to evaluate a 'sprinkle' formulation of amoxicillin PULSYS among paediatric patients with acute pharyngitis/tonsillitis caused by group A streptococcal infections. The drug was administered in multiparticulate granules, designed to be sprinkled over food, in two dosages: 475 mg once daily in patients aged 6 months to 4 years, and 775 mg once daily for children aged 5-12 years. Patients were given either 7 days' treatment with amoxicillin PULSYS or penicillin VK four times daily for 10 days. The primary endpoint of the paediatric trial was the same as the adult one. However, in July 2005, the company reported that the product failed to achieve its desired microbiological and clinical endpoints (primary and secondary) in this trial. MiddleBrook was to review the full data and evaluate what steps, if any, could be taken to improve future outcomes. PMID:17963430

  13. The Contribution of Whole Gene Deletions and Large Rearrangements to the Mutation Spectrum in Inherited Tumor Predisposing Syndromes.

    PubMed

    Smith, Miriam J; Urquhart, Jill E; Harkness, Elaine F; Miles, Emma K; Bowers, Naomi L; Byers, Helen J; Bulman, Michael; Gokhale, Carolyn; Wallace, Andrew J; Newman, William G; Evans, D Gareth

    2016-03-01

    Heterozygous whole gene deletions (WGDs), and intragenic microdeletions, account for a significant proportion of mutations underlying cancer predisposition syndromes. We analyzed the frequency and genotype-phenotype correlations of microdeletions in 12 genes (BRCA1, BRCA2, TP53, MSH2, MLH1, MSH6, PMS2, NF1, NF2, APC, PTCH1, and VHL) representing seven tumor predisposition syndromes in 5,897 individuals (2,611 families) from our center. Overall, microdeletions accounted for 14% of identified mutations. As expected, smaller deletions or duplications were more common (12%) than WGDs (2.2%). Where a WGD was identified in the germline in NF2, the mechanism of somatic second hit was not deletion, as previously described for NF1. For neurofibromatosis type 1 and 2, we compared the mechanism of germline deletion. Unlike NF1, where three specific deletion sizes account for most germline WGDs, NF2 deletion breakpoints were different across seven samples tested. One of these deletions was 3.93 Mb and conferred a severe phenotype, thus refining the region for a potential NF2 modifier gene to a 2.04-Mb region on chromosome 22. The milder phenotype of NF2 WGDs may be due to the apparent absence of chromosome 22 loss as the second hit. These observations of WGD phenotypes will be helpful for interpreting incidental findings from microarray analysis and next-generation sequencing. PMID:26615784

  14. Los Alamos Fires From Landsat 7

    NASA Technical Reports Server (NTRS)

    2002-01-01

    On May 9, 2000, the Landsat 7 satellite acquired an image of the area around Los Alamos, New Mexico. The Landsat 7 satellite acquired this image from 427 miles in space through its sensor called the Enhanced Thematic Mapper Plus (ETM+). Evident within the imagery is a view of the ongoing Cerro Grande fire near the town of Los Alamos and the Los Alamos National Laboratory. Combining the high-resolution (30 meters per pixel in this scene) imaging capacity of ETM+ with its multi-spectral capabilities allows scientists to penetrate the smoke plume and see the structure of the fire on the surface. Notice the high-level of detail in the infrared image (bottom), in which burn scars are clearly distinguished from the hotter smoldering and flaming parts of the fire. Within this image pair several features are clearly visible, including the Cerro Grande fire and smoke plume, the town of Los Alamos, the Los Alamos National Laboratory and associated property, and Cerro Grande peak. Combining ETM+ channels 7, 4, and 2 (one visible and two infrared channels) results in a false color image where vegetation appears as bright to dark green (bottom image). Forested areas are generally dark green while herbaceous vegetation is light green. Rangeland or more open areas appear pink to light purple. Areas with extensive pavement or urban development appear light blue or white to purple. Less densely-developed residential areas appear light green and golf courses are very bright green. The areas recently burned appear black. Dark red to bright red patches, or linear features within the burned area, are the hottest and possibly actively burning areas of the fire. The fire is spreading downslope and the front of the fire is readily detectable about 2 kilometers to the west and south of Los Alamos. Combining ETM+ channels 3, 2, and 1 provides a true-color image of the greater Los Alamos region (top image). Vegetation is generally dark to medium green. Forested areas are very dark green

  15. Germline Mutations in Predisposition Genes in Pediatric Cancer

    PubMed Central

    Edmonson, Michael N.; Gruber, Tanja A.; Easton, John; Hedges, Dale; Ma, Xiaotu; Zhou, Xin; Yergeau, Donald A.; Wilkinson, Mark R.; Vadodaria, Bhavin; Chen, Xiang; McGee, Rose B.; Hines-Dowell, Stacy; Nuccio, Regina; Quinn, Emily; Shurtleff, Sheila A.; Rusch, Michael; Patel, Aman; Becksfort, Jared B.; Wang, Shuoguo; Weaver, Meaghann S.; Ding, Li; Mardis, Elaine R.; Wilson, Richard K.; Gajjar, Amar; Ellison, David W.; Pappo, Alberto S.; Pui, Ching-Hon; Downing, James R.

    2016-01-01

    BACKGROUND The prevalence and spectrum of predisposing mutations among children and adolescents with cancer are largely unknown. Knowledge of such mutations may improve the understanding of tumorigenesis, direct patient care, and enable genetic counseling of patients and families. METHODS In 1120 patients younger than 20 years of age, we sequenced the whole genomes (in 595 patients), whole exomes (in 456), or both (in 69). We analyzed the DNA sequences of 565 genes, including 60 that have been associated with autosomal dominant cancer-predisposition syndromes, for the presence of germline mutations. The pathogenicity of the mutations was determined by a panel of medical experts with the use of cancer-specific and locus-specific genetic databases, the medical literature, computational predictions, and second hits identified in the tumor genome. The same approach was used to analyze data from 966 persons who did not have known cancer in the 1000 Genomes Project, and a similar approach was used to analyze data from an autism study (from 515 persons with autism and 208 persons without autism). RESULTS Mutations that were deemed to be pathogenic or probably pathogenic were identified in 95 patients with cancer (8.5%), as compared with 1.1% of the persons in the 1000 Genomes Project and 0.6% of the participants in the autism study. The most commonly mutated genes in the affected patients were TP53 (in 50 patients), APC (in 6), BRCA2 (in 6), NF1 (in 4), PMS2 (in 4), RB1 (in 3), and RUNX1 (in 3). A total of 18 additional patients had protein-truncating mutations in tumor-suppressor genes. Of the 58 patients with a predisposing mutation and available information on family history, 23 (40%) had a family history of cancer. CONCLUSIONS Germline mutations in cancer-predisposing genes were identified in 8.5% of the children and adolescents with cancer. Family history did not predict the presence of an underlying predisposition syndrome in most patients. (Funded by the American

  16. Gene doping: gene delivery for olympic victory

    PubMed Central

    Gould, David

    2013-01-01

    With one recently recommended gene therapy in Europe and a number of other gene therapy treatments now proving effective in clinical trials it is feasible that the same technologies will soon be adopted in the world of sport by unscrupulous athletes and their trainers in so called ‘gene doping’. In this article an overview of the successful gene therapy clinical trials is provided and the potential targets for gene doping are highlighted. Depending on whether a doping gene product is secreted from the engineered cells or is retained locally to, or inside engineered cells will, to some extent, determine the likelihood of detection. It is clear that effective gene delivery technologies now exist and it is important that detection and prevention plans are in place. PMID:23082866

  17. Prevalence of Germline Mutations in Cancer Predisposition Genes in Patients with Pancreatic Cancer

    PubMed Central

    Grant, Robert C.; Selander, Iris; Connor, Ashton A.; Selvarajah, Shamini; Borgida, Ayelet; Briollais, Laurent; Petersen, Gloria M.; Lerner-Ellis, Jordan; Holter, Spring; Gallinger, Steven

    2015-01-01

    Background & Aims We investigated the prevalence of germline mutations in APC, ATM, BRCA1, BRCA2, CDKN2A, MLH1, MSH2, MSH6, PALB2, PMS2, PRSS1, STK11, and TP53 in patients with pancreatic cancer. Methods The Ontario Pancreas Cancer Study enrolls consenting participants with pancreatic cancer from a province-wide electronic pathology database; 708 probands were enrolled from April 2003 through August 2012. To improve precision of BRCA2 prevalence estimates, 290 probands were randomly selected from 3 strata, based on family history of breast and/or ovarian cancer, pancreatic cancer, or neither. Germline DNA was analyzed by next-generation sequencing using a custom multiple-gene panel. Mutation prevalence estimates were calculated from the sample for the entire cohort. Results Eleven pathogenic mutations were identified: 3 in ATM, 1 in BRCA1, 2 in BRCA2, 1 in MLH1, 2 in MSH2, 1 in MSH6, and 1 in TP53. The prevalence of mutations in all 13 genes was 3.8% (95% confidence interval, 2.1%–5.6%). Carrier status was significantly associated with breast cancer in the proband or first-degree relative (P<.01), and colorectal cancer in the proband or first-degree relative (P<.01), but not family history of pancreatic cancer, age of diagnosis, or stage at diagnosis. Of patients with a personal or family history of breast and colorectal cancer, 10.7% (4.4%–17.0%) and 11.1% (3.0%–19.1%) carried pathogenic mutations, respectively. Conclusions A small but clinically important proportion of pancreatic cancer is associated with mutations in known predisposition genes. The heterogeneity of mutations identified in this study demonstrates the value of using a multiple-gene panel in pancreatic cancer. PMID:25479140

  18. Autism and Genes

    ERIC Educational Resources Information Center

    National Institutes of Health, 2005

    2005-01-01

    This document defines and discusses autism and how genes play a role in the condition. Answers to the following questions are covered: (1) What are genes? (2) What is autism? (3) What causes autism? (4) Why study genes to learn about autism? (5) How do researchers look for the genes involved in autism? (screen the whole genome; conduct cytogenetic…

  19. Compare Gene Profiles

    Energy Science and Technology Software Center (ESTSC)

    2014-05-31

    Compare Gene Profiles (CGP) performs pairwise gene content comparisons among a relatively large set of related bacterial genomes. CGP performs pairwise BLAST among gene calls from a set of input genome and associated annotation files, and combines the results to generate lists of common genes, unique genes, homologs, and genes from each genome that differ substantially in length from corresponding genes in the other genomes. CGP is implemented in Python and runs in a Linuxmore » environment in serial or parallel mode.« less

  20. Los Angeles County Poor Farm, Patient Wards 201, 202, 203, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Los Angeles County Poor Farm, Patient Wards 201, 202, 203, 204, 205, 206, 207, 208 & 209 - Type A Plan, 7601 Imperial Highway; bounded by Esperanza Street, Hawthorn Avenue, Laurel Street, and Descanso Street, Downey, Los Angeles County, CA