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Sample records for 1p loh 19q

  1. Shared allelic losses on chromosomes 1p and 19q suggest a common origin of oligodendroglioma and oligoastrocytoma

    SciTech Connect

    Kraus, J.A.; Koopmann, J.; Kaskel, P.

    1995-01-01

    Loss of heterozygosity (LOH) in specific chromosomal regions, which are likely to harbor tumor suppressor genes, has been associated with human gliomas. In this study we have analyzed astrocytic and oligodendroglial tumors for LOH on chromosomes 1 and 19. By microsatellite analysis LOH was found on chromosome arm 1p in 6/15 oligodendrogliomas WHO grade II and III, 12/25 oligoastrocytomas WHO grade II and III, 6/79 glioblastomas WHO grade IV, 5/44 astrocytomas WHO grade II and III and 0/23 pilocystic astrocytomas WHO grade I. The high incidence of LOH on chromosome arm 1p in oligodendrogliomas and oligoastrocytomas indicates that a putative tumor suppressor gene in this region is involved in the formation of gliomas with oligodendroglial features. Furthermore, the frequent involvement of chromosome arm 1p in oligodendrogliomas and oligoastrocytomas, but not in astrocytomas, suggests that genetically oligoastrocytoma is more similar to oligodendroglioma than to astrocytoma. In order to support this hypothesis, oligodendroglial and astrocytic areas in three mixed oligoastrocytomas were examined differentially for LOH 1p and for LOH 19q, the second genetic region believed to be affected in these tumors. All three tumors had LOH of 1p and LOH of 19q in both areas of oligodendroglial and of astrocytic differentiation. These findings show that the astrocytic and oligodendroglial portions of oligoastrocytoma share molecular genetic features and probably are of monoclonal origin. 32 refs., 2 figs., 1 tab.

  2. Not all 1p/19q non-codeleted oligodendroglial tumors are astrocytic

    PubMed Central

    Aibaidula, Abudumijiti; Chen, Hong; Tang, Qisheng; Li, Kay Ka-Wai; Chung, Nellie Yuk-Fei; Chan, Danny Tat-Ming; Poon, Wai Sang; Mao, Ying; Wu, Jinsong; Zhou, Liangfu; Chan, Aden Ka-yin; Ng, Ho-Keung

    2016-01-01

    Although 1p/19q codeletion is the genetic hallmark defining oligodendrogliomas, approximately 30-40% of oligodendroglial tumors have intact 1p/19q in the literature and they demonstrate a worse prognosis. This group of 1p/19q intact oligodendroglial tumors is frequently suggested to be astrocytic in nature with TP53 and ATRX mutations but actually remains under-investigated. In the present study, we provided evidence that not all 1p/19q intact oligodendroglial tumors are astrocytic through histologic and molecular approaches. We examined 1p/19q status by FISH in a large cohort of 337 oligodendroglial tumors and identified 39.8% lacking 1p/19q codeletion which was independently associated with poor prognosis. Among this 1p/19q intact oligodendroglial tumor cohort, 58 cases demonstrated classic oligodendroglial histology which showed older patient age, better prognosis, association with grade III histology, PDGFRA expression, TERTp mutation, as well as frequent IDH mutation. More than half of the 1p/19q intact oligodendroglial tumors showed lack of astrocytic defining markers, p53 expression and ATRX loss. TP53 mutational analysis was additionally conducted in 45 cases of the 1p/19q intact oligodendroglial tumors. Wild-type TP53 was detected in 71.1% of cases which was associated with classic oligodendroglial histology. Importantly, IDH and TERTp co-occurred in 75% of 1p/19q intact, TP53 wild-type oligodendrogliomas, highlighting the potential of the co-mutations in assisting diagnosis of oligodendrogliomas in tumors with clear cell morphology and non-codeleted 1p/19q status. In summary, our study demonstrated that not all 1p/19q intact oligodendroglial tumors are astrocytic and co-evaluation of IDH and TERTp mutation could potentially serve as an adjunct for diagnosing 1p/19q intact oligodendrogliomas. PMID:27556304

  3. Codeletion of 1p and 19q determines distinct gene methylation and expression profiles in IDH-mutated oligodendroglial tumors.

    PubMed

    Mur, Pilar; Mollejo, Manuela; Ruano, Yolanda; de Lope, Ángel Rodríguez; Fiaño, Concepción; García, Juan Fernando; Castresana, Javier S; Hernández-Laín, Aurelio; Rey, Juan A; Meléndez, Bárbara

    2013-08-01

    Oligodendroglial tumors (OTs) are primary brain tumors that show variable clinical and biological behavior. The 1p/19q codeletion is frequent in these tumors, indicating a better prognosis and/or treatment response. Recently, the prognostically favorable CpG island methylator phenotype (CIMP) in gliomas (G-CIMP+) was associated with mutations in the isocitrate dehydrogenase 1 and isocitrate dehydrogenase 2 (IDH) genes, as opposed to G-CIMP- tumors, highlighting the relevance of epigenetic mechanisms. We performed a whole-genome methylation study in 46 OTs, and a gene expression study of 25 tumors, correlating the methylation and transcriptomic profiles with molecular and clinical variables. Here, we identified two different epigenetic patterns within the previously described main G-CIMP+ profile. Both IDH mutation-associated methylation profiles featured one group of OTs with 1p/19q loss (CD-CIMP+), most of which were pure oligodendrogliomas, and a second group with intact 1p/19q and frequent TP53 mutation (CIMP+), most of which exhibited a mixed histopathology. A third group of OTs lacking the CIMP profile (CIMP-), and with a wild-type IDH and an intact 1p/19q, similar to the G-CIMP- subgroup, was also observed. The three CIMP groups presented a significantly better (CD-CIMP+), intermediate (CIMP+) or worse (CIMP-) prognosis. Furthermore, transcriptomic analyses revealed CIMP-specific gene expression signatures, indicating the impact of genetic status (IDH mutation, 1p/19q codeletion, TP53 mutation) on gene expression, and pointing to candidate biomarkers. Therefore, the CIMP profiles contributed to the identification of subgroups of OTs characterized by different prognoses, histopathologies, molecular features and gene expression signatures, which may help in the classification of OTs.

  4. Allelic loss of 9p21.3 is a prognostic factor in 1p/19q codeleted anaplastic gliomas

    PubMed Central

    Alentorn, Agustí; Dehais, Caroline; Ducray, François; Carpentier, Catherine; Mokhtari, Karima; Figarella-Branger, Dominique; Chinot, Olivier; Cohen-Moyal, Elisabeth; Ramirez, Carole; Loiseau, Hugues; Elouahdani-Hamdi, Selma; Beauchesne, Patrick; Langlois, Olivier; Desenclos, Christine; Guillamo, Jean-Sébastien; Dam-Hieu, Phong; Ghiringhelli, François; Colin, Philippe; Godard, Joel; Parker, Fabrice; Dhermain, Frédéric; Carpentier, Antoine F.; Frenel, Jean-Sebastien; Menei, Philippe; Bauchet, Luc; Faillot, Thierry; Fesneau, Mélanie; Fontaine, Denys; Motuo-Fotso, Marie-Jeannette; Vauleon, Elodie; Gaultier, Claude; Le Guerinel, Caroline; Gueye, Edouard-Marcel; Noel, Georges; Desse, Nicolas; Durando, Xavier; Barrascout, Eduardo; Wager, Michel; Ricard, Damien; Carpiuc, Ioana; Delattre, Jean-Yves

    2015-01-01

    Objectives: We aimed to study the potential clinical relevance of 9p allelic loss, with or without copy number variation, in 1p/19q codeleted anaplastic oligodendroglial tumors (AOTs). Methods: This study enrolled 216 patients with 1p/19q codeleted AOT. The prognostic value of 9p allelic loss was investigated using a French nation-wide prospective registry, POLA (prise en charge des tumeurs oligodendrogliales anaplasiques) and high-density single nucleotide polymorphism arrays. We validated our results using the Repository of Molecular Brain Neoplasia Data (REMBRANDT) dataset. Results: The minimal common region of allelic loss in chromosome arm 9p was 9p21.3. Allelic loss of 9p21.3, detected in 41.7% of tumors, was associated with shorter progression-free and overall survival rates in univariate (p = 0.008 and p < 0.001, respectively) and multivariate analyses (p = 0.009 and p = 0.009, respectively). This finding was validated in the REMBRANDT dataset in univariate and multivariate analysis (p = 0.01 and p = 0.01, respectively). Conclusion: Our study highlights a novel potential prognostic biomarker in 1p/19q codeleted AOT. Further prospective studies are warranted to investigate our finding. PMID:26385879

  5. ImmunoFISH Is a Reliable Technique for the Assessment of 1p and 19q Status in Oligodendrogliomas

    PubMed Central

    Duval, Céline; de Tayrac, Marie; Sanschagrin, François; Michaud, Karine; Gould, Peter Vincent; Saikali, Stéphan

    2014-01-01

    Objective To develop a new ImmunoFISH technique for the study of oligodendrogliomas by combining a standard immunohistochemical stain using MIB-1 antibody with a standard FISH technique using commercial 1p36 and 19q13 chromosomal probes. Methods Validation was performed by two observers on a series of 36 pre-selected oligodendrogliomas and compared to the results previously determined by FISH alone. Results The ImFISH technique is easy to perform and to analyze and is no more time-consuming than the usual FISH technique. Our results show that the inter-observer reliability of ImFISH is high (κ = 0.86 and 0.95 respectively for 1p and 19q). Compared to FISH, the ImFISH exhibits a very high sensitivity (∼100%) and specificity (∼90%) for 1p and/or 19q deleted cases. The sensitivity is high for normal cases (∼85%) and imbalanced cases (∼90%) with a specificity ranging between 50 and 85%. Finally, there were no significant differences between FISH and ImFISH results calculated on 60, 40 or 20 cells. Conclusion Our study demonstrates the reliability of the ImFISH technique in oligodendrogliomas and emphasizes its advantage in poorly cellular tumoral specimen. PMID:24949947

  6. Automated Analysis of 1p/19q Status by FISH in Oligodendroglial Tumors: Rationale and Proposal of an Algorithm

    PubMed Central

    Duval, Céline; de Tayrac, Marie; Michaud, Karine; Cabillic, Florian; Paquet, Claudie; Gould, Peter Vincent; Saikali, Stéphan

    2015-01-01

    Objective To propose a new algorithm facilitating automated analysis of 1p and 19q status by FISH technique in oligodendroglial tumors with software packages available in the majority of institutions using this technique. Methods We documented all green/red (G/R) probe signal combinations in a retrospective series of 53 oligodendroglial tumors according to literature guidelines (Algorithm 1) and selected only the most significant combinations for a new algorithm (Algorithm 2). This second algorithm was then validated on a prospective internal series of 45 oligodendroglial tumors and on an external series of 36 gliomas. Results Algorithm 2 utilizes 24 G/R combinations which represent less than 40% of combinations observed with Algorithm 1. The new algorithm excludes some common G/R combinations (1/1, 3/2) and redefines the place of others (defining 1/2 as compatible with normal and 3/3, 4/4 and 5/5 as compatible with imbalanced chromosomal status). The new algorithm uses the combination + ratio method of signal probe analysis to give the best concordance between manual and automated analysis on samples of 100 tumor cells (91% concordance for 1p and 89% concordance for 19q) and full concordance on samples of 200 tumor cells. This highlights the value of automated analysis as a means to identify cases in which a larger number of tumor cells should be studied by manual analysis. Validation of this algorithm on a second series from another institution showed a satisfactory concordance (89%, κ = 0.8). Conclusion Our algorithm can be easily implemented on all existing FISH analysis software platforms and should facilitate multicentric evaluation and standardization of 1p/19q assessment in gliomas with reduction of the professional and technical time required. PMID:26135922

  7. Chromogenic in situ hybridization is a reliable alternative to fluorescence in situ hybridization for diagnostic testing of 1p and 19q loss in paraffin-embedded gliomas.

    PubMed

    Lass, Ulrike; Hartmann, Christian; Capper, David; Herold-Mende, Christel; von Deimling, Andreas; Meiboom, Maren; Mueller, Wolf

    2013-05-01

    Recent studies imply the importance of rapid and reliable diagnostic assessment of 1p/19q status in oligodendroglial tumors. To date, fluorescence in situ hybridization (FISH) is the most commonly applied technique. FISH, however, has several technical shortcomings that are suboptimal for diagnostic applications: results must be viewed in a fluorescence microscope, results are usually evaluated by a single investigator only, and signal fading excludes physical archiving. Also, in gliomas, the distinction of diffusely infiltrating tumor cells from reactively altered normal tissue may be challenging in fluorescence microscopy. Dual-color chromogenic in situ hybridization (CISH) has started to replace FISH in some diagnostic tests performed in pathology. Here, we present the first single institute experience with a side-by-side analysis of 1p/19q FISH and CISH in a series of 42 consecutive gliomas. FISH and CISH produced identical results for 1p and 19q in 93% of cases (n = 39/42). Discrepant results were reevaluated by repeated FISH and a polymerase chain reaction (PCR)-based microsatellite marker analysis for loss of heterozygosity. Reevaluation confirmed CISH data in all three cases. We conclude that CISH is a reliable alternative in 1p/19q testing in paraffin-embedded tissues likely to be more sensitive to detect 1p/19q status than FISH analysis.

  8. Tumor cells with neuronal intermediate progenitor features define a subgroup of 1p/19q co-deleted anaplastic gliomas.

    PubMed

    Bielle, Franck; Ducray, François; Mokhtari, Karima; Dehais, Caroline; Adle-Biassette, Homa; Carpentier, Catherine; Chanut, Anaïs; Polivka, Marc; Poggioli, Sylvie; Rosenberg, Shai; Giry, Marine; Marie, Yannick; Duyckaerts, Charles; Sanson, Marc; Figarella-Branger, Dominique; Idbaih, Ahmed

    2016-08-20

    The integrated diagnosis of anaplastic oligodendroglioma, IDH mutant and 1p/19q co-deleted, grade III (O3(id) ) is a histomolecular entity that WHO 2016 classification distinguished from other diffuse gliomas by specific molecular alterations. In contrast, its cell portrait is less well known. The present study is focused on intertumor and intratumor, cell lineage-oriented, heterogeneity in O3(id) . Based on pathological, transcriptomic and immunophenotypic studies, a novel subgroup of newly diagnosed O3(id) overexpressing neuronal intermediate progenitor (NIP) genes was identified. This NIP overexpression pattern in O3(id) is associated with: (i) morphological and immunohistochemical similarities with embryonic subventricular zone, (ii) proliferating tumor cell subpopulation with NIP features including expression of INSM1 and no expression of SOX9, (iii) mutations in critical genes involved in NIP biology and, (iv) increased tumor necrosis. Interestingly, NIP tumor cell subpopulation increases in O3(id) recurrence compared with paired newly diagnosed tumors. Our results, validated in an independent cohort, emphasize intertumor and intratumor heterogeneity in O3(id) and identified a tumor cell subpopulation exhibiting NIP characteristics that is potentially critical in oncogenesis of O3(id) . A better understanding of spatial and temporal intratumor cell heterogeneity in O3(id) will open new therapeutic avenues overcoming resistance to current antitumor treatments.

  9. Impact of gross total resection in patients with WHO grade III glioma harboring the IDH 1/2 mutation without the 1p/19q co-deletion.

    PubMed

    Kawaguchi, Tomohiro; Sonoda, Yukihiko; Shibahara, Ichiyo; Saito, Ryuta; Kanamori, Masayuki; Kumabe, Toshihiro; Tominaga, Teiji

    2016-09-01

    The prognosis of patients with WHO grade III gliomas is highly dependent on their genomic status such as the isocitrate dehydrogenase (IDH) 1/2 mutation and1p/19q co-deletion. However, difficulties have been associated with determining which tumors have certain genomic profiles by preoperative radiographical modalities, and the role of surgical resection in achieving better outcomes remains unclear. This retrospective study included 124 consecutive patients with newly diagnosed grade III gliomas. The genomic status of IDH1/2 and 1p/19q was analyzed in these patients. Tumors were then divided into 3 subgroups based on their genomic status; the IDH 1/2 mutation with the 1p/19q co-deletion (1p/19q co-del), the IDH 1/2 mutation without the 1p/19q co-deletion (non-1p/19q co-del), and the IDH 1/2 wild type (IDH wt). Survival times were compared between patients who underwent gross total resection and those who did not (GTR versus non-GTR). The relationships between genomic statuses and MR imaging characteristics such as ring-like or nodular enhancements by gadolinium, and very low intensity on T1-weighted images with blurry enhancements (T1VL) were also examined. Among all patients with grade III gliomas, GTR patients had longer median survival and progression-free times than those of non-GTR patients (undefined versus 87 months, p = 0.097, and 124 versus 34 months, p = 0.059, respectively). No significant differences were observed in survival between GTR and non-GTR patients in the 1p/19q co-del group (p = 0.14), or between GTR and non-GTR patients in the IDH wt group (26 and 27 months, p = 0.29). On the other hand, in non-1p/19q co-del group, survival was significantly longer in GTR patients than in non-GTR patients (undefined versus 77 months, p = 0.005). Radiographically, T1VL was detected in most tumors in the non-1p/19q co-del group (78.2 %), but only 6 (21.4 %) and 17 (41.5 %) tumors in the 1p/19q co-del and IDH wt groups

  10. Impact of 1p/19q Codeletion and Histology on Outcomes of Anaplastic Gliomas Treated With Radiation Therapy and Temozolomide

    SciTech Connect

    Speirs, Christina K.; Simpson, Joseph R.; Robinson, Clifford G.; DeWees, Todd A.; Tran, David D.; Linette, Gerry; Chicoine, Michael R.; Dacey, Ralph G.; Rich, Keith M.; Dowling, Joshua L.; Leuthardt, Eric C.; Zipfel, Gregory J.; Kim, Albert H.; Huang, Jiayi

    2015-02-01

    Purpose: Anaplastic gliomas represent a heterogeneous group of primary high-grade brain tumors, and the optimal postoperative treatment remains controversial. In this report, we present our institutional data on the clinical outcomes of radiation therapy (RT) plus temozolomide (RT + TMZ) for anaplastic gliomas, stratified by histology and 1p/19q codeletion. Methods and Materials: A single-institution retrospective review was conducted of patients with supratentorial anaplastic oligodendroglioma (AO), mixed anaplastic oligoastrocytoma (AOA), and anaplastic astrocytoma (AA). After surgery, RT was delivered at a median total dose of 60 Gy (range, 31.6-63 Gy) in daily fractions. All patients received standard concurrent TMZ, with or without adjuvant TMZ. Histological/molecular subtypes were defined as codeleted AO/AOA, non-codeleted AO/AOA, and AA. Results: From 2000 to 2012, 111 cases met study criteria and were evaluable. Codeleted AO/AOA had superior overall survival (OS) to non-codeleted AO/AOA (91% vs 68% at 5 years, respectively, P=.02), whereas progression-free survival (PFS) was not significantly different (70% vs 46% at 5 years, respectively, P=.10). AA had inferior OS to non-codeleted AO/AOA (37% vs 68% at 5 years, respectively, P=.007) and inferior PFS (27% vs 46%, respectively, P=.03). On multivariate analysis, age, performance status, and histological or molecular subtype were independent predictors for both PFS and OS. Compared to historical controls, RT + TMZ provided comparable OS to RT with procarbazine, lomustine, and vincristine (RT + PCV) for codeleted AO/AOA, superior OS to RT alone for non-codeleted AO/AOA, and similar OS to RT alone for AA. Conclusions: RT + TMZ may be a promising treatment for both codeleted and non-codeleted AO/AOA, but its role for AA remains unclear.

  11. Mitotic index, microvascular proliferation, and necrosis define 3 groups of 1p/19q codeleted anaplastic oligodendrogliomas associated with different genomic alterations

    PubMed Central

    Figarella-Branger, Dominique; Mokhtari, Karima; Dehais, Caroline; Jouvet, Anne; Uro-Coste, Emmanuelle; Colin, Carole; Carpentier, Catherine; Forest, Fabien; Maurage, Claude-Alain; Vignaud, Jean-Michel; Polivka, Marc; Lechapt-Zalcman, Emmanuelle; Eimer, Sandrine; Viennet, Gabriel; Quintin-Roué, Isabelle; Aubriot-Lorton, Marie-Hélène; Diebold, Marie-Danièle; Loussouarn, Delphine; Lacroix, Catherine; Rigau, Valérie; Laquerrière, Annie; Vandenbos, Fanny; Michalak, Sophie; Sevestre, Henri; Peoch, Michel; Labrousse, François; Christov, Christo; Kemeny, Jean-Louis; Chenard, Marie-Pierre; Chiforeanu, Danchristian; Ducray, François; Idbaih, Ahmed; Desenclos, Christine; Menei, Philippe; Al Nader, Edmond; Godard, Joel; Servagi-Vernat, Stéphanie; Carpentier, Antoine; Loiseau, Hugues; Dam-Hieu, Phong; Guillamo, Jean Sebastien; Emery, Evelyne; Verelle, Pierre; Durando, Xavier; Faillot, Thierry; Le Guerinel, Caroline; Ghiringhelli, François; Parker, Fabrice; Adam, Clovis; Dubois, François; Ramirez, Carole; Gueye, Edouard Marcel; Honnorat, Jerome; Chinot, Olivier; Bauchet, Luc; Beauchesne, Patrick; Campone, Mario; Frenel, Jean Sébastien; Fontaine, Denys; Campello, Chantal; Roger, Pascal; Heitzmann, Anne; Fesneau, Mélanie; Delattre, Jean Yves; Elouadhani-Hamdi, Selma; Ricard, Damien; Colin, Philippe; Vauléon, Elodie; Langlois, Olivier; Fotso, Marie Janette Motsuo; Andraud, Marie; Mouton, Servane; Noel, Georges; Desse, Nicolas; Soulard, Raoulin; Cohen-Moyal, Elisabeth; Lubrano, Vincent; Dhermain, Frederic

    2014-01-01

    Background The aim of this study was to correlate histological features and molecular characteristics in anaplastic oligodendrogliomas (AOs). Methods The histological characteristics of 203 AO patients, enrolled in the French national network POLA, were analyzed. The genomic profiles of 191 cases were studied using genomic arrays. IDH mutational status was assessed by immunohistochemistry and direct sequencing. Results 1p/19q codeletion was present in 79% of cases and was associated with alpha-internexin expression (P < 10−4), IDH1/2 mutation (P < 10−4), chromosome 4 loss (P < 10−3), and better overall survival (P < 10−4). Based on mitotic index, microvascular proliferation (MVP), and necrosis, 3 groups of 1p/19q codeleted AOs were identified: (group 1) AO with more than 5 mitoses per 10-HPF, no MVP, and no necrosis; (group 2) AO with MVP and no necrosis; and (group 3) AO with MVP and necrosis. Compared with group 1, groups 2 and 3 AOs had a higher mean Ki-67 proliferation index and a higher rate of 9p and 9q losses. Compared with group 2, group 3 AOs had a higher number of chromosomal alterations including chromosome 4 loss. In the subgroup of 157 1p/19q codeleted AOs, chromosomal instability was associated with shorter progression-free survival (P = .024) and shorter overall survival (P = .023). Conclusions The present study shows that oligodendrogliomas with classic histological features remain a molecularly heterogeneous entity and should be stratified according to 1p/19q status because of its major prognostic relevance. Moreover, 1p/19q codeleted AOs are also heterogeneous. Interestingly, mitotic index, MVP, and necrosis help to classify them into 3 groups associated with distinct genomic alterations. PMID:24723566

  12. Radio-chemotherapy improves survival in IDH-mutant, 1p/19q non-codeleted secondary high-grade astrocytoma patients.

    PubMed

    Juratli, Tareq A; Lautenschläger, Tim; Geiger, Kathrin D; Pinzer, Thomas; Krause, Mechthild; Schackert, Gabriele; Krex, Dietmar

    2015-09-01

    Isocitrate dehydrogenase (IDH) mutations are beginning to drive decisions on therapy for glioma patients. Here we sought to determine the impact of adjuvant treatment in patients with IDH-mutant, 1p/19q non-codeleted secondary high-grade astrocytoma (sHGA) WHO grades III/IV. Clinical data of 109 sHGA patients grades III/IV, in addition to IDH mutation-, 1p/19q-codeletion- and MGMT-promoter methylation status-were retrospectively analyzed. Survival analysis in relation to adjuvant treatment modalities and molecular profiling were performed. Out of 109 patients, 88 patients (80.7 %) harbored IDH mutations, 30 patients had a 1p/19q-codeletion (27.5 %) and 69 patients (63.3 %) exhibited a methylated MGMT-promoter status. At a median follow-up of 9.8 years, 62 patients (57 %) died. The postsurgical treatment included: radio-chemotherapy (RT-CT; 54.5 %), RT alone (19.3 %), and CT alone (22.7 %). The median overall survival (OS) in the entire group was 3.4 years (1.9-6.7 years). Patients who received RT-CT had a significantly longer OS compared with those who underwent RT alone (6.5 vs. 1.2 years, HR 0.35, CI 0.32-0.51, p = 0.011). In the IDH-mutant 1p/19q non-codeleted sHGA subgroup the RT-CT cohort had a significantly longer OS in comparison to the RT cohort (6.4 vs. 1.2 years, HR 2.7, CI 1.1-6.5, p = 0.022). In the stepwise multivariable Cox model for OS of all 88 IDH-mutant sHGA patients, survival was strongly associated with only one factor, namely, adjuvant RT-CT at diagnosis of a sHGA. This retrospective long-term study demonstrates that RT and CT (mostly PCV) significantly improves progression-free and overall survival in IDH-mutant secondary high-grade astrocytoma patients, regardless of 1p/19q-codeletion status.

  13. Contribution of 1p, 19q, 9p and 10q Automated Analysis by FISH to the Diagnosis and Prognosis of Oligodendroglial Tumors According to WHO 2016 Guidelines

    PubMed Central

    Michaud, Karine; de Tayrac, Marie; D’Astous, Myreille; Duval, Céline; Paquet, Claudie; Samassekou, Oumar; Gould, Peter Vincent; Saikali, Stéphan

    2016-01-01

    Objective To study the feasibility and the diagnostic and prognostic interest of automated analysis of 1p, 19q, 9p and 10q status by FISH technique in oligodendroglial tumors. Methods We analyzed a retrospective series of 33 consecutive gliomas with oligodendroglial histology (originally diagnosed as 24 oligodendrogliomas and 9 oligoastrocytomas). For all cases, automated FISH analysis of 1p, 19q, 9p and 10q status were performed and compared to clinical and histological data, ATRX, IDH1R132H and alpha-internexin status (studied by immunohistochemistry) and overall survival (OS). Manual analysis of 9p and 10q status were also performed and compared to automated analysis to verify the concordance of the two methods. Results The 33 gliomas were reclassified into 13 low-grade oligodendrogliomas (OII), 10 anaplastic oligodendrogliomas (OIII), 3 diffuse astrocytomas (AII), 3 anaplastic astrocytomas (AIII) and 4 glioblastomas (GBM) according to the WHO 2016 histological criteria. The 1p and/or 19q imbalanced status were restricted to astrocytomas with no correlation to their grade or their OS. Chromosome 9p deletion was restricted to OIII (70%) and GBM (100%) and was correlated with a shorter OS in the total cohort (p = 0.0007), the oligodendroglioma cohort (p = 0.03) and the astrocytoma cohort (p = 0.001). Concordance between 9p manual and automated analysis was satisfactory (81%, κ = 0.69). Chromosome 10q deletion was restricted to GBMs (50%) and was correlated with a poor OS in both the total cohort (p = 0.003) and the astrocytoma (AS) cohort (p = 0.04). Concordance between manual and automated analysis was satisfactory (79%, κ = 0.62). Conclusion Automated analysis of 1p, 19q, 9p and 10q status by FISH is a reliable technique which allows for refined classification of oligodendroglial tumors. 1p and/or 19q imbalanced status is evidence of astrocytic differentiation. 9p deletion is found in high grade oligodendrogliomas and astrocytomas with a poor OS. 10q is

  14. Deferred radiotherapy and upfront procarbazine–ACNU–vincristine administration for 1p19q codeleted oligodendroglial tumors are associated with favorable outcome without compromising patient performance, regardless of WHO grade

    PubMed Central

    Hata, Nobuhiro; Yoshimoto, Koji; Hatae, Ryusuke; Kuga, Daisuke; Akagi, Yojiro; Suzuki, Satoshi O; Iwaki, Toru; Shono, Tadahisa; Mizoguchi, Masahiro; Iihara, Koji

    2016-01-01

    Recently updated phase III trials revealed the favorable effect of add-on procarbazine-lomustine-vincristine chemotherapy (CT) to radiotherapy (RT) in treating anaplastic oligodendrogliomas with 1p19q codeletion (codel). However, the underlying rationality of deferring RT and upfront CT administration for these tumors is yet to be elucidated. Here, we retrospectively analyzed the long-term outcome of our case series with oligodendroglial tumors treated with deferred RT and upfront procarbazine+nimustine+vincristine (PAV) in the introduction administration. We enrolled 36 patients with newly diagnosed oligodendroglial tumors (17, grade II and 19, grade III) treated during 1999–2012 and followed up for a median period of 69.0 months. Their clinical and genetic prognostic factors were analyzed, and progression-free survival, overall survival (OS), and deterioration-free survival (DFS) were evaluated. Regardless of the WHO grade, the 25 patients with 1p19q codel tumors never received RT initially, and of these 25, 23 received PAV treatment upfront. The 75% OS of patients with 1p19q codel tumor was 135.3 months (did not reach the median OS), indicating a favorable outcome. Multivariate analysis revealed that IDH mutation and 1p19q, not WHO grade, are independent prognostic factors; furthermore, IDH and 1p19q status stratified the cohort into 3 groups with significantly different OS. The DFS explained the prolonged survival without declining performance in patients with both grade II and III 1p19q codel tumors. Deferred RT and upfront PAV treatment for 1p19q codel oligodendrogliomas were associated with favorable outcomes without compromising performance status, regardless of WHO grade. PMID:27895504

  15. Assessing CpG island methylator phenotype, 1p/19q codeletion, and MGMT promoter methylation from epigenome-wide data in the biomarker cohort of the NOA-04 trial

    PubMed Central

    Wiestler, Benedikt; Capper, David; Hovestadt, Volker; Sill, Martin; Jones, David T.W.; Hartmann, Christian; Felsberg, Joerg; Platten, Michael; Feiden, Wolfgang; Keyvani, Kathy; Pfister, Stefan M.; Wiestler, Otmar D.; Meyermann, Richard; Reifenberger, Guido; Pietsch, Thorsten; von Deimling, Andreas; Weller, Michael; Wick, Wolfgang

    2014-01-01

    Background Molecular biomarkers including isocitrate dehydrogenase 1 or 2 (IDH1/2) mutation, 1p/19q codeletion, and O6-methylguanine-DNA-methyltransferase (MGMT) promoter methylation may improve prognostication and guide treatment decisions for patients with World Health Organization (WHO) anaplastic gliomas. At present, each marker is individually tested by distinct assays. Illumina Infinium HumanMethylation450 BeadChip arrays (HM450) enable the determination of large-scale methylation profiles and genome-wide DNA copy number changes. Algorithms have been developed to detect the glioma CpG island methylator phenotype (G-CIMP) associated with IDH1/2 mutation, 1p/19q codeletion, and MGMT promoter methylation using a single assay. Methods Here, we retrospectively investigated the diagnostic and prognostic performance of these algorithms in comparison to individual marker testing and patient outcome in the biomarker cohort (n = 115 patients) of the NOA-04 trial. Results Concordance for IDH and 1p/19q status was very high: In 92% of samples, the HM450 and reference data agreed. In discordant samples, survival analysis by Kaplan-Meier and Cox regression analyses suggested a more accurate assessment of biological phenotype by the HM450 analysis. The HM450-derived MGMT-STP27 model to calculate MGMT promoter methylation probability revealed this aberration in a significantly higher fraction of samples than conventional methylation-specific PCR, with 87 of 91 G-CIMP tumors predicted as MGMT promoter-methylated. Pyrosequencing of discordant samples confirmed the HM450 assessment in 14 of 17 cases. Conclusions G-CIMP and 1p/19q codeletion are reliably detectable by HM450 analysis and are associated with prognosis in the NOA-04 trial. For MGMT, HM450 suggests promoter methylation in the vast majority of G-CIMP tumors, which is supported by pyrosequencing. PMID:25028501

  16. Loss-of-heterozygosity on chromosome 19q in early-stage serous ovarian cancer is associated with recurrent disease

    PubMed Central

    2012-01-01

    Background Ovarian cancer is a heterogeneous disease and prognosis for apparently similar cases of ovarian cancer varies. Recurrence of the disease in early stage (FIGO-stages I-II) serous ovarian cancer results in survival that is comparable to those with recurrent advanced-stage disease. The aim of this study was to investigate if there are specific genomic aberrations that may explain recurrence and clinical outcome. Methods Fifty-one women with early stage serous ovarian cancer were included in the study. DNA was extracted from formalin fixed samples containing tumor cells from ovarian tumors. Tumor samples from thirty-seven patients were analysed for allele-specific copy numbers using OncoScan single nucleotide polymorphism arrays from Affymetrix and the bioinformatic tool Tumor Aberration Prediction Suite. Genomic gains, losses, and loss-of-heterozygosity that associated with recurrent disease were identified. Results The most significant differences (p < 0.01) in Loss-of-heterozygosity (LOH) were identified in two relatively small regions of chromosome 19; 8.0-8,8 Mbp (19 genes) and 51.5-53.0 Mbp (37 genes). Thus, 56 genes on chromosome 19 were potential candidate genes associated with clinical outcome. LOH at 19q (51-56 Mbp) was associated with shorter disease-free survival and was an independent prognostic factor for survival in a multivariate Cox regression analysis. In particular LOH on chromosome 19q (51-56 Mbp) was significantly (p < 0.01) associated with loss of TP53 function. Conclusions The results of our study indicate that presence of two aberrations in TP53 on 17p and LOH on 19q in early stage serous ovarian cancer is associated with recurrent disease. Further studies related to the findings of chromosomes 17 and 19 are needed to elucidate the molecular mechanism behind the recurring genomic aberrations and the poor clinical outcome. PMID:22967087

  17. Alterations of the RRAS and ERCC1 genes at 19q13 in gemistocytic astrocytomas.

    PubMed

    Ohta, Takashi; Kim, Young-Ho; Oh, Ji-Eun; Satomi, Kaishi; Nonoguchi, Naosuke; Keyvani, Kathy; Pierscianek, Daniela; Sure, Ulrich; Mittelbronn, Michel; Paulus, Werner; Vital, Anne; Yokoo, Hideaki; McDonald, Kerrie; Kleihues, Paul; Nazaret, Nicolas; Barbet, Fabienne; Lachuer, Joel; Ohgaki, Hiroko

    2014-10-01

    Gemistocytic astrocytoma (World Health Organization grade II) is a rare variant of diffuse astrocytoma that is characterized by the presence of neoplastic gemistocytes and has a significantly less favorable prognosis. Other than frequent TP53 mutations (>80%), little is known about its molecular profile. Here, we show that gemistocytic astrocytomas carry a lower frequency of IDH mutations than fibrillary astrocytomas (74% vs 92%; p = 0.0255) but have profiles similar to those of fibrillary astrocytomas with respect to TERT promoter mutations (5% vs 0%), 1p/19q loss (10% vs 8%), and loss of heterozygosity 10q (10% vs 12%). Exome sequencing in 5 gemistocytic astrocytomas revealed homozygous deletion of genes at 19q13 (i.e. RRAS [related RAS viral oncogene homolog; 2 cases] and ERCC1 [excision repair cross-complementing rodent repair deficiency, complementation group 1; 1 case]). Further screening showed RRAS homozygous deletion in 7 of 42 (17%) gemistocytic astrocytomas and in 3 of 24 (13%) IDH1 mutated secondary glioblastomas. Patients with gemistocytic astrocytoma and secondary glioblastoma with an RRAS deletion tended to have shorter survival rates than those without deletion. Differential polymerase chain reaction and methylation-specific polymerase chain reaction revealed an ERCC1 homozygous deletion or promoter methylation in 10 of 42 (24%) gemistocytic astrocytomas and in 8 of 24 (33%) secondary glioblastomas. Alterations in RRAS and ERCC1 appear to be typical in gemistocytic astrocytomas and secondary glioblastomas, since they were not present in 49 fibrillary astrocytomas or 30 primary glioblastomas.

  18. Life Origination Hydrate Hypothesis (LOH-Hypothesis)

    PubMed Central

    Ostrovskii, Victor; Kadyshevich, Elena

    2012-01-01

    The paper develops the Life Origination Hydrate Hypothesis (LOH-hypothesis), according to which living-matter simplest elements (LMSEs, which are N-bases, riboses, nucleosides, nucleotides), DNA- and RNA-like molecules, amino-acids, and proto-cells repeatedly originated on the basis of thermodynamically controlled, natural, and inevitable processes governed by universal physical and chemical laws from CH4, niters, and phosphates under the Earth's surface or seabed within the crystal cavities of the honeycomb methane-hydrate structure at low temperatures; the chemical processes passed slowly through all successive chemical steps in the direction that is determined by a gradual decrease in the Gibbs free energy of reacting systems. The hypothesis formulation method is based on the thermodynamic directedness of natural movement and consists ofan attempt to mentally backtrack on the progression of nature and thus reveal principal milestones alongits route. The changes in Gibbs free energy are estimated for different steps of the living-matter origination process; special attention is paid to the processes of proto-cell formation. Just the occurrence of the gas-hydrate periodic honeycomb matrix filled with LMSEs almost completely in its final state accounts for size limitation in the DNA functional groups and the nonrandom location of N-bases in the DNA chains. The slowness of the low-temperature chemical transformations and their “thermodynamic front” guide the gross process of living matter origination and its successive steps. It is shown that the hypothesis is thermodynamically justified and testable and that many observed natural phenomena count in its favor. PMID:25382120

  19. Detailed comparative map of human chromosome 19q and related regions of the mouse genome.

    PubMed

    Stubbs, L; Carver, E A; Shannon, M E; Kim, J; Geisler, J; Generoso, E E; Stanford, B G; Dunn, W C; Mohrenweiser, H; Zimmermann, W; Watt, S M; Ashworth, L K

    1996-08-01

    One of the larger contiguous blocks of mouse-human genomic homology includes the proximal portion of mouse chromosome 7 and the long arm of human chromosome 19. Previous studies have demonstrated the close relationship between the two regions, but have also indicated significant rearrangements in the relative orders of homologous mouse and human genes. Here we present the genetic locations of the homologs of 42 human chromosome 19q markers in the mouse, with an emphasis on genes also included in the human chromosome 19 physical map. Our results demonstrate that despite an overall inversion of sequences relative to the centromere, apparent "transpositions" of three gene-rich segments, and a local inversion of markers mapping near the 19q telomere, gene content, order, and spacing are remarkably well conserved throughout the lengths of these related mouse and human regions. Although most human 19q markers have remained genetically linked in mouse, one small human segment forms a separate region of homology between human chromosome 19q and mouse chromosome 17. Three of the four rearrangements of mouse versus human 19q sequences involve segments that are located directly adjacent to each other in 19q13.3-q13.4, suggesting either the coincident occurrence of these events or their common association with unstable DNA sequences. These data permit an unusually in-depth examination of this large region of mouse-human genomic homology and provide an important new tool to aid in the mapping of genes and associated phenotypes in both species.

  20. Functional characterization of the 19q12 amplicon in grade III breast cancers

    PubMed Central

    2012-01-01

    Introduction The 19q12 locus is amplified in a subgroup of oestrogen receptor (ER)-negative grade III breast cancers. This amplicon comprises nine genes, including cyclin E1 (CCNE1), which has been proposed as its 'driver'. The aim of this study was to identify the genes within the 19q12 amplicon whose expression is required for the survival of cancer cells harbouring their amplification. Methods We investigated the presence of 19q12 amplification in a series of 313 frozen primary breast cancers and 56 breast cancer cell lines using microarray comparative genomic hybridisation (aCGH). The nine genes mapping to the smallest region of amplification on 19q12 were silenced using RNA interference in phenotypically matched breast cancer cell lines with (MDA-MB-157 and HCC1569) and without (Hs578T, MCF7, MDA-MB-231, ZR75.1, JIMT1 and BT474) amplification of this locus. Genes whose silencing was selectively lethal in amplified cells were taken forward for further validation. The effects of cyclin-dependent kinase 2 (CDK2) silencing and chemical inhibition were tested in cancer cells with and without CCNE1 amplification. Results 19q12 amplification was identified in 7.8% of ER-negative grade III breast cancer. Of the nine genes mapping to this amplicon, UQCRFS1, POP4, PLEKHF1, C19ORF12, CCNE1 and C19ORF2 were significantly over-expressed when amplified in primary breast cancers and/or breast cancer cell lines. Silencing of POP4, PLEKHF1, CCNE1 and TSZH3 selectively reduced cell viability in cancer cells harbouring their amplification. Cancer cells with CCNE1 amplification were shown to be dependent on CDK2 expression and kinase activity for their survival. Conclusions The 19q12 amplicon may harbour more than a single 'driver', given that expression of POP4, PLEKHF1, CCNE1 and TSZH3 is required for the survival of cancer cells displaying their amplification. The observation that cancer cells harbouring CCNE1 gene amplification are sensitive to CDK2 inhibitors provides a

  1. A genome-wide association study of COPD identifies a susceptibility locus on chromosome 19q13.

    PubMed

    Cho, Michael H; Castaldi, Peter J; Wan, Emily S; Siedlinski, Mateusz; Hersh, Craig P; Demeo, Dawn L; Himes, Blanca E; Sylvia, Jody S; Klanderman, Barbara J; Ziniti, John P; Lange, Christoph; Litonjua, Augusto A; Sparrow, David; Regan, Elizabeth A; Make, Barry J; Hokanson, John E; Murray, Tanda; Hetmanski, Jacqueline B; Pillai, Sreekumar G; Kong, Xiangyang; Anderson, Wayne H; Tal-Singer, Ruth; Lomas, David A; Coxson, Harvey O; Edwards, Lisa D; MacNee, William; Vestbo, Jørgen; Yates, Julie C; Agusti, Alvar; Calverley, Peter M A; Celli, Bartolome; Crim, Courtney; Rennard, Stephen; Wouters, Emiel; Bakke, Per; Gulsvik, Amund; Crapo, James D; Beaty, Terri H; Silverman, Edwin K

    2012-02-15

    The genetic risk factors for chronic obstructive pulmonary disease (COPD) are still largely unknown. To date, genome-wide association studies (GWASs) of limited size have identified several novel risk loci for COPD at CHRNA3/CHRNA5/IREB2, HHIP and FAM13A; additional loci may be identified through larger studies. We performed a GWAS using a total of 3499 cases and 1922 control subjects from four cohorts: the Evaluation of COPD Longitudinally to Identify Predictive Surrogate Endpoints (ECLIPSE); the Normative Aging Study (NAS) and National Emphysema Treatment Trial (NETT); Bergen, Norway (GenKOLS); and the COPDGene study. Genotyping was performed on Illumina platforms with additional markers imputed using 1000 Genomes data; results were summarized using fixed-effect meta-analysis. We identified a new genome-wide significant locus on chromosome 19q13 (rs7937, OR = 0.74, P = 2.9 × 10(-9)). Genotyping this single nucleotide polymorphism (SNP) and another nearby SNP in linkage disequilibrium (rs2604894) in 2859 subjects from the family-based International COPD Genetics Network study (ICGN) demonstrated supportive evidence for association for COPD (P = 0.28 and 0.11 for rs7937 and rs2604894), pre-bronchodilator FEV(1) (P = 0.08 and 0.04) and severe (GOLD 3&4) COPD (P = 0.09 and 0.017). This region includes RAB4B, EGLN2, MIA and CYP2A6, and has previously been identified in association with cigarette smoking behavior.

  2. A region of consistent deletion in neuroblastoma maps within human chromosome 1p36.2-36.3

    SciTech Connect

    White, P.S.; Maris, J.M.; Beltinger, C.

    1995-06-06

    Deletion of the short arm of human chromosome 1 is the most common cytogenetic abnormality observed in neuroblastoma. To characterize the region of consistent deletion, we performed loss of heterozygosity (LOH) studies on 122 neuroblastoma tumor samples with 30 distal chromosome 1p polymorphisms. LOH was detected in 32 of the 122 tumors (26%). A single region of LOH, marked distally by D1Z2 and proximally by D1S228, was detected in all tumors demonstrating loss. Also, cells from a patient with a constitutional deletion of 1p36, and from a neuroblastoma cell line with a small 1p36 deletion, were analyzed by fluorescence in situ hybridization. Cells from both sources had interstitial deletions of 1p36.2-36.3 which overlapped the consensus region of LOH defined by the tumors. Interstitial deletion in the constitutional case was confirmed by allelic loss studies using the panel of polymorphic markers. Four proposed candidate genes-DAN, ID3 (heir-1), CDC2L1 (p58), and TNFR2-were shown to lie outside of the consensus region of allelic loss, as defined by the above deletions. These results more precisely define the location of a neuroblastoma suppressor gene within 1p36.2-36.3, eliminating 33 centimorgans of proximal 1p36 from consideration. Furthermore, a consensus region of loss, which excludes the four leading candidate genes, was found in all tumors with 1p36 LOH. 31 refs., 4 figs.

  3. The 19q12 bladder cancer GWAS signal: association with cyclin E function and aggressive disease

    PubMed Central

    Fu, Yi-Ping; Kohaar, Indu; Moore, Lee E.; Lenz, Petra; Figueroa, Jonine D.; Tang, Wei; Porter-Gill, Patricia; Chatterjee, Nilanjan; Scott-Johnson, Alexandra; Garcia-Closas, Montserrat; Muchmore, Brian; Baris, Dalsu; Paquin, Ashley; Ylaya, Kris; Schwenn, Molly; Apolo, Andrea B.; Karagas, Margaret R.; Tarway, McAnthony; Johnson, Alison; Mumy, Adam; Schned, Alan; Guedez, Liliana; Jones, Michael A.; Kida, Masatoshi; Monawar Hosain, GM; Malats, Nuria; Kogevinas, Manolis; Tardon, Adonina; Serra, Consol; Carrato, Alfredo; Garcia-Closas, Reina; Lloreta, Josep; Wu, Xifeng; Purdue, Mark; Andriole, Gerald L.; Grubb, Robert L.; Black, Amanda; Landi, Maria T.; Caporaso, Neil E.; Vineis, Paolo; Siddiq, Afshan; Bueno-de-Mesquita, H. Bas; Trichopoulos, Dimitrios; Ljungberg, Börje; Severi, Gianluca; Weiderpass, Elisabete; Krogh, Vittorio; Dorronsoro, Miren; Travis, Ruth C.; Tjønneland, Anne; Brennan, Paul; Chang-Claude, Jenny; Riboli, Elio; Prescott, Jennifer; Chen, Constance; De Vivo, Immaculata; Govannucci, Edward; Hunter, David; Kraft, Peter; Lindstrom, Sara; Gapstur, Susan M.; Jacobs, Eric J.; Diver, W. Ryan; Albanes, Demetrius; Weinstein, Stephanie J.; Virtamo, Jarmo; Kooperberg, Charles; Hohensee, Chancellor; Rodabough, Rebecca J.; Cortessis, Victoria K.; Conti, David V.; Gago-Dominguez, Manuela; Stern, Mariana C.; Pike, Malcolm C.; Van Den Berg, David; Yuan, Jian-Min; Haiman, Christopher A.; Cussenot, Olivier; Cancel-Tassin, Geraldine; Roupret, Morgan; Comperat, Eva; Porru, Stefano; Carta, Angela; Pavanello, Sofia; Arici, Cecilia; Mastrangelo, Giuseppe; Grossman, H. Barton; Wang, Zhaoming; Deng, Xiang; Chung, Charles C.; Hutchinson, Amy; Burdette, Laurie; Wheeler, William; Fraumeni, Joseph; Chanock, Stephen J.; Hewitt, Stephen M.; Silverman, Debra T.; Rothman, Nathaniel; Prokunina-Olsson, Ludmila

    2014-01-01

    A genome-wide association study (GWAS) of bladder cancer identified a genetic marker rs8102137 within the 19q12 region as a novel susceptibility variant. This marker is located upstream of the CCNE1 gene, which encodes cyclin E, a cell cycle protein. We performed genetic fine mapping analysis of the CCNE1 region using data from two bladder cancer GWAS (5,942 cases and 10,857 controls). We found that the original GWAS marker rs8102137 represents a group of 47 linked SNPs (with r2≥0.7) associated with increased bladder cancer risk. From this group we selected a functional promoter variant rs7257330, which showed strong allele-specific binding of nuclear proteins in several cell lines. In both GWAS, rs7257330 was associated only with aggressive bladder cancer, with a combined per-allele odds ratio (OR) =1.18 (95%CI=1.09-1.27, p=4.67×10−5 vs. OR =1.01 (95%CI=0.93-1.10, p=0.79) for non-aggressive disease, with p=0.0015 for case-only analysis. Cyclin E protein expression analyzed in 265 bladder tumors was increased in aggressive tumors (p=0.013) and, independently, with each rs7257330-A risk allele (ptrend=0.024). Over-expression of recombinant cyclin E in cell lines caused significant acceleration of cell cycle. In conclusion, we defined the 19q12 signal as the first GWAS signal specific for aggressive bladder cancer. Molecular mechanisms of this genetic association may be related to cyclin E over-expression and alteration of cell cycle in carriers of CCNE1 risk variants. In combination with established bladder cancer risk factors and other somatic and germline genetic markers, the CCNE1 variants could be useful for inclusion into bladder cancer risk prediction models. PMID:25320178

  4. A genome-wide association study of COPD identifies a susceptibility locus on chromosome 19q13

    PubMed Central

    Cho, Michael H.; Castaldi, Peter J.; Wan, Emily S.; Siedlinski, Mateusz; Hersh, Craig P.; Demeo, Dawn L.; Himes, Blanca E.; Sylvia, Jody S.; Klanderman, Barbara J.; Ziniti, John P.; Lange, Christoph; Litonjua, Augusto A.; Sparrow, David; Regan, Elizabeth A.; Make, Barry J.; Hokanson, John E.; Murray, Tanda; Hetmanski, Jacqueline B.; Pillai, Sreekumar G.; Kong, Xiangyang; Anderson, Wayne H.; Tal-Singer, Ruth; Lomas, David A.; Coxson, Harvey O.; Edwards, Lisa D.; MacNee, William; Vestbo, Jørgen; Yates, Julie C.; Agusti, Alvar; Calverley, Peter M.A.; Celli, Bartolome; Crim, Courtney; Rennard, Stephen; Wouters, Emiel; Bakke, Per; Gulsvik, Amund; Crapo, James D.; Beaty, Terri H.; Silverman, Edwin K.

    2012-01-01

    The genetic risk factors for chronic obstructive pulmonary disease (COPD) are still largely unknown. To date, genome-wide association studies (GWASs) of limited size have identified several novel risk loci for COPD at CHRNA3/CHRNA5/IREB2, HHIP and FAM13A; additional loci may be identified through larger studies. We performed a GWAS using a total of 3499 cases and 1922 control subjects from four cohorts: the Evaluation of COPD Longitudinally to Identify Predictive Surrogate Endpoints (ECLIPSE); the Normative Aging Study (NAS) and National Emphysema Treatment Trial (NETT); Bergen, Norway (GenKOLS); and the COPDGene study. Genotyping was performed on Illumina platforms with additional markers imputed using 1000 Genomes data; results were summarized using fixed-effect meta-analysis. We identified a new genome-wide significant locus on chromosome 19q13 (rs7937, OR = 0.74, P = 2.9 × 10−9). Genotyping this single nucleotide polymorphism (SNP) and another nearby SNP in linkage disequilibrium (rs2604894) in 2859 subjects from the family-based International COPD Genetics Network study (ICGN) demonstrated supportive evidence for association for COPD (P = 0.28 and 0.11 for rs7937 and rs2604894), pre-bronchodilator FEV1 (P = 0.08 and 0.04) and severe (GOLD 3&4) COPD (P = 0.09 and 0.017). This region includes RAB4B, EGLN2, MIA and CYP2A6, and has previously been identified in association with cigarette smoking behavior. PMID:22080838

  5. Assessment of the frequency of genetic alterations (LOH/MSI) in patients with intraepithelial cervical lesions with HPV infection: a pilot study.

    PubMed

    Migdalska-Sęk, Monika; Karowicz-Bilińska, Agata; Pastuszak-Lewandoska, Dorota; Czarnecka, Karolina H; Nawrot, Ewa; Domańska-Senderowska, Daria; Kiszałkiewicz, Justyna; Brzeziańska-Lasota, Ewa

    2016-05-01

    In the present study, we analyzed (1) the type of HPV infection and (2) the frequency of loss of heterozygosity and microsatellite imbalance (LOH/MSI) in normal cytology and cervical intraepithelial neoplasia (CIN1-3). The cytological material included: low-grade squamous intraepithelial lesions (CIN1, n = 11), high-grade lesions (CIN2 and CIN3, n = 13), and cytologically normal cells from non-neoplastic cervical samples (n = 8). HPV genotyping was done using RealLine HPV 16/18 kit. We used 20 microsatellite markers from: 1p31.2, 3p14.3, 3p21.3, 3p22.2, 3p24.2, 3p25.3, 7q32.2, 9p21.3, 11p15.5, 12q23.2, and 16q22.1. LOH/MSI was correlated with clinicopathological parameters. The presence of HPV DNA was revealed in 78.13 % samples, including normal cytology. LOH/MSI was the most frequent for: 3p25.3 (39 %), 3p22.2 (20.83 %), 3p24.2 (20 %), and 3p14.3 (16.67 %). It was demonstrated that D3S1234 (FHIT; 3p14.3), D3S1611 (MLH1; 3p22.2), D3S1583 (RARB; 3p24.2), D3S1317 and D3S3611 (VHL; 3p25.3) could differentiate patients with CIN2/CIN3 versus CIN1, showing significantly higher frequency in CIN2/CIN3. LOH/MSI frequency for other than 3p markers was lower, 10-22.2 %. The simultaneous occurrence of LOH/MSI for several markers (OFAL) was higher in CIN2/CIN3. Significant differences in OFAL were found between samples with versus without HPV infection. In HPV-positive patients, significant differences in OFAL were found between normal cytology, CIN1 and CIN2/CIN3. HPV infection influences the increase in LOH/MSI frequency, especially in tumor suppressor gene loci. Several studied microsatellite markers seem to be useful for CIN grading. Hopefully, the obtained results, if confirmed on larger patient cohort, would allow creating a panel of markers supporting clinical diagnosis in patients with HPV infection.

  6. Identifying Human Genome-Wide CNV, LOH and UPD by Targeted Sequencing of Selected Regions.

    PubMed

    Wang, Yu; Li, Wei; Xia, Yingying; Wang, Chongzhi; Tang, Y Tom; Guo, Wenying; Li, Jinliang; Zhao, Xia; Sun, Yepeng; Hu, Juan; Zhen, Hefu; Zhang, Xiandong; Chen, Chao; Shi, Yujian; Li, Lin; Cao, Hongzhi; Du, Hongli; Li, Jian

    2014-01-01

    Copy-number variations (CNV), loss of heterozygosity (LOH), and uniparental disomy (UPD) are large genomic aberrations leading to many common inherited diseases, cancers, and other complex diseases. An integrated tool to identify these aberrations is essential in understanding diseases and in designing clinical interventions. Previous discovery methods based on whole-genome sequencing (WGS) require very high depth of coverage on the whole genome scale, and are cost-wise inefficient. Another approach, whole exome genome sequencing (WEGS), is limited to discovering variations within exons. Thus, we are lacking efficient methods to detect genomic aberrations on the whole genome scale using next-generation sequencing technology. Here we present a method to identify genome-wide CNV, LOH and UPD for the human genome via selectively sequencing a small portion of genome termed Selected Target Regions (SeTRs). In our experiments, the SeTRs are covered by 99.73%~99.95% with sufficient depth. Our developed bioinformatics pipeline calls genome-wide CNVs with high confidence, revealing 8 credible events of LOH and 3 UPD events larger than 5M from 15 individual samples. We demonstrate that genome-wide CNV, LOH and UPD can be detected using a cost-effective SeTRs sequencing approach, and that LOH and UPD can be identified using just a sample grouping technique, without using a matched sample or familial information.

  7. Identifying Human Genome-Wide CNV, LOH and UPD by Targeted Sequencing of Selected Regions

    PubMed Central

    Guo, Wenying; Li, Jinliang; Zhao, Xia; Sun, Yepeng; Hu, Juan; Zhen, Hefu; Zhang, Xiandong; Chen, Chao; Shi, Yujian; Li, Lin; Cao, Hongzhi; Du, Hongli; Li, Jian

    2015-01-01

    Copy-number variations (CNV), loss of heterozygosity (LOH), and uniparental disomy (UPD) are large genomic aberrations leading to many common inherited diseases, cancers, and other complex diseases. An integrated tool to identify these aberrations is essential in understanding diseases and in designing clinical interventions. Previous discovery methods based on whole-genome sequencing (WGS) require very high depth of coverage on the whole genome scale, and are cost-wise inefficient. Another approach, whole exome genome sequencing (WEGS), is limited to discovering variations within exons. Thus, we are lacking efficient methods to detect genomic aberrations on the whole genome scale using next-generation sequencing technology. Here we present a method to identify genome-wide CNV, LOH and UPD for the human genome via selectively sequencing a small portion of genome termed Selected Target Regions (SeTRs). In our experiments, the SeTRs are covered by 99.73%~99.95% with sufficient depth. Our developed bioinformatics pipeline calls genome-wide CNVs with high confidence, revealing 8 credible events of LOH and 3 UPD events larger than 5M from 15 individual samples. We demonstrate that genome-wide CNV, LOH and UPD can be detected using a cost-effective SeTRs sequencing approach, and that LOH and UPD can be identified using just a sample grouping technique, without using a matched sample or familial information. PMID:25919136

  8. The human glia maturation factor-gamma gene: genomic structure and mutation analysis in gliomas with chromosome 19q loss.

    PubMed

    Peters, N; Smith, J S; Tachibana, I; Lee, H K; Pohl, U; Portier, B P; Louis, D N; Jenkins, R B

    1999-09-01

    Human glia maturation factor-gamma (hGMF-gamma) is a recently identified gene that may be involved in glial differentiation, neural regeneration, and inhibition of tumor cell proliferation. The gene maps to the long arm of chromosome 19 at band q13.2, a region that is frequently deleted in human malignant gliomas and is thus suspected to harbor a glioma tumor suppressor gene. Given the putative role of hGMF-gamma in cell differentiation and proliferation and its localization to chromosome 19q13, this gene is an interesting candidate for the chromosome 19q glioma tumor suppressor gene. To evaluate this possibility, we determined the genomic structure of human hGMF-gamma and performed mutation screening in a series of 41 gliomas with and without allelic loss of chromosome 19q. Mutations were not detected, which suggests that hGMF-gamma is not the chromosome 19q glioma suppressor gene. However, the elucidation of the genomic structure of hGMF-gamma may prove useful in future investigations of hGMF-gamma in the normal adult and developing human nervous system.

  9. Simultaneous de novo interstitial deletion of 16q21 and intercalary duplication of 19q in a retarded infant with minor dysmorphic features.

    PubMed Central

    Trautmann, U; Pfeiffer, R A; Seufert-Satomi, U; Tietze, H U

    1993-01-01

    We report on a retarded infant with minor dysmorphic features in whom deletion 16 and duplication 19q were discovered. The karyotype is 46,XX,del(16) (q13.08-21.05),dup(19)(q13.13-13.2). The origin and significance of the aberrant chromosomes are unknown. Images PMID:8487285

  10. Genomic Instability and Copy-Number Heterogeneity of Chromosome 19q, Including the Kallikrein Locus, in Ovarian Carcinomas

    PubMed Central

    Bayani, Jane; Marrano, Paula; Graham, Cassandra; Zheng, Yingye; Li, Lin; Katsaros, Dionyssios; Lassus, Heini; Butzow, Ralf; Squire, Jeremy A.; Diamandis, Eleftherios P.

    2011-01-01

    Many tissue kallikrein (KLK) genes and proteins are candidate diagnostic, prognostic and predictive biomarkers for ovarian cancer (OCa). We previously demonstrated that the KLK locus (19q13.3/4) is subject to copy-number gains and structural rearrangements in a pilot study of cell lines and ovarian cancer primary tissues, shown to overexpress KLK gene family members. To determine the overall frequency of genomic instability and copy-number changes, a retrospective study was conducted using formalin-fixed paraffin embedded (FFPE) tissues. Eighty-one chemotherapy naïve serous OCas were examined using 3-colour fluorescence in situ hybridization (FISH) to identify structural and numerical changes on 19q, including the KLK locus; in addition to immunohistochemistry (IHC) for KLK6, which has been shown to be overexpressed in OCa. The KLK locus was subject to copy-number changes in ~83% of cases: net gain in 51%, net loss in 30% and amplified in 2%; and found to be chromosomally unstable (p<0.001). All cases showed a wide range of immuoreactivity for KLK6 by IHC. Although no strong correlation could be found with copy number, the latter was contributing factor to the observed KLK6 protein overexpression. Moreover, univariate and multivariate analyses showed an association between the net loss of the KLK locus with longer disease-free survival. Interestingly, FISH analyses indicated that chromosome 19q was subject to structural rearrangement in 62% of cases and was significantly correlated to tumor grade (p<0.001). We conclude that numerical and structural aberrations of chromosome 19q, affect genes including the KLK gene members, may contributing to ovarian carcinoma progression and aggressiveness. PMID:20800559

  11. Genomic alterations in cervical carcinoma: Losses of chromosome heterozygosity (LOH) correlated with cytogenetic, HPV, and p53 evaluations

    SciTech Connect

    Klinger, H.P.; Mullokandov, M.; Khollodilov, N.G.

    1994-09-01

    This study was undertaken to obtain indications of chromosomes likely to carry tumorigenicity suppressor genes the loss of function of which play a role in the origin or progression of cervical carcinomas. PCR and electrophoresis with primers for 73 highly polymorphic microsatellite chromosome markers were used to determine the incidence of LOH of all of the autosomes in 38 cervical carcinomas. According to these criteria 14 of the autosomes are involved in LOH in 24% to 42% of the tumors. This involves chromosomes 1, 2, 3, 4, 5, 6, 7, 8, 9, 11, 13, 16, 18 and 19. Most frequently involved are chromosomes 3 and 6 with LOH in 42% of the tumors. The chromosomes next most frequently involved are 4, 7, 11 and 18, with LOH in 31-32% of cases. Chromosomes 1, 2, 5, 8 and 16 each had LOH in 29% of the tumors; 9 and 13 each in 26%; and 19 in 24% of the tumors. All other autosomes had LOH in 18% or fewer of the tumors. Cytogenetic analyses performed on direct preparations from many of the same tumors agreed well with the molecular LOH assays. Correlation of the information obtained with both of these methods provides considerable insight into the mechanisms involved in the occurrence of these chromosome alterations. Chromosome 3 is the third most frequent chromosome involved in LOH in all types of cancer. In cervical carcinomas the region most frequently involved is 3p13-p25, which is a segment within which suppressors have been implicated in several other types of malignancies. Chromosome 6 on the other hand is rarely involved in other neoplasias and this appears to be unique to cervical carcinomas. Of interest was the finding that many of the HPV-negative tumors had LOH of chromosome 17 and many of these expressed mutant p53. The latter tumors occur in older women and are on the average much more aggressive than the HPV-positive tumors.

  12. Contribution of retinoblastoma LOH and the p53 Arg/Pro polymorphism to cervical cancer.

    PubMed

    Eltahir, Huda A; Elhassan, Ahmed M; Ibrahim, Muntaser E

    2012-09-01

    Epidemiological studies indicate that infections by certain types of human papillomaviruses (HPVs) are causally linked to the development of cervical cancer. It is also known that HPV infections alone do not cause progression to cervical cancer, as additional genetic changes such as loss of distinct chromosomal regions, inactivation of tumor-suppressor genes and activation of oncogenes must also occur in order for malignant transformation to take place. In the present study, 78 patients diagnosed with cervical cancer and 36 cervical cancer-free cases (control) were analyzed for high-risk HPV genotypes (16 and 18) by polymerase chain reaction (PCR). Loss of heterozygosity (LOH) of the retinoblastoma gene (Rb) at two polymorphic intronic sites (intron 1 and 17) and the p53 polymorphism in codon 72 were detected by RFLP and allele-specific PCR, respectively. HPV 16 and 18 were found at frequencies of 93.6 and 8.3% in the cervical cancer and control samples, respectively. LOH was detected in 63% of patients in intron 1 and/or intron 17. p53 allele frequency for Arg/Arg was 43.6% (34/78), for Arg/Pro 37.2% (29/78) and for Pro/Pro 19.2% (15/78). The relative risk (RR) of LOH and Arg/Arg alone was 1.7 and 1.1, respectively, while the combined RR for Rb LOH and p53 Arg/Arg was 2.5. The present study showed a significant association of the chromosomal allelic loss of Rb in Sudanese cervical cancer patients, while no such association was observed with other parameters, such as clinical stage and degree of differentiation; hence, it cannot be a determinant of tumor behavior in cervical carcinoma. Although the p53 arginine allele is itself an important risk factor for cervical cancer, the combined risk with LOH of Rb, which appears to be greater, might indicate a possible epistatic effect of the two genes/polymorphisms.

  13. Phytophthora capsici - Loss of Heterozygosity (LOH): A Widespread Mechanism for Rapid Adaptation ( 7th Annual SFAF Meeting, 2012)

    ScienceCinema

    Mudge, Joanne [NCGR

    2016-07-12

    Joanne Mudge on "Phytophthora capsici - Loss of Heterozygosity (LOH): A Widespread Mechanism for Rapid Mutation" at the 2012 Sequencing, Finishing, Analysis in the Future Meeting held June 5-7, 2012 in Santa Fe, New Mexico.

  14. Comparative analysis of a conserved zinc finger gene cluster on human chromosome 19q and mouse chromosome 7.

    PubMed

    Shannon, M; Ashworth, L K; Mucenski, M L; Lamerdin, J E; Branscomb, E; Stubbs, L

    1996-04-01

    Several lines of evidence now suggest that many of the zinc-finger-containing (ZNF) genes in the human genome are arranged in clusters. However, little is known about the structure or function of the clusters or about their conservation throughout evolution. Here, we report the analysis of a conserved ZNF gene cluster located in human chromosome 19q13.2 and mouse chromosome 7. Our results indicate that the human cluster consists of at least 10 related Kruppel-associated box (KRAB)-containing ZNF genes organized in tandem over a distance of 350-450 kb. Two cDNA clones representing genes in the murine cluster have been studied in detail. The KRAB A domains of these genes are nearly identical and are highly similar to human 19q13.2-derived KRAB sequences, but DNA-binding ZNF domains and other portions of the genes differ considerably. The two murine genes display distinct expression patterns, but are coexpressed in some adult tissues. These studies pave the way for a systematic analysis of the evolution of structure and function of genes within the numerous clustered ZNF families located on human chromosome 19 and elsewhere in the human and mouse genomes.

  15. Localization, by linkage analysis, of the cystinuria type III gene to chromosome 19q13.1

    SciTech Connect

    Bisceglia, L.; Totaro, A.; Melchionda, S.

    1997-03-01

    Cystinuria is an autosomal recessive aminoaciduria in which three urinary phenotypes (I, II, and III) have been described. An amino acid transporter gene, SLC3A1 (formerly rBAT), was found to be responsible for this disorder. Mutational and linkage analysis demonstrated the presence of genetic heterogeneity in which the SLC3A1 gene is responsible for type I cystinuria but not for type II or type III. In this study, we report the identification of the cystinuria type III locus on the long arm of chromosome 19 (19q13.1), obtained after a genomewide search. Pairwise linkage analysis in a series of type III or type II families previously excluded from linkage to the cystinuria type I locus (SLC3A1 gene) revealed a significant maximum LOD score (Z{sub max}) of 13.11 at a maximum recombination fraction ({theta}{sub max}) of .00, with marker D19S225. Multipoint linkage analysis performed with the use of additional markers from the region placed the cystinuria type III locus between D19S414 and D19S220. Preliminary data on type II families also seem to place the disease locus for this rare type of cystinuria at 19q13.1 (significant Z{sub max} = 3.11 at {theta}{sub max} of .00, with marker D19S225). 33 refs., 2 figs., 1 tab.

  16. Life Origination Hydrate Theory (LOH-Theory) and the explanation of the biological diversification.

    PubMed

    Ostrovskii, Victor E; Kadyshevich, Elena A

    2014-12-01

    The Life Origination Hydrate Theory (LOH-Theory) considers the life origination process as a sequence of thermodynamically caused regular and inevitable chemical transformations regulated by universal physical and chemical laws. The LOH-Theory bears on a number of experimental, thermodynamic, observation, and simulation researches. N-bases, riboses, nucleosides, and nucleotides and DNAs and RNAs are formed repeatedly within structural cavities of localizations of underground and underseabed honeycomb CH4-hydrate deposits from CH4 and nitrate and phosphate ions that diffused into the hydrate structures; proto-cells and their agglomerates originated from these DNAs and from the same minerals in the semi-liquid soup after liquation of the hydrate structures. Each localization gave rise to a multitude of different DNAs and living organisms. The species diversity is caused by the spatial and temporal repeatability of the processes of living matter origination under similar but not identical conditions, multiplicity of the DNA forms in each living matter origination event, variations in the parameters of the native medium, intraspecific variations, and interspecific variations. The contribution of the last to the species diversity is, likely, significant for prokaryotes and those eukaryotes that are only at low steps of their biological organization; however, in the light of the LOH-Theory, of available long-term paleontological investigations, and of studies of reproduction of proliferous organisms, we conclude that, in toto, the contribution of interspecific variations to the species diversity was earlier overestimated by some researchers. The reason of this overestimation is that origination of scores of «spores» of different organisms in any one event and multiple reproductions of such events in time and Earth's space were not taken into consideration.

  17. An application of LOH analysis for detecting the genetic influences of space environmental radiation

    NASA Astrophysics Data System (ADS)

    Yatagai, F.; Umebayashi, Y.; Honma, M.; Abe, T.; Suzuki, H.; Shimazu, T.; Ishioka, N.; Iwaki, M.

    To detect the genetic influence of space environmental radiation at the chromosome level we proposed an application of loss of heterozygosity LOH analysis system for the mutations induced in human lymphoblastoid TK6 cells Surprisingly we succeeded the mutation detection in the frozen dells which were exposed to a low-dose 10 cGy of carbon-ion beam irradiation Mutation assays were performed within a few days or after about one month preservation at --80 r C following irradiation The results showed an increase in mutation frequency at the thymidine kinase TK gene locus 1 6-fold 2 5 X 10 -6 to 3 9 X 10 -6 and 2 1-fold 2 5 X 10 -6 to 5 3 X 10 -6 respectively Although the relative distributions of mutation classes were not changed by the radiation exposure in either assay an interesting characteristic was detected using this LOH analysis system two TK locus markers and eleven microsatellite loci spanning chromosome 17 The radiation-specific patterns of interstitial deletions were observed in the hemizygous LOH mutants which were considered as a result of end-joining repair of carbon ion-induced DNA double-strand breaks These results clearly demonstrate that this analysis can be used for the detection of low-dose ionizing radiation effects in the frozen cells In addition we performed so called adaptive response experiments in which TK6 cells were pre-irradiated with low-dose 2 5 sim 10 cGy of X-ray and then exposed to challenging dose 2Gy of X-rays Interestingly the

  18. Assignment of the human ZNF83 (HPF1) zinc finger gene to chromosome 19q13. 3-q13. 4

    SciTech Connect

    Marine, J.C.; Lecoq, P.J.; Poncelet, D.A.; Martial, J.A. ); Bellefroid, E.J.; Bourguignon, C. ); Riviere, M.; Szpirer, J.; Szpirer, C. )

    1994-05-01

    The authors have isolated a collection of human ZFPs encoding cDNAs (HPF1 to -9) by hybridization with a finger motif oligonucleotide probe. Here, they describe the localization of a chromosome 19-linked human ZFP gene (HPF1/ZNF83). They first assigned the ZNF83 gene on chromosome 19 by the screening of a human x rodent hybrid panel by DNA hybridization with a fragment of a previously cloned cDNA (data not shown). To further localize the gene within chromosome 19, the regional assignment of the ZNF83 gene was determined by fluorescence in situ hybridization and digital imaging microscopy as described elsewhere. Human metaphase spreads were hybridized with biotinylated ZNF83 cDNA, and hybridization was detected with fluorescein isothiocyanate-conjugated avidin-DCS. Chromosomes were identified by staining with 4,6-diamino-2-phenylindol dihydrochloride. The fractional length (Flpter) distance of the signal to the p arm terminus relative to the total chromosome length gave a Flpter value between 82.8 and 89.9, which is consistent with an assignment of the ZNF83 gene in ISCN region 19q13.3-q13.4. 14 refs., 1 fig.

  19. Assignment of the locus for PLO-SL, a frontal-lobe dementia with bone cysts, to 19q13.

    PubMed Central

    Pekkarinen, P; Hovatta, I; Hakola, P; Järvi, O; Kestilä, M; Lenkkeri, U; Adolfsson, R; Holmgren, G; Nylander, P O; Tranebjaerg, L; Terwilliger, J D; Lönnqvist, J; Peltonen, L

    1998-01-01

    PLO-SL (polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy) is a recessively inherited disorder characterized by systemic bone cysts and progressive presenile frontal-lobe dementia, resulting in death at <50 years of age. Since the 1960s, approximately 160 cases have been reported, mainly in Japan and Finland. The pathogenesis of the disease is unknown. In this article, we report the assignment of the locus for PLO-SL, by random genome screening using a modification of the haplotype-sharing method, in patients from a genetically isolated population. By screening five patient samples from 2 Finnish families, followed by linkage analysis of 12 Finnish families, 3 Swedish families, and 1 Norwegian family, we were able to assign the PLO-SL locus to a 9-cM interval between markers D19S191 and D19S420 on chromosome 19q13. The critical region was further restricted, to approximately 1.8 Mb, by linkage-disequilibrium analysis of the Finnish families. According to the haplotype analysis, one Swedish and one Norwegian PLO-SL family are not linked to the chromosome 19 locus, suggesting that PLO-SL is a heterogeneous disease. In this chromosomal region, one potential candidate gene for PLO-SL, the gene encoding amyloid precursor-like protein 1, was analyzed, but no mutations were detected in the coding region. PMID:9463329

  20. Assignment of the gastric inhibitory polypeptide receptor gene (GIPR) to chromosome bands 19q13.2-q13.3 by fluorescence in situ hybridization

    SciTech Connect

    Stoffel, M.; Fernald, A.A.; Bell, G.I.; Le Beau, M.M.

    1995-08-10

    The gastric inhibitory polypeptide receptor gene (GIPR) was localized, using fluorescence in situ hybridization (FISH), to human chromosome bands 19q13.2-q13.3. Gastric inhibitory polypeptide (GIP) is a potent stimulator of insulin secretion and mutations in the GIPR gene may be related to non-insulin-dependent diabetes mellitus (NIDDM). 13 refs., 1 fig.

  1. Chromosomal localization of the human natural killer cell class I receptor family genes to 19q13.4 by fluorescence in situ hybridization

    SciTech Connect

    Suto, Yumiko; Maenaka, Katsumi; yabe, Toshio

    1996-07-01

    This report describes the localization of the human natural killer cell I receptor family genes to human chromosome 19q13.4 using fluorescence in situ hybridization. These genes mediate the inhibition of the cytotoxicity of subsets of natural killer cells. 8 refs., 1 fig.

  2. Single Nucleotide Polymorphism (SNP)-Based Loss of Heterozygosity (LOH) Testing by Real Time PCR in Patients Suspect of Myeloproliferative Disease

    PubMed Central

    Huijsmans, Cornelis J. J.; Poodt, Jeroen; Damen, Jan; van der Linden, Johannes C.; Savelkoul, Paul H. M.; Pruijt, Johannes F. M.; Hilbink, Mirrian; Hermans, Mirjam H. A.

    2012-01-01

    During tumor development, loss of heterozygosity (LOH) often occurs. When LOH is preceded by an oncogene activating mutation, the mutant allele may be further potentiated if the wild-type allele is lost or inactivated. In myeloproliferative neoplasms (MPN) somatic acquisition of JAK2V617F may be followed by LOH resulting in loss of the wild type allele. The occurrence of LOH in MPN and other proliferative diseases may lead to a further potentiating the mutant allele and thereby increasing morbidity. A real time PCR based SNP profiling assay was developed and validated for LOH detection of the JAK2 region (JAK2LOH). Blood of a cohort of 12 JAK2V617F-positive patients (n = 6 25–50% and n = 6>50% JAK2V617F) and a cohort of 81 patients suspected of MPN was stored with EDTA and subsequently used for validation. To generate germ-line profiles, non-neoplastic formalin-fixed paraffin-embedded tissue from each patient was analyzed. Results of the SNP assay were compared to those of an established Short Tandem Repeat (STR) assay. Both assays revealed JAK2LOH in 1/6 patients with 25–50% JAK2V617F. In patients with >50% JAK2V617F, JAK2LOH was detected in 6/6 by the SNP assay and 5/6 patients by the STR assay. Of the 81 patients suspected of MPN, 18 patients carried JAK2V617F. Both the SNP and STR assay demonstrated the occurrence of JAK2LOH in 5 of them. In the 63 JAK2V617F-negative patients, no JAK2LOH was observed by SNP and STR analyses. The presented SNP assay reliably detects JAK2LOH and is a fast and easy to perform alternative for STR analyses. We therefore anticipate the SNP approach as a proof of principle for the development of LOH SNP-assays for other clinically relevant LOH loci. PMID:22768290

  3. HHV-8-unrelated primary effusion-like lymphoma associated with clonal loss of inherited chromosomally-integrated human herpesvirus-6A from the telomere of chromosome 19q.

    PubMed

    Zhang, Enjie; Cotton, Victoria E; Hidalgo-Bravo, Alberto; Huang, Yan; Bell, Adam J; Jarrett, Ruth F; Wilkie, Gavin S; Davison, Andrew J; Nacheva, Ellie P; Siebert, Reiner; Majid, Aneela; Kelpanides, Inga; Jayne, Sandrine; Dyer, Martin J; Royle, Nicola J

    2016-03-07

    Primary effusion lymphomas (PEL) are associated with human herpesvirus-8 (HHV-8) and usually occur in immunocompromised individuals. However, there are numerous reports of HHV-8-unrelated PEL-like lymphomas with unknown aetiology. Here we characterize an HHV-8-unrelated PEL-like lymphoma in an elderly woman who was negative for human immunodeficiency viruses 1 and 2, and hepatitis B and C. The woman was, however, a carrier of an inherited-chromosomally-integrated human herpesvirus-6A (iciHHV-6A) genome in one 19q telomere. The iciHHV-6A genome was complete in blood DNA, encoding a full set of protein-coding genes. Interestingly, the entire iciHHV-6A genome was absent from the HHV-8-unrelated-PEL-like lymphoma cells despite retention of both copies of chromosome 19. The somatic loss of the 19q-iciHHV-6A genome occurred very early during lymphoma development and we propose it occurred via telomere-loop formation and excision to release a circular viral genome that was subsequently lost. Whether release of the HHV-6A genome from the telomere contributed to lymphomagenesis, or was coincidental, remains unclear but this event may have deregulated the expression of HHV-6A or 19q genes or else disrupted telomere function. To establish the frequency and importance of iciHHV-6 loss from telomeres, the HHV-6 copy number should be assessed in tumours that arise in iciHHV-6 carriers.

  4. HHV-8-unrelated primary effusion-like lymphoma associated with clonal loss of inherited chromosomally-integrated human herpesvirus-6A from the telomere of chromosome 19q

    PubMed Central

    Zhang, Enjie; Cotton, Victoria E.; Hidalgo-Bravo, Alberto; Huang, Yan; J. Bell, Adam; F. Jarrett, Ruth; Wilkie, Gavin S.; Davison, Andrew J.; P. Nacheva, Ellie; Siebert, Reiner; Majid, Aneela; Kelpanides, Inga; Jayne, Sandrine; Dyer, Martin J.; Royle, Nicola J.

    2016-01-01

    Primary effusion lymphomas (PEL) are associated with human herpesvirus-8 (HHV-8) and usually occur in immunocompromised individuals. However, there are numerous reports of HHV-8-unrelated PEL-like lymphomas with unknown aetiology. Here we characterize an HHV-8-unrelated PEL-like lymphoma in an elderly woman who was negative for human immunodeficiency viruses 1 and 2, and hepatitis B and C. The woman was, however, a carrier of an inherited-chromosomally-integrated human herpesvirus-6A (iciHHV-6A) genome in one 19q telomere. The iciHHV-6A genome was complete in blood DNA, encoding a full set of protein-coding genes. Interestingly, the entire iciHHV-6A genome was absent from the HHV-8-unrelated-PEL-like lymphoma cells despite retention of both copies of chromosome 19. The somatic loss of the 19q-iciHHV-6A genome occurred very early during lymphoma development and we propose it occurred via telomere-loop formation and excision to release a circular viral genome that was subsequently lost. Whether release of the HHV-6A genome from the telomere contributed to lymphomagenesis, or was coincidental, remains unclear but this event may have deregulated the expression of HHV-6A or 19q genes or else disrupted telomere function. To establish the frequency and importance of iciHHV-6 loss from telomeres, the HHV-6 copy number should be assessed in tumours that arise in iciHHV-6 carriers. PMID:26947392

  5. Life origination and development hydrate theory (LOH-Theory) in the context of biological, physicochemical, astrophysical, and paleontological studies

    NASA Astrophysics Data System (ADS)

    Ostrovskii, V. E.; Kadyshevich, E. A.

    2014-04-01

    Till now, we formulated and developed the Life Origination Hydrate Theory (LOH-Theory) and Mitosis and Replication Hydrate Theory (MRHTheory) as the instruments for understanding the physical and chemical mechanisms applied by Nature for the living matter origination and propagation. This work is aimed at coordination of these theories with the paleontological and astrophysical knowledges and hypotheses of the Earth and Solar System remote histories.

  6. Life Origination Hydrate Theory (LOH-Theory) and Mitosis and Replication Hydrate Theory (MRH-Theory): three-dimensional PC validation

    NASA Astrophysics Data System (ADS)

    Kadyshevich, E. A.; Dzyabchenko, A. V.; Ostrovskii, V. E.

    2014-04-01

    Size compatibility of the CH4-hydrate structure II and multi-component DNA fragments is confirmed by three-dimensional simulation; it is validation of the Life Origination Hydrate Theory (LOH-Theory).

  7. High-resolution genome-wide analysis of irradiated (UV and γ-rays) diploid yeast cells reveals a high frequency of genomic loss of heterozygosity (LOH) events.

    PubMed

    St Charles, Jordan; Hazkani-Covo, Einat; Yin, Yi; Andersen, Sabrina L; Dietrich, Fred S; Greenwell, Patricia W; Malc, Ewa; Mieczkowski, Piotr; Petes, Thomas D

    2012-04-01

    In diploid eukaryotes, repair of double-stranded DNA breaks by homologous recombination often leads to loss of heterozygosity (LOH). Most previous studies of mitotic recombination in Saccharomyces cerevisiae have focused on a single chromosome or a single region of one chromosome at which LOH events can be selected. In this study, we used two techniques (single-nucleotide polymorphism microarrays and high-throughput DNA sequencing) to examine genome-wide LOH in a diploid yeast strain at a resolution averaging 1 kb. We examined both selected LOH events on chromosome V and unselected events throughout the genome in untreated cells and in cells treated with either γ-radiation or ultraviolet (UV) radiation. Our analysis shows the following: (1) spontaneous and damage-induced mitotic gene conversion tracts are more than three times larger than meiotic conversion tracts, and conversion tracts associated with crossovers are usually longer and more complex than those unassociated with crossovers; (2) most of the crossovers and conversions reflect the repair of two sister chromatids broken at the same position; and (3) both UV and γ-radiation efficiently induce LOH at doses of radiation that cause no significant loss of viability. Using high-throughput DNA sequencing, we also detected new mutations induced by γ-rays and UV. To our knowledge, our study represents the first high-resolution genome-wide analysis of DNA damage-induced LOH events performed in any eukaryote.

  8. A multi-ethnic genome-wide association study identifies novel loci for non-syndromic cleft lip with or without cleft palate on 2p24.2, 17q23 and 19q13.

    PubMed

    Leslie, Elizabeth J; Carlson, Jenna C; Shaffer, John R; Feingold, Eleanor; Wehby, George; Laurie, Cecelia A; Jain, Deepti; Laurie, Cathy C; Doheny, Kimberly F; McHenry, Toby; Resick, Judith; Sanchez, Carla; Jacobs, Jennifer; Emanuele, Beth; Vieira, Alexandre R; Neiswanger, Katherine; Lidral, Andrew C; Valencia-Ramirez, Luz Consuelo; Lopez-Palacio, Ana Maria; Valencia, Dora Rivera; Arcos-Burgos, Mauricio; Czeizel, Andrew E; Field, L Leigh; Padilla, Carmencita D; Cutiongco-de la Paz, Eva Maria C; Deleyiannis, Frederic; Christensen, Kaare; Munger, Ronald G; Lie, Rolv T; Wilcox, Allen; Romitti, Paul A; Castilla, Eduardo E; Mereb, Juan C; Poletta, Fernando A; Orioli, Iêda M; Carvalho, Flavia M; Hecht, Jacqueline T; Blanton, Susan H; Buxó, Carmen J; Butali, Azeez; Mossey, Peter A; Adeyemo, Wasiu L; James, Olutayo; Braimah, Ramat O; Aregbesola, Babatunde S; Eshete, Mekonen A; Abate, Fikre; Koruyucu, Mine; Seymen, Figen; Ma, Lian; de Salamanca, Javier Enríquez; Weinberg, Seth M; Moreno, Lina; Murray, Jeffrey C; Marazita, Mary L

    2016-07-01

    Orofacial clefts (OFCs), which include non-syndromic cleft lip with or without cleft palate (CL/P), are among the most common birth defects in humans, affecting approximately 1 in 700 newborns. CL/P is phenotypically heterogeneous and has a complex etiology caused by genetic and environmental factors. Previous genome-wide association studies (GWASs) have identified at least 15 risk loci for CL/P. As these loci do not account for all of the genetic variance of CL/P, we hypothesized the existence of additional risk loci. We conducted a multiethnic GWAS in 6480 participants (823 unrelated cases, 1700 unrelated controls and 1319 case-parent trios) with European, Asian, African and Central and South American ancestry. Our GWAS revealed novel associations on 2p24 near FAM49A, a gene of unknown function (P = 4.22 × 10(-8)), and 19q13 near RHPN2, a gene involved in organizing the actin cytoskeleton (P = 4.17 × 10(-8)). Other regions reaching genome-wide significance were 1p36 (PAX7), 1p22 (ARHGAP29), 1q32 (IRF6), 8q24 and 17p13 (NTN1), all reported in previous GWASs. Stratification by ancestry group revealed a novel association with a region on 17q23 (P = 2.92 × 10(-8)) among individuals with European ancestry. This region included several promising candidates including TANC2, an oncogene required for development, and DCAF7, a scaffolding protein required for craniofacial development. In the Central and South American ancestry group, significant associations with loci previously identified in Asian or European ancestry groups reflected their admixed ancestry. In summary, we have identified novel CL/P risk loci and suggest new genes involved in craniofacial development, confirming the highly heterogeneous etiology of OFCs.

  9. The human homolog of a mouse-imprinted gene, Peg3, maps to a zinc finger gene-rich region of human chromosome 19q13.4.

    PubMed

    Kim, J; Ashworth, L; Branscomb, E; Stubbs, L

    1997-05-01

    Peg3 (paternally expressed gene 3) is the first imprinted gene detected in the proximal region of mouse chromosome 7. Because imprinting is a trait that is generally conserved among mammals, and imprinted domains generally encompass several adjacent genes, expression patterns and chromosomal environment of the human counterpart of Peg3 are of special interest. In this study we have localized human PEG3 approximately 2 Mb proximal of the telomere of chromosome 19q, within a region known to carry large numbers of tandemly clustered Krüppel-type zinc finger-containing (ZNF) genes. Peg3 also encodes a Krüppel-type ZNF protein but one that is distinguished from other ZNF gene products by the fact that it carries two novel proline-rich motifs. Comparison between mouse Peg3 and partial human PEG3 gene sequences revealed a high level of conservation between the two species, despite the fact that one of the two proline-rich repeats is absent from the human gene. Our data demonstrate that the human gene is expressed at highest levels in ovary and placenta; mouse Peg3, by contrast, is transcribed at highest levels in the adult brain. These comparative mapping, sequencing, and expression data provide the first clues to the potential activities of PEG3, and generate new tools to aid in the analysis of structure and function of a potentially new imprinted domain located in human chromosome 19q13.4 and mouse chromosome 7.

  10. Assembly of a 1-Mb restriction-mapped cosmid contig spanning the candidate region for Finnish congenital nephrosis (NPHS1) in 19q13.1

    SciTech Connect

    Olsen, A.S.; Georgescu, A.; Johnson, S.; Carrano, A.V.

    1996-06-01

    We describe the assembly of a 1-Mb cosmid contig and restriction map spanning the candidate region for Finnish congenital nephrosis (NPHS1) in 19q13.1. The map was constructed from 16 smaller contigs assembled by fingerprinting, a BAC and a PAC clone, and 42 previously unmapped cosmids. In most cases, single-step cosmid walks were sufficient to join two previously assembled contigs, and all but one gap was filled from this cosmid contig library. The remaining gap of about 19 kb was spanned with a single BAC and a single PAC clone. EcoRI mapping of a dense set of overlapping clones validated the assembly of the map and indicated a length of 1040 kb for the contig. This high-resolution clone map provides an ideal resource for gene identification through cDNA selection, exon trapping, and DNA sequencing. 10 refs., 1 fig.

  11. Life Origination and Development Hydrate theory (LOH-Theory): new approaches to the problems of the optimal nutrition and life prolongation

    NASA Astrophysics Data System (ADS)

    Kadyshevich, E. A.; Ostrovskii, V. E.

    2014-04-01

    Life Origination Hydrate Theory (LOH-Theory) and Mitosis and Replication Hydrate Theory (MRHTheory), both grounded on the notion of honeycomb gas-hydrate structures formation/destruction as the physicochemical phenomenon governing the DNA origination and replication, allow new approaches to the optimal nutrition and life prolongation problems.

  12. Characterization of the human and rat phospholemman (PLM) cDNAs and localization of the human PLM gene to chromosome 19q13.1

    SciTech Connect

    Chen, Ling-Sing K.; Lo, C.F.; Numann, R.; Cuddy, M.

    1997-05-01

    Previous reports have demonstrated that the phospholemman (PLM), a 72-residue plasma-membrane protein enriched in skeletal muscle and heart, is a major substrate phosphorylated in response to insulin and adrenergic stimulation. Here we describe the isolation and characterization of human and rat PLM cDNA from the heart. Both PLM proteins share significant nucleotide and amino acid sequence and structural similarities with the previously published canine PLM and, to a lesser degree, with Na{sup +}/K{sup +}-ATPase {gamma} subunit, Mat-8 protein, and CHIF protein. Despite the functional diversity, all these proteins are quite small and possess a single transmembrane domain. Human PLM appears to be a unique gene localized on chromosome 19q13.1. The PLM mRNA is widely distributed in human tissues, with the highest expression in skeletal muscle and heart, suggesting a functional role in muscle contraction. Like canine PLM, both human and rat PLM induce a hyperpolarization-activated chloride current when expressed in Xenopus oocytes. The high degree of sequence and functional conservation among the mammalian PLM proteins indicates that this gene is conserved throughout evolution. 34 refs., 4 figs.

  13. Characterization of the human and rat phospholemman (PLM) cDNAs and localization of the human PLM gene to chromosome 19q13.1.

    PubMed

    Chen, L S; Lo, C F; Numann, R; Cuddy, M

    1997-05-01

    Previous reports have demonstrated that the phospholemman (PLM), a 72-residue plasma-membrane protein enriched in skeletal muscle and heart, is a major substrate phosphorylated in response to insulin and adrenergic stimulation. Here we describe the isolation and characterization of human and rat PLM cDNA from the heart. Both PLM proteins share significant nucleotide and amino acid sequence and structural similarities with the previously published canine PLM and, to a lesser degree, with Na+/K(+)-ATPase gamma subunit, Mat-8 protein, and CHIF protein. Despite the functional diversity, all these proteins are quite small and possess a single transmembrane domain. Human PLM appears to be a unique gene localized on chromosome 19q13.1. The PLM mRNA is widely distributed in human tissues, with the highest expression in skeletal muscle and heart, suggesting a functional role in muscle contraction. Like canine PLM, both human and rat PLM induce a hyperpolarization-activated chloride current when expressed in Xenopus oocytes. The high degree of sequence and functional conservation among the mammalian PLM proteins indicates that this gene is conserved throughout evolution.

  14. Mechanisms leading to Prader-Willi syndrome in a patient with a de novo 46, XY, t(15; 19)(q12; q13.41)

    SciTech Connect

    Sun, Y.; Hainline, B.E.; Palmer, C.G.

    1994-09-01

    A three year and six month-old boy with Prader-Willi syndrome (PWS) was found to have a de novo 46, XY, t(15; 19) (q12; q13.41) karyotype. PCR studies of microsatellite loci showed heterozygosity, including biparental inheritance. Fluorescence in situ hybridization (FISH) studies were performed with cosmid probes D15S11, SNRPN, D15S10, and GABRB3 and no deletion was found. The chromosomal breakage occurred inside the SNRPN contig, which contains two overlapping cosmids. Each cosmid shows signals with FISH on both the der(15) and the der(19), and on the normal chromosome 15. Additional FISH studies using cosmid subfragments demonstrated that the breakage occurred upstream to coding exons of the SNRPN gene. SNRPN contains 10 exons, including two recently identified upstream exons, exon-1 and exon-0. A probe from an RT-PCR product (1020bp) of total human brain mRNA spanning exons 1-8 and an exon1-specific probe were used on genome DNA Southern hybridizaiton. An extra DNA band 20kb in size was detected specifically from our patients genomic DNA using BamHl when compared to his normal parents and normal individuals. Further studies revealed that the breakage occurred between exon 0 and exon 1 of the SNRPN gene.

  15. Organization of the human gene for nucleobindin (NUC) and its chromosomal assignment to 19q13.2-q13.4

    SciTech Connect

    Miura, Keiji; Kurosawa, Yoshikazu; Hirai, Momoki

    1996-06-01

    Nucleobindin (Nuc) was first identified as a secreted protein of 55 kDa that promotes production of DNA-specific antibodies in lupus-prone MRL/lpr mice. Analysis of cDNA that encoded Nuc revealed that the protein is composed of a signal peptide, a DNA-binding site, two calcium-binding motifs (EF-hand motifs), and a leucine zipper. In the present study, we analysed the organization of the human gene for Nuc (NUC). It consists of 13 exons that are distributed in a region of 32 kb. The functional motifs listed above are encoded in corresponding exons. NUC was expressed in all organs examined. Comparison of nucleotide sequences in the promotre regions between human and mouse NCU genes revealed several conserved sequences. Among them, two Sp1-binding sites and a CCAAT box are of particular interest. The promoter is of the TATA-less type, and transcription starts at multiple sites in both the human and the mouse genes. These features suggest that NUC might normally play a role as a housekeeping gene. NUC was located at human chromosome 19q13.2-q13.4. 25 refs., 4 figs., 1 tab.

  16. Dnm1p-dependent peroxisome fission requires Caf4p, Mdv1p and Fis1p.

    PubMed

    Motley, Alison M; Ward, Gemma P; Hettema, Ewald H

    2008-05-15

    Yeast peroxisomes multiply by fission. Fission requires two dynamin-related proteins, Dnm1p and Vps1p. Using an in vivo fission assay, we show that Dnm1p-dependent peroxisome fission requires Fis1p, Caf4p and Mdv1p. Fluorescence microscopy of cells expressing GFP-tagged Caf4p and Mdv1p revealed that their association with peroxisomes relies on Fis1p. Vps1p-dependent peroxisome fission occurs independently of these factors. Vps1p contributes most to fission of peroxisomes when cells are grown on glucose. Overexpression of Dnm1p suppresses the fission defect as long as Fis1p and either Mdv1p or Caf4p are present. Conversely, overexpression of Dnm1p does not restore the vacuolar fusion defect of vps1 cells and Vps1p overexpression does not restore the mitochondrial fission defect of dnm1 cells. These data show that Vps1p and Dnm1p are part of independent fission machineries. Because the contribution of Dnm1p to peroxisome fission appears to be more pronounced in cells that proliferate peroxisomes in response to mitochondrial dysfunction, Dnm1p might be part of the mechanism that coordinates mitochondrial and peroxisomal biogenesis.

  17. Cloning of ELL, a gene that fuses to MLL in a t(11; 19)(q23; p13. 1) in acute myeloid leukemia

    SciTech Connect

    Thirman, M.J.; Levitan, D.A.; Kobayashi, H.; Simon, M.C.; Rowley, J.D. )

    1994-12-06

    To characterize the functions of MLL fusion transcripts, we cloned the gene that fuses to MLL in the translocation t(11;19)(q23;p13.1). This translocation is distinct from another type of 11;19 translocation with a 19p13.3 breakpoint that results in the fusion of MLL to the ENL gene. By PCR screening of a cDNA library prepared from a patient's leukemia cells with this translocation, we obtained a fusion transcript containing exon 7 of MLL and sequence of an unknown gene. The sequence of this gene was amplified and used as a probe to screen a fetal brain cDNA library. On Northern blot analysis, this cDNA detected a 4.4-kb transcript that was abundant in peripheral blood leukocytes, skeletal muscle, placenta, and testis and expressed at lower levels in spleen, thymus, heart, brain, lung, kidney, liver, and ovary. In addition, a 2.8-kb transcript was present in peripheral blood, testis, and placenta. On [open quotes]zoo blots,[close quotes] this gene was shown to be evolutionarily conserved in 10 mammalian species as well as in chicken, frog, and fish. We have named this gene ELL (for eleven-nineteen lysine-rich leukemia gene). A highly basic, lysine-rich motif of the predicted ELL protein is homologous to similar regions of several proteins, including the DNA-binding domain of poly(ADP-ribose) polymerase. The characterization of the normal functions of ELL as well as its altered function when fused to MLL will be critical to further our understanding of the mechanisms of leukemogenesis. 30 refs., 7 figs.

  18. A molecular cytogenetic study of imprinting effect in childhood acute lymphoblastic leukemia with the t(1;19)(q23;p13)

    SciTech Connect

    Knops, J.F.; Han, J.; Finley, W.H.

    1994-09-01

    Genomic imprinting (the functional difference between homologous alleles dependent upon parent of origin) seems to play an important role in some forms of recessive tumors as well as in the genetic predisposition to cancer. Recently, this phenomenon has been implicated in reciprocal translocations found in hematological malignancies. Cytogenetic polymorphism studies on the t(9;22) chromosome in chronic myelogenous leukemia have shown chromosome 9 to be paternal in origin while chromosome 22 is maternally derived in all the cases studied. We developed a molecular technique using chromosome microdissection and polymerase chain reaction (CMPCR) to study the possible involvement of genomic imprinting in childhood acute lymphoblastic leukemia (ALL) with the t(1;19). This technique has an advantage over conventional molecular techniques because it can distinguish the translocated chromosome from the normal homologue. Amplification of highly polymorphic microsatellite loci was performed on the microdissected translocated chromosomes and compared with parental alleles to assess the parent of origin of the chromosome 1 and chromosome 19 involved in the translocation. Parental origin of the derivative chromosomes 1 and 19 has been evaluated in six children with pre-B cell ALL and t(1;19)(q23;p13). The ratio of maternal:paternal:uncertain origin was 3:2:1 for chromosome 1 and 1:3:2 for chromosome 19. Thus far, the data do not indicate a strong parent of origin bias for this specific ALL associated translocation. Although genomic imprinting does not appear to be involved in this subgroup of childhood ALL patients, this technique can be used to determine parental origin of aberrant chromosomes in other hematological malignancies.

  19. [The diagnostic value of microsatellite LOH analysis and the prognostic relevance of angiogenic gene expression in urinary bladder cancer].

    PubMed

    Szarvas, Tibor

    2009-12-01

    Bladder cancer is the second most common malignancy affecting the urinary system. Currently, histology is the only tool that determines therapy and patients' prognosis. As the treatment of non-invasive (Ta/T1) and muscle invasive (T2-T4) bladder tumors are completely different, correct staging is important, although it is often hampered by disturbing factors. Molecular methods offer new prospects for early disease detection, confirmation of unclear histological findings and prognostication. Applying molecular biological methods, the present study is searching for answers to current diagnostic and prognostic problems in bladder carcinoma. We analyzed tumor, blood and/or urine samples of 334 bladder cancer patients and 117 control individuals. Genetic alterations were analyzed in urine samples of patients and controls, both by PCR-based microsatellite loss of heterozigosity (LOH) analysis using 12 fluorescently labeled primers and by DNA hybridization based UroVysion FISH technique using 4 probes, to assess the diagnostic values of these methods. Whole genome microsatellite analysis (with 400 markers) was performed in tumor and blood specimens of bladder cancer patients to find chromosomal regions, the loss of which may be associated with tumor stage. Furthermore, we assessed the prognostic value of Tie2, VEGF, Angiopoietin-1 and -2. We concluded that DNA analysis of voided urine samples by microsatellite analysis and FISH are sensitive and non-invasive methods to detect bladder cancer. Furthermore, we established a panel of microsatellite markers that could differentiate between non-invasive and invasive bladder cancer. However, further analyses in a larger cohort of patients are needed to assess their specificity and sensitivity. Finally, we identified high Ang-2 and low Tie2 gene expression as significant and independent risk factors of tumor recurrence and cancer related survival.

  20. Fine-mapping markers of lung cancer susceptibility in a sub-region of chromosome 19q13.3 among Chinese

    PubMed Central

    Yin, Jiaoyang; Wang, Huiwen; Vogel, Ulla; Wang, Chunhong; Ma, Yegang; Hou, Wei; Zhang, Ying; Guo, Li; Li, Xinxin

    2016-01-01

    Linkage disequilibrium-mapping studies in Caucasians have indicated anassociation of Chr19q13.3 sub-region spanning ERCC2, PPP1R13L, CD3EAP and ERCC1 with several cancers. To refine the region of association and identify potential causal variations among Asians, we performed a fine-mapping study using 32 (39) SNPs in a 71.654kb sub-region. The study included 384 Chinese lung cancer cases and 387 controls. Seven closely situated SNPs showed significant associations with lung cancer risk in five different genetic models of single-locus associations (adjusted for smoking duration). These were PPP1R13L rs1970764 [OR (95% CI) = 1.58 (1.09-2.29), P = 0.014] in a recessive model and PPP1R13L rs1005165 [OR (95% CI) = 1.25 (1.01-1.54), P = 0.036], CD3EAP rs967591 [OR (95% CI) = 1.40 (1.13-1.75), P = 0.0023], rs735482 [OR (95% CI) = 1.29 (1.03-1.61), P = 0.026], rs1007616 [OR (95% CI) = 0.78 (0.61-1.00), P = 0.046], and rs62109563 [OR (95% CI) = 1.28 (1.03-1.59), P = 0.024] in a log-additive model and ERCC1 rs3212965 [OR (95% CI) = 0.70 (0.52-0.94), P = 0.019] in an over-dominant model. Six-haplotype blocks were determined in the sub-region. Using an alternative approach where we performed a haplotype analysis of all significant polymorphisms, rs1970764 was found to be most consistently associated with lung cancer risk. The combined data suggest that the sub-region with the strongest association to lung cancer susceptibility might locate to the 23.173kb from PPP1R13L intron8 rs1970764 to rs62109563 3′ to CD3EAP. Limited risk loci and span on lung cancer in this sub-region are initially defined among Asians. PMID:27183913

  1. S1P control of endothelial integrity.

    PubMed

    Xiong, Yuquan; Hla, Timothy

    2014-01-01

    Sphingosine 1-phosphate (S1P), a lipid mediator produced by sphingolipid metabolism, promotes endothelial cell spreading, vascular maturation/stabilization, and barrier function. S1P is present at high concentrations in the circulatory system, whereas in tissues its levels are low. This so-called vascular S1P gradient is essential for S1P to regulate much physiological and pathophysiological progress such as the modulation of vascular permeability. Cellular sources of S1P in blood has only recently begun to be identified. In this review, we summarize the current understanding of S1P in regulating vascular integrity. In particular, we discuss the recent discovery of the endothelium-protective functions of HDL-bound S1P which is chaperoned by apolipoprotein M.

  2. Acquired copy-neutral loss of heterozygosity of chromosome 1p as a molecular event associated with marrow fibrosis in MPL-mutated myeloproliferative neoplasms.

    PubMed

    Rumi, Elisa; Pietra, Daniela; Guglielmelli, Paola; Bordoni, Roberta; Casetti, Ilaria; Milanesi, Chiara; Sant'Antonio, Emanuela; Ferretti, Virginia; Pancrazzi, Alessandro; Rotunno, Giada; Severgnini, Marco; Pietrelli, Alessandro; Astori, Cesare; Fugazza, Elena; Pascutto, Cristiana; Boveri, Emanuela; Passamonti, Francesco; De Bellis, Gianluca; Vannucchi, Alessandro; Cazzola, Mario

    2013-05-23

    We studied mutations of MPL exon 10 in patients with essential thrombocythemia (ET) or primary myelofibrosis (PMF), first investigating a cohort of 892 consecutive patients. MPL mutation scanning was performed on granulocyte genomic DNA by using a high-resolution melt assay, and the mutant allele burden was evaluated by using deep sequencing. Somatic mutations of MPL, all but one involving codon W515, were detected in 26/661 (4%) patients with ET, 10/187 (5%) with PMF, and 7/44 (16%) patients with post-ET myelofibrosis. Comparison of JAK2 (V617F)-mutated and MPL-mutated patients showed only minor phenotypic differences. In an extended group of 62 MPL-mutated patients, the granulocyte mutant allele burden ranged from 1% to 95% and was significantly higher in patients with PMF or post-ET myelofibrosis compared with those with ET. Patients with higher mutation burdens had evidence of acquired copy-neutral loss of heterozygosity (CN-LOH) of chromosome 1p in granulocytes, consistent with a transition from heterozygosity to homozygosity for the MPL mutation in clonal cells. A significant association was found between MPL-mutant allele burden greater than 50% and marrow fibrosis. These observations suggest that acquired CN-LOH of chromosome 1p involving the MPL location may represent a molecular mechanism of fibrotic transformation in MPL-mutated myeloproliferative neoplasms.

  3. Acquired copy-neutral loss of heterozygosity of chromosome 1p as a molecular event associated with marrow fibrosis in MPL-mutated myeloproliferative neoplasms

    PubMed Central

    Pietra, Daniela; Guglielmelli, Paola; Bordoni, Roberta; Casetti, Ilaria; Milanesi, Chiara; Sant’Antonio, Emanuela; Ferretti, Virginia; Pancrazzi, Alessandro; Rotunno, Giada; Severgnini, Marco; Pietrelli, Alessandro; Astori, Cesare; Fugazza, Elena; Pascutto, Cristiana; Boveri, Emanuela; Passamonti, Francesco; De Bellis, Gianluca; Vannucchi, Alessandro; Cazzola, Mario

    2013-01-01

    We studied mutations of MPL exon 10 in patients with essential thrombocythemia (ET) or primary myelofibrosis (PMF), first investigating a cohort of 892 consecutive patients. MPL mutation scanning was performed on granulocyte genomic DNA by using a high-resolution melt assay, and the mutant allele burden was evaluated by using deep sequencing. Somatic mutations of MPL, all but one involving codon W515, were detected in 26/661 (4%) patients with ET, 10/187 (5%) with PMF, and 7/44 (16%) patients with post-ET myelofibrosis. Comparison of JAK2 (V617F)–mutated and MPL-mutated patients showed only minor phenotypic differences. In an extended group of 62 MPL-mutated patients, the granulocyte mutant allele burden ranged from 1% to 95% and was significantly higher in patients with PMF or post-ET myelofibrosis compared with those with ET. Patients with higher mutation burdens had evidence of acquired copy-neutral loss of heterozygosity (CN-LOH) of chromosome 1p in granulocytes, consistent with a transition from heterozygosity to homozygosity for the MPL mutation in clonal cells. A significant association was found between MPL-mutant allele burden greater than 50% and marrow fibrosis. These observations suggest that acquired CN-LOH of chromosome 1p involving the MPL location may represent a molecular mechanism of fibrotic transformation in MPL-mutated myeloproliferative neoplasms. PMID:23575445

  4. 1p36 deletion syndrome: an update.

    PubMed

    Jordan, Valerie K; Zaveri, Hitisha P; Scott, Daryl A

    2015-01-01

    Deletions of chromosome 1p36 affect approximately 1 in 5,000 newborns and are the most common terminal deletions in humans. Medical problems commonly caused by terminal deletions of 1p36 include developmental delay, intellectual disability, seizures, vision problems, hearing loss, short stature, distinctive facial features, brain anomalies, orofacial clefting, congenital heart defects, cardiomyopathy, and renal anomalies. Although 1p36 deletion syndrome is considered clinically recognizable, there is significant phenotypic variation among affected individuals. This variation is due, at least in part, to the genetic heterogeneity seen in 1p36 deletions which include terminal and interstitial deletions of varying lengths located throughout the 30 Mb of DNA that comprise chromosome 1p36. Array-based copy number variant analysis can easily identify genomic regions of 1p36 that are deleted in an affected individual. However, predicting the phenotype of an individual based solely on the location and extent of their 1p36 deletion remains a challenge since most of the genes that contribute to 1p36-related phenotypes have yet to be identified. In addition, haploinsufficiency of more than one gene may contribute to some phenotypes. In this article, we review recent successes in the effort to map and identify the genes and genomic regions that contribute to specific 1p36-related phenotypes. In particular, we highlight evidence implicating MMP23B, GABRD, SKI, PRDM16, KCNAB2, RERE, UBE4B, CASZ1, PDPN, SPEN, ECE1, HSPG2, and LUZP1 in various 1p36 deletion phenotypes.

  5. 1p36 deletion syndrome: an update

    PubMed Central

    Jordan, Valerie K; Zaveri, Hitisha P; Scott, Daryl A

    2015-01-01

    Deletions of chromosome 1p36 affect approximately 1 in 5,000 newborns and are the most common terminal deletions in humans. Medical problems commonly caused by terminal deletions of 1p36 include developmental delay, intellectual disability, seizures, vision problems, hearing loss, short stature, distinctive facial features, brain anomalies, orofacial clefting, congenital heart defects, cardiomyopathy, and renal anomalies. Although 1p36 deletion syndrome is considered clinically recognizable, there is significant phenotypic variation among affected individuals. This variation is due, at least in part, to the genetic heterogeneity seen in 1p36 deletions which include terminal and interstitial deletions of varying lengths located throughout the 30 Mb of DNA that comprise chromosome 1p36. Array-based copy number variant analysis can easily identify genomic regions of 1p36 that are deleted in an affected individual. However, predicting the phenotype of an individual based solely on the location and extent of their 1p36 deletion remains a challenge since most of the genes that contribute to 1p36-related phenotypes have yet to be identified. In addition, haploinsufficiency of more than one gene may contribute to some phenotypes. In this article, we review recent successes in the effort to map and identify the genes and genomic regions that contribute to specific 1p36-related phenotypes. In particular, we highlight evidence implicating MMP23B, GABRD, SKI, PRDM16, KCNAB2, RERE, UBE4B, CASZ1, PDPN, SPEN, ECE1, HSPG2, and LUZP1 in various 1p36 deletion phenotypes. PMID:26345236

  6. Carboxyarabinitol-1-P phosphatase of Phaseolus vulgaris

    SciTech Connect

    Kobza, J.; Moore, B.d.; Seemann, J.R. )

    1990-05-01

    The activity of carboxyarabinitol-1-P (CA1P) phosphatase was detected in clarified stromal extracts by the generation of {sup 14}C-carboxyarabinitol from {sup 14}C-CA1P. Carboxyribitol-1-P dependent activity was 3% of the CA1P dependent activity, indicating the enzyme was specific for CA1P. Inclusion of DTT in the assay was required for maximum velocity, but it appears that the enzyme is not regulated by thioredoxin in vivo. Activity o f the CA1P phosphatase was stimulated by RuBP, NADPH and FBP, though the latter two metabolites were required at nonphysiological concentrations in order to achieve significant stimulation. Contrary to a previous report on purified tobacco enzyme, ATP stimulated the CA1P phosphatase activity. In the presence of 1 mM RuBP or ATP, rates of 2 or 3 {mu}mol mg{sup {minus}1} Chl h{sup {minus}1}, respectively, were observed at 1 mM CA1P. These rates were 3-4 fold higher than the rate observed in the absence of effectors and are 2-4 times the in vivo rate of degradation of CA1P during dark/light transitions. The rates from bean were about 7 fold higher than rates reported for the enzyme from tobacco. Changes in the levels of ATP and RuBP associated with dark/light transitions could modulate the enzyme activity in vivo, but it remains to be established if this is the only mechanism for the required regulation of the enzyme.

  7. Loss of expression of the metastasis suppressor gene KiSS1 during melanoma progression and its association with LOH of chromosome 6q16.3-q23.

    PubMed

    Shirasaki, F; Takata, M; Hatta, N; Takehara, K

    2001-10-15

    KiSS1 is a putative melanoma metastasis suppressor gene, the expression of which may be regulated by another gene(s) mapping to chromosome 6q16.3-q23. To additionally elucidate the role of KiSS1 in the progression of human melanoma in vivo, we examined KiSS1 mRNA expression in 51 melanocytic tumors with various stages of progression by in situ hybridization. We also examined a correlation between loss of KiSS1 mRNA expression and loss of heterozygosity (LOH) of 6q16.3-q23 in 27 melanoma metastases. All of the four nevocellular nevi and eight primary melanomas <4 mm in thickness showed KiSS1 mRNA expression, whereas only 50% (6 of 12) of primary melanomas >4 mm in thickness expressed KiSS1. Loss of KiSS1 mRNA was equally frequent in metastases; 44% (12 of 27) of tumors lost KiSS1 expression. LOH of 6q16.3-q23 was observed in 52% (14 of 27) of metastases. There was a strong association between LOH and loss of KiSS1 expression (P = 0.03); nine metastases with LOH of 6q16.3-q23 lost KiSS1 expression, whereas 10 tumors with no LOH showed positive KiSS1 mRNA expression. The findings in this study show, for the first time, KiSS1 down-regulation during the progression of melanoma in vivo and strongly suggest that inactivation of a tumor suppressor gene(s) mapping to 6q16.3-q23 by deletion or mutation coupled with LOH may lead to the down-regulation of KiSS1.

  8. Loss of heterozygosity (LOH) for markers on chromosome 8q in a human chondrosarcoma cell line and in a tumor that developed in a man with Hereditary Multiple Exostoses (HME)

    SciTech Connect

    Raskind, W.H.; Conrad, E.U.; Robbins, J.R.

    1994-09-01

    HME is an autosomal dominant disorder in which multiple benign cartilage-capped lesions develop on otherwise histologically normal bones. The majority of chondrosarcomas are sporadic, but the presence of HME greatly increases the risk to develop this tumor. Sarcoma may also arise in sporadically-occurring exostoses. The study of inherited disorders that predispose to malignant diseases has led to discoveries regarding molecular changes involved in carcinogenesis in general. In an analogous manner, somatic mutations in HME genes may be responsible for sporadic exostoses and/or chondrosarcomas. HME is genetically heterogeneous; EXT genes have been assigned to 8q24, the pericentromeric region of 11 and 19p11-p13. We compared chondrosarcoma cell DNA to DNA from white blood cells for LOH at polymorphic loci in these 3 regions. LOH for 4 of 5 markers in 8q24 was detected in a chondrosarcoma that arose in a man with HME. Heterozygosity was retained for markers and chromosomes 11 and 19. We then evaluated cultured cells from 10 sporadic chondrosarcomas. LOH for multiple markers in 8q24 was detected in a cell line, Ch-1, established from an aggressive tumor, but not in 9 other tumors. Of the 9 tumors studied, only the Ch-1 line exhibited LOH for chromosome 11 markers. LOH for a 19p marker was not detected in any of 6 tumors examined, including Ch-1. The karyotype of Ch-1 contains many structurally rearranged chromosomes. The two chromosome 11s appear normal but both chromosome 8 homologues have been replaced by der(8)t(5;8)(q22;q21.2). LOH at 8q24 was also detected in the uncultured tumor. These results suggest that genes responsible for HME may have tumor suppressor functions whose loss may be related to the development of a subset of chondrosarcomas.

  9. 1p36 is a preferential target of chromosome 1 deletions in astrocytic tumours and homozygously deleted in a subset of glioblastomas

    PubMed Central

    Ichimura, K; Vogazianou, AP.; Liu, L; Pearson, DM.; Bäcklund, LM; Plant, K; Baird, K; Langford, CF.; Gregory, SG.; Collins, VP

    2009-01-01

    Astrocytic, oligodendroglial and mixed gliomas are the commonest gliomas in adults. They have distinct phenotypes and clinical courses, but as they exist as a continuous histological spectrum differentiating them can be difficult. Co-deletions of total 1p and 19q are found in the majority of oligodendrogliomas and considered as a diagnostic marker and a prognostic indicator. The 1p status of astrocytomas has not yet been thoroughly examined. Using a chromosome 1 tile path array, we investigated 108 adult astrocytic tumours for copy number alterations. Total 1p deletions were rare (2%), however partial deletions involving 1p36 were frequently identified in anaplastic astrocytomas (22%) and glioblastomas (34%). Multivariate analysis showed that patients with total 1p deletions had significantly longer survival (p=0.005). In 9 glioblastomas homozygous deletions at 1p36 were identified. No somatic mutations were found among the 5 genes located in the homozygously deleted region. However, the CpG island of TNFRSF9 was hypermethylated in 19% of astrocytic tumours and 87% of glioma cell lines. TNFRSF9 expression was upregulated after demethylation of glioma cell lines. Akt3 amplifications were found in four glioblastomas. Our results indicate that 1p deletions are common anaplastic astrocytomas and glioblastomas but are distinct from the 1p abnormalities in oligodendrogliomas. PMID:17934521

  10. Monosomy 1p36 deletion syndrome.

    PubMed

    Gajecka, Marzena; Mackay, Katherine L; Shaffer, Lisa G

    2007-11-15

    Monosomy 1p36 results from a heterozygous deletion of the most distal chromosomal band on the short arm of chromosome 1. Occurring in approximately 1 in 5,000 live births, monosomy 1p36 is the most common terminal deletion observed in humans. Monosomy 1p36 is associated with mental retardation, developmental delay, hearing impairment, seizures, growth impairment, hypotonia, and heart defects. The syndrome is also characterized by several distinct dysmorphic features, including large anterior fontanels, microcephaly, brachycephaly, deep-set eyes, flat nose and nasal bridge, and pointed chin. Several genes have been proposed as causative for individual features of the phenotype. In addition, based upon molecular characterization of subjects with monosomy 1p36, several mechanisms for the generation and stabilization of terminal deletions have been proposed.

  11. Sphingosine-1-phosphate (S1P) displays sustained S1P1 receptor agonism and signaling through S1P lyase-dependent receptor recycling.

    PubMed

    Gatfield, John; Monnier, Lucile; Studer, Rolf; Bolli, Martin H; Steiner, Beat; Nayler, Oliver

    2014-07-01

    The sphingosine-1-phosphate (S1P) type 1 receptor (S1P1R) is a novel therapeutic target in lymphocyte-mediated autoimmune diseases. S1P1 receptor desensitization caused by synthetic S1P1 receptor agonists prevents T-lymphocyte egress from secondary lymphoid organs into the circulation. The selective S1P1 receptor agonist ponesimod, which is in development for the treatment of autoimmune diseases, efficiently reduces peripheral lymphocyte counts and displays efficacy in animal models of autoimmune disease. Using ponesimod and the natural ligand S1P, we investigated the molecular mechanisms leading to different signaling, desensitization and trafficking behavior of S1P1 receptors. In recombinant S1P1 receptor-expressing cells, ponesimod and S1P triggered Gαi protein-mediated signaling and β-arrestin recruitment with comparable potency and efficiency, but only ponesimod efficiently induced intracellular receptor accumulation. In human umbilical vein endothelial cells (HUVEC), ponesimod and S1P triggered translocation of the endogenous S1P1 receptor to the Golgi compartment. However, only ponesimod treatment caused efficient surface receptor depletion, receptor accumulation in the Golgi and degradation. Impedance measurements in HUVEC showed that ponesimod induced only short-lived Gαi protein-mediated signaling followed by resistance to further stimulation, whereas S1P induced sustained Gαi protein-mediated signaling without desensitization. Inhibition of S1P lyase activity in HUVEC rendered S1P an efficient S1P1 receptor internalizing compound and abrogated S1P-mediated sustained signaling. This suggests that S1P lyase - by facilitating S1P1 receptor recycling - is essential for S1P-mediated sustained signaling, and that synthetic agonists are functional antagonists because they are not S1P lyase substrates.

  12. Cometary gas relations 1P/Halley

    NASA Astrophysics Data System (ADS)

    Voelzke, Marcos R.

    Photographic and photoelectric observations of comet 1P/Halley's ionised gas coma from CO+ at 4,250 Å and neutral gas coma from CN at 3,880 Å were part of the Bochum Halley Monitoring Program, conducted at the European Southern Observatory, La Silla, Chile, from February 17 to April 17, 1986. In this spectral range it is possible to see the continuum formation, motion and expansion of plasma and neutral gas structures. To observe the morphology of these structures, 32 CO+ photos (glass plates) from comet 1P/Halley obtained by means of an interference filter have been analysed. They have a field of view of 28.6 × 28.6 degrees and were obtained from March 29 to April 17, 1986 with exposure times between 20 and 120 minutes. All photos were digitised with a PDS 2020 GM microdensitometer. After digitisation, the data were reduced to relative intensities, and those with proper calibrations were also converted to absolute intensities, expressed in terms of column densities. The CO+ absolute intensity values still contain the continuum intensity. To calculate the CO+ column density it is necessary to subtract this continuum intensity. The relations between CO+ and CN in average column density values (NCO+/NCN) are 11.6 for a circular diaphragm with average diameter (Φ) of 6.1 arcminutes which corresponds to a distance from the nucleus (ρ) equal to 6.3 × 104 km; 20.0 for Φ = 7.1 arcminutes and ρ = 7.3 × 104 km; 8.1 for Φ = 8.5 arcminutes and ρ = 8.7 × 104 km; 35.6 for Φ = 11.9 arcminutes and ρ = 1.2 × 105 km; and 31.3 for Φ = 16.7 arcminutes and ρ = 1.7 × 105 km. These values are in perfect agreement with the data for short distances (ρ from 3.9 × 103 to 1.2 × 104 km) and small slit diameters (Φ from 0.4 to 1.2 arcminutes). With the use of diaphragms with large diameters it is possible to get some information about the outer coma of the comet (in this paper, from 60,000 until 170,000 km away from the nucleus). At these distances, the CO+ column density

  13. Chromosome 19q13 disruption alters expressions of CYP2A7, MIA and MIA-RAB4B lncRNA and contributes to FAP-like phenotype in APC mutation-negative familial colorectal cancer patients.

    PubMed

    Thean, Lai Fun; Wong, Yu Hui; Lo, Michelle; Loi, Carol; Chew, Min Hoe; Tang, Choong Leong; Cheah, Peh Yean

    2017-01-01

    Familial adenomatous polyposis (FAP) is an autosomal-dominantly inherited form of colorectal cancer (CRC) caused by mutation in the adenomatous polyposis coli (APC) gene. Our ability to exhaustively screen for APC mutations identify microsatellite-stable and APC-mutation negative familial CRC patients, enabling us to search for novel genes. We performed genome-wide scan on two affected siblings of one family and 88 ethnicity- and gender-matched healthy controls to identify deletions shared by the siblings. Combined loss of heterozygosity, copy number and allelic-specific copy number analysis uncovered 5 shared deletions. Long-range polymerase chain reaction (PCR) confirmed chromosome 19q13 deletion, which was subsequently found in one other family. The 32 kb deleted region harbors the CYP2A7 gene and was enriched with enhancer, repressor and insulator sites. The wildtype allele was lost in the polyps of the proband. Further, real-time RT-PCR assays showed that expressions of MIA and MIA-RAB4B located 35 kb upstream of the deletion, were up-regulated in the polyps compared to the matched mucosa of the proband. MIA-RAB4B, the read-through long non-coding RNA (lncRNA), RAB4B, PIM2 and TAOK1 share common binding site of a microRNA, miR-24, in their 3'UTRs. PIM2 and TAOK1, two target oncogenes of miR-24, were co-ordinately up-regulated with MIA-RAB4B in the polyps, suggesting that MIA-RAB4B could function as competitive endogenous RNA to titrate miR-24 away from its other targets. The data suggest that the 19.13 deletion disrupted chromatin boundary, leading to altered expression of several genes and lncRNA, could contribute to colorectal cancer via novel genetic and epigenetic mechanisms.

  14. Chromosome 19q13 disruption alters expressions of CYP2A7, MIA and MIA-RAB4B lncRNA and contributes to FAP-like phenotype in APC mutation-negative familial colorectal cancer patients

    PubMed Central

    Thean, Lai Fun; Wong, Yu Hui; Lo, Michelle; Loi, Carol; Chew, Min Hoe; Tang, Choong Leong; Cheah, Peh Yean

    2017-01-01

    Familial adenomatous polyposis (FAP) is an autosomal-dominantly inherited form of colorectal cancer (CRC) caused by mutation in the adenomatous polyposis coli (APC) gene. Our ability to exhaustively screen for APC mutations identify microsatellite-stable and APC-mutation negative familial CRC patients, enabling us to search for novel genes. We performed genome-wide scan on two affected siblings of one family and 88 ethnicity- and gender-matched healthy controls to identify deletions shared by the siblings. Combined loss of heterozygosity, copy number and allelic-specific copy number analysis uncovered 5 shared deletions. Long-range polymerase chain reaction (PCR) confirmed chromosome 19q13 deletion, which was subsequently found in one other family. The 32 kb deleted region harbors the CYP2A7 gene and was enriched with enhancer, repressor and insulator sites. The wildtype allele was lost in the polyps of the proband. Further, real-time RT-PCR assays showed that expressions of MIA and MIA-RAB4B located 35 kb upstream of the deletion, were up-regulated in the polyps compared to the matched mucosa of the proband. MIA-RAB4B, the read-through long non-coding RNA (lncRNA), RAB4B, PIM2 and TAOK1 share common binding site of a microRNA, miR-24, in their 3’UTRs. PIM2 and TAOK1, two target oncogenes of miR-24, were co-ordinately up-regulated with MIA-RAB4B in the polyps, suggesting that MIA-RAB4B could function as competitive endogenous RNA to titrate miR-24 away from its other targets. The data suggest that the 19.13 deletion disrupted chromatin boundary, leading to altered expression of several genes and lncRNA, could contribute to colorectal cancer via novel genetic and epigenetic mechanisms. PMID:28306719

  15. The functional roles of S1P in immunity.

    PubMed

    Hisano, Yu; Nishi, Tsuyoshi; Kawahara, Atsuo

    2012-10-01

    The lipid mediator sphingosine-1-phosphate (S1P) is generated within cells from sphingosine by two sphingosine kinases (SPHK1 and SPHK2). Intracellularly synthesized S1P is released into the extracellular fluid by S1P transporters, including SPNS2. Released S1P binds specifically to the G protein-coupled S1P receptors (S1PR1/S1P(1)-S1PR5/S1P(5)), which activate a diverse range of downstream signalling pathways. Recent studies have proposed that one of the central physiological functions of intercellular S1P signalling is in lymphocyte trafficking in vivo because genetic disruption of SPHK1/2, SPNS2 or S1PR1/S1P(1) in mice induces a lymphopenia phenotype. In this review, we discuss the current understanding of intercellular S1P signalling in the context of immunity.

  16. The IPP gene is assigned to human chromosome 1p32-1p22

    SciTech Connect

    Chang-Yeh, A.; Huang, R.C.C. ); Jabs, E.W.; Li, Xiang ); Dracopoli, N.C. )

    1993-01-01

    We previously reported the isolation and characterization of a novel mouse gene that is promoted by an intracisternal A-particle (IAP) LTR and is expressed in placental tissue (mouse IAP-promoted placenta gene, Ipp). Based on restriction fragment length polymorphism (RFLP) studies using inbred strains and recombinant inbred (RI) mice, we have established the linkage between the Ipp gene and several loci on distal mouse chromosome 4. In this publication, we report the partial sequence of a human cDNA clone isolated from a human placental library that has 83% identity to the 3[prime]region of the Ipp cDNA. For human chromosome mapping, two 20-base oligonucleotides within the homologous region were used as primers for polymerase chain reactions (PCR) to chromosome-specific DNAs from two somatic cell hybrid panels and several hybrid cell lines carrying breakpoints on human chromosome 1p. We have assigned this human homolog of the Ipp (IPP) gene to chromosome 1 at 1p32-1p22, based on analysis of PCR products. With this assignment, the homology between mouse chromosome 4 and human chromosome 1 is maintained (5). 7 refs., 1 fig.

  17. Yet1p-Yet3p interacts with Scs2p-Opi1p to regulate ER localization of the Opi1p repressor.

    PubMed

    Wilson, Joshua D; Thompson, Sarah L; Barlowe, Charles

    2011-05-01

    Lipid sensing mechanisms at the endoplasmic reticulum (ER) coordinate an array of biosynthetic pathways. A major phospholipid regulatory circuit in yeast is controlled by Scs2p, an ER membrane protein that binds the transcriptional repressor protein Opi1p. Cells grown in the absence of inositol sequester Scs2p-Opi1p at the ER and derepress target genes including INO1. We recently reported that Yet1p and Yet3p, the yeast homologues of BAP29 and BAP31, are required for normal growth in the absence of inositol. Here we show that the Yet1p-Yet3p complex acts in derepression of INO1 through physical association with Scs2p-Opi1p. Yet complex binding to Scs2p-Opi1p was enhanced by inositol starvation, although the interaction between Scs2p and Opi1p was not influenced by YET1 or YET3 deletion. Interestingly, live-cell imaging analysis indicated that Opi1p does not efficiently relocalize to the ER during inositol starvation in yet3Δ cells. Together our data demonstrate that a physical association between the Yet complex and Scs2p-Opi1p is required for proper localization of the Opi1p repressor to ER membranes and subsequent INO1 derepression.

  18. Exogenous S1P Exposure Potentiates Ischemic Stroke Damage That Is Reduced Possibly by Inhibiting S1P Receptor Signaling.

    PubMed

    Moon, Eunjung; Han, Jeong Eun; Jeon, Sejin; Ryu, Jong Hoon; Choi, Ji Woong; Chun, Jerold

    2015-01-01

    Initial and recurrent stroke produces central nervous system (CNS) damage, involving neuroinflammation. Receptor-mediated S1P signaling can influence neuroinflammation and has been implicated in cerebral ischemia through effects on the immune system. However, S1P-mediated events also occur within the brain itself where its roles during stroke have been less well studied. Here we investigated the involvement of S1P signaling in initial and recurrent stroke by using a transient middle cerebral artery occlusion/reperfusion (M/R) model combined with analyses of S1P signaling. Gene expression for S1P receptors and involved enzymes was altered during M/R, supporting changes in S1P signaling. Direct S1P microinjection into the normal CNS induced neuroglial activation, implicating S1P-initiated neuroinflammatory responses that resembled CNS changes seen during initial M/R challenge. Moreover, S1P microinjection combined with M/R potentiated brain damage, approximating a model for recurrent stroke dependent on S1P and suggesting that reduction in S1P signaling could ameliorate stroke damage. Delivery of FTY720 that removes S1P signaling with chronic exposure reduced damage in both initial and S1P-potentiated M/R-challenged brain, while reducing stroke markers like TNF-α. These results implicate direct S1P CNS signaling in the etiology of initial and recurrent stroke that can be therapeutically accessed by S1P modulators acting within the brain.

  19. Blocking S1P interaction with S1P{sub 1} receptor by a novel competitive S1P{sub 1}-selective antagonist inhibits angiogenesis

    SciTech Connect

    Fujii, Yasuyuki; Ueda, Yasuji; Ohtake, Hidenori; Ono, Naoya; Takayama, Tetsuo; Nakazawa, Kiyoshi; Igarashi, Yasuyuki; Goitsuka, Ryo

    2012-03-23

    Highlights: Black-Right-Pointing-Pointer The effect of a newly developed S1P{sub 1}-selective antagonist on angiogenic responses. Black-Right-Pointing-Pointer S1P{sub 1} is a critical component of VEGF-related angiogenic responses. Black-Right-Pointing-Pointer S1P{sub 1}-selective antagonist showed in vitro activity to inhibit angiogenesis. Black-Right-Pointing-Pointer S1P{sub 1}-selective antagonist showed in vivo activity to inhibit angiogenesis. Black-Right-Pointing-Pointer The efficacy of S1P{sub 1}-selective antagonist for anti-cancer therapies. -- Abstract: Sphingosine 1-phosphate receptor type 1 (S1P{sub 1}) was shown to be essential for vascular maturation during embryonic development and it has been demonstrated that substantial crosstalk exists between S1P{sub 1} and other pro-angiogenic growth factors, such as vascular endothelial growth factor (VEGF) and basic fibroblast growth factor. We developed a novel S1P{sub 1}-selective antagonist, TASP0277308, which is structurally unrelated to S1P as well as previously described S1P{sub 1} antagonists. TASP0277308 inhibited S1P- as well as VEGF-induced cellular responses, including migration and proliferation of human umbilical vein endothelial cells. Furthermore, TASP0277308 effectively blocked a VEGF-induced tube formation in vitro and significantly suppressed tumor cell-induced angiogenesis in vivo. These findings revealed that S1P{sub 1} is a critical component of VEGF-related angiogenic responses and also provide evidence for the efficacy of TASP0277308 for anti-cancer therapies.

  20. Genetics Home Reference: 1p36 deletion syndrome

    MedlinePlus

    ... Understand Genetics Home Health Conditions 1p36 deletion syndrome 1p36 deletion syndrome Enable Javascript to view the expand/collapse boxes. Download PDF Open All Close All Description 1p36 deletion syndrome is a disorder that typically causes severe intellectual ...

  1. Chemical and genetic tools to explore S1P biology.

    PubMed

    Cahalan, Stuart M

    2014-01-01

    The zwitterionic lysophospholipid Sphingosine 1-Phosphate (S1P) is a pleiotropic mediator of physiology and pathology. The synthesis, transport, and degradation of S1P are tightly regulated to ensure that S1P is present in the proper concentrations in the proper location. The binding of S1P to five G protein-coupled S1P receptors regulates many physiological systems, particularly the immune and vascular systems. Our understanding of the functions of S1P has been aided by the tractability of the system to both chemical and genetic manipulation. Chemical modulators have been generated to affect most of the known components of S1P biology, including agonists of S1P receptors and inhibitors of enzymes regulating S1P production and degradation. Genetic knockouts and manipulations have been similarly engineered to disrupt the functions of individual S1P receptors or enzymes involved in S1P metabolism. This chapter will focus on the development and utilization of these chemical and genetic tools to explore the complex biology surrounding S1P and its receptors, with particular attention paid to the in vivo findings that these tools have allowed for.

  2. The MAPK Hog1p Modulates Fps1p-dependent Arsenite Uptake and Tolerance in Yeast

    PubMed Central

    Thorsen, Michael; Di, Yujun; Tängemo, Carolina; Morillas, Montserrat; Ahmadpour, Doryaneh; Van der Does, Charlotte; Wagner, Annemarie; Johansson, Erik; Boman, Johan; Posas, Francesc; Wysocki, Robert

    2006-01-01

    Arsenic is widely distributed in nature and all organisms possess regulatory mechanisms to evade toxicity and acquire tolerance. Yet, little is known about arsenic sensing and signaling mechanisms or about their impact on tolerance and detoxification systems. Here, we describe a novel role of the S. cerevisiae mitogen-activated protein kinase Hog1p in protecting cells during exposure to arsenite and the related metalloid antimonite. Cells impaired in Hog1p function are metalloid hypersensitive, whereas cells with elevated Hog1p activity display improved tolerance. Hog1p is phosphorylated in response to arsenite and this phosphorylation requires Ssk1p and Pbs2p. Arsenite-activated Hog1p remains primarily cytoplasmic and does not mediate a major transcriptional response. Instead, hog1Δ sensitivity is accompanied by elevated cellular arsenic levels and we demonstrate that increased arsenite influx is dependent on the aquaglyceroporin Fps1p. Fps1p is phosphorylated on threonine 231 in vivo and this phosphorylation critically affects Fps1p activity. Moreover, Hog1p is shown to affect Fps1p phosphorylation. Our data are the first to demonstrate Hog1p activation by metalloids and provides a mechanism by which this kinase contributes to tolerance acquisition. Understanding how arsenite/antimonite uptake and toxicity is modulated may prove of value for their use in medical therapy. PMID:16885417

  3. Targeting sphingosine 1-phosphate (S1P) levels and S1P receptor functions for therapeutic immune interventions.

    PubMed

    Gräler, Markus H

    2010-01-01

    Sphingosine 1-phosphate (S1P) is an important regulator of many different immune functions including lymphocyte circulation, antigen presentation, and T cell development. It stimulates five G protein-coupled receptors designated S1P(1-5), which are also expressed by immune cells. S1P receptors couple to different heterotrimeric G proteins including G alpha i, q, and 12/13, and elicit cellular signalling events by activating the small GTPases Rac and Rho and protein kinases Akt, ERK, and JNK, and by inducing cellular calcium flux and inhibiting cAMP accumulation, amongst others. S1P is the exit signal for lymphocytes leaving lymphoid organs and present in blood and lymph at high nanomolar concentrations due to the S1P-producing activity of sphingosine kinases (SK). The S1P-degrading enzyme S1P-lyase maintains low amounts of S1P in lymphoid organs. Disrupting this concentration difference by S1P receptor agonists and antagonists like FTY720, SEW2871, and VPC23019, by an anti-S1P antibody, or by inhibiting the S1P-lyase has therapeutic potential for autoimmune diseases like multiple sclerosis (MS) and rheumatoid arthritis and for many other disorders like cancer, fibrosis, inflammation, macular degeneration, diabetic retinopathy, and glaucoma. This report aims to provide a brief overview of concepts, approaches, pharmaceutical compounds, and targets that are currently used to modulate S1P-driven immune functions.

  4. Exit Strategies: S1P Signaling and T Cell Migration.

    PubMed

    Baeyens, Audrey; Fang, Victoria; Chen, Cynthia; Schwab, Susan R

    2015-12-01

    Whereas the role of sphingosine 1-phosphate receptor 1 (S1PR1) in T cell egress and the regulation of S1P gradients between lymphoid organs and circulatory fluids in homeostasis are increasingly well understood, much remains to be learned about S1P signaling and distribution during an immune response. Recent data suggest that the role of S1PR1 in directing cells from tissues into circulatory fluids is reprised again and again, particularly in guiding activated T cells from non-lymphoid tissues into lymphatics. Conversely, S1P receptor 2 (S1PR2), which antagonizes migration towards chemokines, confines cells within tissues. Here we review the current understanding of the roles of S1P signaling in activated T cell migration. In this context, we outline open questions, particularly regarding the shape of S1P gradients in different tissues in homeostasis and inflammation, and discuss recent strategies to measure S1P.

  5. S1P and the birth of platelets.

    PubMed

    Hla, Timothy; Galvani, Sylvain; Rafii, Shahin; Nachman, Ralph

    2012-11-19

    Recent work has highlighted the multitude of biological functions of sphingosine 1-phosphate (S1P), which include roles in hematopoietic cell trafficking, organization of immune organs, vascular development, and neuroinflammation. Indeed, a functional antagonist of S1P(1) receptor, FTY720/Gilenya, has entered the clinic as a novel therapeutic for multiple sclerosis. In this issue of the JEM, Zhang et al. highlight yet another function of this lipid mediator: thrombopoiesis. The S1P(1) receptor is required for the growth of proplatelet strings in the bloodstream and the shedding of platelets into the circulation. Notably, the sharp gradient of S1P between blood and the interstitial fluids seems to be essential to ensure the production of platelets, and S1P appears to cooperate with the CXCL12-CXCR4 axis. Pharmacologic modulation of the S1P(1) receptor altered circulating platelet numbers acutely, suggesting a potential therapeutic strategy for controlling thrombocytopenic states. However, the S1P(4) receptor may also regulate thrombopoiesis during stress-induced accelerated platelet production. This work reveals a novel physiological action of the S1P/S1P(1) duet that could potentially be harnessed for clinical translation.

  6. Functional Characterization of Candida albicans ABC Transporter Cdr1p

    PubMed Central

    Shukla, Suneet; Saini, Preeti; Smriti; Jha, Sudhakar; Ambudkar, Suresh V.; Prasad, Rajendra

    2003-01-01

    In view of the importance of Candida drug resistance protein (Cdr1p) in azole resistance, we have characterized it by overexpressing it as a green fluorescent protein (GFP)-tagged fusion protein (Cdr1p-GFP). The overexpressed Cdr1p-GFP in Saccharomyces cerevisiae is shown to be specifically labeled with the photoaffinity analogs iodoarylazidoprazosin (IAAP) and azidopine, which have been used to characterize the drug-binding sites on mammalian drug-transporting P-glycoproteins. While nystatin could compete for the binding of IAAP, miconazole specifically competed for azidopine binding, suggesting that IAAP and azidopine bind to separate sites on Cdr1p. Cdr1p was subjected to site-directed mutational analysis. Among many mutant variants of Cdr1p, the phenotypes of F774A and ΔF774 were particularly interesting. The analysis of GFP-tagged mutant variants of Cdr1p revealed that a conserved F774, in predicted transmembrane segment 6, when changed to alanine showed increased binding of both photoaffinity analogues, while its deletion (ΔF774), as revealed by confocal microscopic analyses, led to mislocalization of the protein. The mislocalized ΔF774 mutant Cdr1p could be rescued to the plasma membrane as a functional transporter by growth in the presence of a Cdr1p substrate, cycloheximide. Our data for the first time show that the drug substrate-binding sites of Cdr1p exhibit striking similarities with those of mammalian drug-transporting P-glycoproteins and despite differences in topological organization, the transmembrane segment 6 in Cdr1p is also a major contributor to drug substrate-binding site(s). PMID:14665469

  7. The caspase-generated cleavage product of Ets-1 p51 and Ets-1 p27, Cp17, induces apoptosis.

    PubMed

    Choul-Li, Souhaila; Tulasne, David; Aumercier, Marc

    2016-11-04

    The transcription factor Ets-1 is involved in various physiological processes and invasive pathologies. Human Ets-1 exists under three isoforms: p51, the predominant full-length isoform, p42 and p27, shorter alternatively spliced isoforms. We have previously demonstrated that Ets-1 p51, but not the spliced variant Ets-1 p42, is processed by caspases in vitro and during apoptosis. However, the caspase cleavage of the second spliced variant Ets-1 p27 remains to investigate. In the present study, we demonstrate that Ets-1 p27 is a cleavage substrate of caspases. We show that Ets-1 p27 is processed in vitro by caspase-3, resulting in three C-terminal fragments Cp20, Cp17 and Cp14. Similarly, Ets-1 p27 was cleaved during apoptotic cell death induced by anisomycin, producing fragments consistent with those observed in in vitro cleavage assay. These fragments are generated by cleavage at three sites located in the exon VII-encoded region of Ets-1 p27. As a functional consequences, Cp17 fragment, the major cleavage product generated during apoptosis, induced itself apoptosis when transfected into cells. Our results show that Ets-1 p27 is cleaved in the same manner as Ets-1 p51 within the exon VII-encoded region, thus generating a stable C-terminal fragment that induces cell death by initiating apoptosis.

  8. Rat1p maintains RNA polymerase II CTD phosphorylation balance

    PubMed Central

    Jimeno-González, Silvia; Schmid, Manfred; Malagon, Francisco; Haaning, Line Lindegaard; Jensen, Torben Heick

    2014-01-01

    In S. cerevisiae, the 5′-3′ exonuclease Rat1p partakes in transcription termination. Although Rat1p-mediated RNA degradation has been suggested to play a role for this activity, the exact mechanisms by which Rat1p helps release RNA polymerase II (RNAPII) from the DNA template are poorly understood. Here we describe a function of Rat1p in regulating phosphorylation levels of the C-terminal domain (CTD) of the largest RNAPII subunit, Rpb1p, during transcription elongation. The rat1-1 mutant exhibits highly elevated levels of CTD phosphorylation as well as RNAPII distribution and transcription termination defects. These phenotypes are all rescued by overexpression of the CTD phosphatase Fcp1p, suggesting a functional relationship between the absence of Rat1p activity, elevated CTD phosphorylation, and transcription defects. We also demonstrate that rat1-1 cells display increased RNAPII transcription kinetics, a feature that may contribute to the cellular phenotypes of the mutant. Consistently, the rat1-1 allele is synthetic lethal with the rpb1-E1103G mutation, causing increased RNAPII speed, and is suppressed by the rpb2-10 mutation, causing slowed transcription. Thus, Rat1p plays more complex roles in controlling transcription than previously thought. PMID:24501251

  9. Sphingosine-1-Phosphate (S1P) and S1P Signaling Pathway: Therapeutic Targets in Autoimmunity and Inflammation.

    PubMed

    Tsai, Hsing-Chuan; Han, May H

    2016-07-01

    Sphingosine-1-phosphate (S1P) and S1P receptors (S1PR) are ubiquitously expressed. S1P-S1PR signaling has been well characterized in immune trafficking and activation in innate and adaptive immune systems. However, the full extent of its involvement in the pathogenesis of autoimmune diseases is not well understood. FTY720 (fingolimod), a non-selective S1PR modulator, significantly decreased annualized relapse rates in relapsing-remitting multiple sclerosis (MS). FTY720, which primarily targets S1P receptor 1 as a functional antagonist, arrests lymphocyte egress from secondary lymphoid tissues and reduces neuroinflammation in the central nervous system (CNS). Recent studies suggest that FTY720 also decreases astrogliosis and promotes oligodendrocyte differentiation within the CNS and may have therapeutic benefit to prevent brain atrophy. Since S1P signaling is involved in multiple immune functions, therapies targeting S1P axis may be applicable to treat autoimmune diseases other than MS. Currently, over a dozen selective S1PR and S1P pathway modulators with potentially superior therapeutic efficacy and better side-effect profiles are in the pipeline of drug development. Furthermore, newly characterized molecules such as apolipoprotein M (ApoM) (S1P chaperon) and SPNS2 (S1P transporter) are also potential targets for treatment of autoimmune diseases. Finally, the application of therapies targeting S1P and S1P signaling pathways may be expanded to treat several other immune-mediated disorders (such as post-infectious diseases, post-stroke and post-stroke dementia) and inflammatory conditions beyond their application in primary autoimmune diseases.

  10. Yhh1p/Cft1p directly links poly(A) site recognition and RNA polymerase II transcription termination.

    PubMed

    Dichtl, Bernhard; Blank, Diana; Sadowski, Martin; Hübner, Wolfgang; Weiser, Stefan; Keller, Walter

    2002-08-01

    RNA polymerase II (pol II) transcription termination requires co-transcriptional recognition of a functional polyadenylation signal, but the molecular mechanisms that transduce this signal to pol II remain unclear. We show that Yhh1p/Cft1p, the yeast homologue of the mammalian AAUAAA interacting protein CPSF 160, is an RNA-binding protein and provide evidence that it participates in poly(A) site recognition. Interestingly, RNA binding is mediated by a central domain composed of predicted beta-propeller-forming repeats, which occurs in proteins of diverse cellular functions. We also found that Yhh1p/Cft1p bound specifically to the phosphorylated C-terminal domain (CTD) of pol II in vitro and in a two-hybrid test in vivo. Furthermore, transcriptional run-on analysis demonstrated that yhh1 mutants were defective in transcription termination, suggesting that Yhh1p/Cft1p functions in the coupling of transcription and 3'-end formation. We propose that direct interactions of Yhh1p/Cft1p with both the RNA transcript and the CTD are required to communicate poly(A) site recognition to elongating pol II to initiate transcription termination.

  11. Dynamin-related proteins Vps1p and Dnm1p control peroxisome abundance in Saccharomyces cerevisiae.

    PubMed

    Kuravi, Kasinath; Nagotu, Shirisha; Krikken, Arjen M; Sjollema, Klaas; Deckers, Markus; Erdmann, Ralf; Veenhuis, Marten; van der Klei, Ida J

    2006-10-01

    Saccharomyces cerevisiae contains three dynamin-related-proteins, Vps1p, Dnm1p and Mgm1p. Previous data from glucose-grown VPS1 and DNM1 null mutants suggested that Vps1p, but not Dnm1p, plays a role in regulating peroxisome abundance. Here we show that deletion of DNM1 also results in reduction of peroxisome numbers. This was not observed in glucose-grown dnm1 cells, but was evident in cells grown in the presence of oleate. Similar observations were made in cells lacking Fis1p, a protein involved in Dnm1p function. Fluorescence microscopy of cells producing Dnm1-GFP or GFP-Fis1p demonstrated that both proteins had a dual localization on mitochondria and peroxisomes. Quantitative analysis revealed a greater reduction in peroxisome number in oleate-induced vps1 cells relative to dnm1 or fis1 cells. A significant fraction of oleate-induced vps1 cells still contained two or more peroxisomes. Conversely, almost all cells of a dnm1 vps1 double-deletion strain contained only one, enlarged peroxisome. This suggests that deletion of DNM1 reinforces the vps1 peroxisome phenotype. Time-lapse imaging indicated that during budding of dnm1 vps1 cells, the single peroxisome present in the mother cell formed long protrusions into the developing bud. This organelle divided at a very late stage of the budding process, possibly during cytokinesis.

  12. A role for Yip1p in COPII vesicle biogenesis

    PubMed Central

    Heidtman, Matthew; Chen, Catherine Z.; Collins, Ruth N.; Barlowe, Charles

    2003-01-01

    Yeast Ypt1p-interacting protein (Yip1p) belongs to a conserved family of transmembrane proteins that interact with Rab GTPases. We encountered Yip1p as a constituent of ER-derived transport vesicles, leading us to hypothesize a direct role for this protein in transport through the early secretory pathway. Using a cell-free assay that recapitulates protein transport from the ER to the Golgi complex, we find that affinity-purified antibodies directed against the hydrophilic amino terminus of Yip1p potently inhibit transport. Surprisingly, inhibition is specific to the COPII-dependent budding stage. In support of this in vitro observation, strains bearing the temperature-sensitive yip1-4 allele accumulate ER membranes at a nonpermissive temperature, with no apparent accumulation of vesicle intermediates. Genetic interaction analyses of the yip1-4 mutation corroborate a function in ER budding. Finally, ordering experiments show that preincubation of ER membranes with COPII proteins decreases sensitivity to anti-Yip1p antibodies, indicating an early requirement for Yip1p in vesicle formation. We propose that Yip1p has a previously unappreciated role in COPII vesicle biogenesis. PMID:14557247

  13. A role for Yip1p in COPII vesicle biogenesis.

    PubMed

    Heidtman, Matthew; Chen, Catherine Z; Collins, Ruth N; Barlowe, Charles

    2003-10-13

    Yeast Ypt1p-interacting protein (Yip1p) belongs to a conserved family of transmembrane proteins that interact with Rab GTPases. We encountered Yip1p as a constituent of ER-derived transport vesicles, leading us to hypothesize a direct role for this protein in transport through the early secretory pathway. Using a cell-free assay that recapitulates protein transport from the ER to the Golgi complex, we find that affinity-purified antibodies directed against the hydrophilic amino terminus of Yip1p potently inhibit transport. Surprisingly, inhibition is specific to the COPII-dependent budding stage. In support of this in vitro observation, strains bearing the temperature-sensitive yip1-4 allele accumulate ER membranes at a nonpermissive temperature, with no apparent accumulation of vesicle intermediates. Genetic interaction analyses of the yip1-4 mutation corroborate a function in ER budding. Finally, ordering experiments show that preincubation of ER membranes with COPII proteins decreases sensitivity to anti-Yip1p antibodies, indicating an early requirement for Yip1p in vesicle formation. We propose that Yip1p has a previously unappreciated role in COPII vesicle biogenesis.

  14. Structural Characterization of Tip20p and Dsl1p, Subunits of the Dsl1p Vesicle Tethering Complex

    SciTech Connect

    Tripathi, A.; Ren, Y; Jeffrey, P; Hughson, F

    2009-01-01

    Multisubunit tethering complexes are essential for intracellular trafficking and have been proposed to mediate the initial interaction between vesicles and the membranes with which they fuse. Here we report initial structural characterization of the Dsl1p complex, whose three subunits are essential for trafficking from the Golgi apparatus to the endoplasmic reticulum (ER). Crystal structures reveal that two of the three subunits, Tip20p and Dsl1p, resemble known subunits of the exocyst complex, establishing a structural connection among several multisubunit tethering complexes and implying that many of their subunits are derived from a common progenitor. We show, moreover, that Tip20p and Dsl1p interact directly via N-terminal alpha-helices. Finally, we establish that different Dsl1p complex subunits bind independently to different ER SNARE proteins. Our results map out two alternative protein-interaction networks capable of tethering COPI-coated vesicles, via the Dsl1p complex, to ER membranes.

  15. Growth patterns of patients with 1p36 deletion syndrome.

    PubMed

    Sangu, Noriko; Shimojima, Keiko; Shimada, Shino; Ando, Tomohiro; Yamamoto, Toshiyuki

    2014-05-01

    1p36 deletion syndrome is one of the most common subtelomeric deletion syndromes. Obesity is frequently observed in patients with this syndrome. Thus, it is important to evaluate the growth status of an individual patient. For this purpose, we accumulated recorded growth data from 44 patients with this syndrome and investigated the growth patterns of patients. Most of the patients showed weight parameters within normal limits, whereas a few of these patients showed intrauterine growth delay and microcephaly. The length of the patients after birth was under the 50th centile in most patients. Many patients showed poor weight gain after birth, and only two female patients were overweight. These findings indicate two different phenotypes of the 1p36 deletion syndrome. The overweight patients with 1p36 deletion started excessive weight gain after two years of life. This characteristic of the patients with 1p36 deletion syndrome is similar to Prader-Willi syndrome.

  16. 1p36 tumor suppression--a matter of dosage?

    PubMed

    Henrich, Kai-Oliver; Schwab, Manfred; Westermann, Frank

    2012-12-01

    A broad range of human malignancies is associated with nonrandom 1p36 deletions, suggesting the existence of tumor suppressors encoded in this region. Evidence for tumor-specific inactivation of 1p36 genes in the classic "two-hit" manner is scarce; however, many tumor suppressors do not require complete inactivation but contribute to tumorigenesis by partial impairment. We discuss recent data derived from both human tumors and functional cancer models indicating that the 1p36 genes CHD5, CAMTA1, KIF1B, CASZ1, and miR-34a contribute to cancer development when reduced in dosage by genomic copy number loss or other mechanisms. We explore potential interactions among these candidates and propose a model where heterozygous 1p36 deletion impairs oncosuppressive pathways via simultaneous downregulation of several dosage-dependent tumor suppressor genes.

  17. Sphingosine-1-Phosphate (S1P) Lyase Inhibition Causes Increased Cardiac S1P Levels and Bradycardia in Rats.

    PubMed

    Harris, Christopher M; Mittelstadt, Scott; Banfor, Patricia; Bousquet, Peter; Duignan, David B; Gintant, Gary; Hart, Michelle; Kim, Youngjae; Segreti, Jason

    2016-10-01

    Inhibition of the sphingosine-1-phosphate (S1P)-catabolizing enzyme S1P lyase (S1PL) elevates the native ligand of S1P receptors and provides an alternative mechanism for immune suppression to synthetic S1P receptor agonists. S1PL inhibition is reported to preferentially elevate S1P in lymphoid organs. Tissue selectivity could potentially differentiate S1PL inhibitors from S1P receptor agonists, the use of which also results in bradycardia, atrioventricular block, and hypertension. But it is unknown if S1PL inhibition would also modulate cardiac S1P levels or cardiovascular function. The S1PL inhibitor 6-[(2R)-4-(4-benzyl-7-chlorophthalazin-1-yl)-2-methylpiperazin-1-yl]pyridine-3-carbonitrile was used to determine the relationship in rats between drug concentration, S1P levels in select tissues, and circulating lymphocytes. Repeated oral doses of the S1PL inhibitor fully depleted circulating lymphocytes after 3 to 4 days of treatment in rats. Full lymphopenia corresponded to increased levels of S1P of 100- to 1000-fold in lymph nodes, 3-fold in blood (but with no change in plasma), and 9-fold in cardiac tissue. Repeated oral dosing of the S1PL inhibitor in telemeterized, conscious rats resulted in significant bradycardia within 48 hours of drug treatment, comparable in magnitude to the bradycardia induced by 3 mg/kg fingolimod. These results suggest that S1PL inhibition modulates cardiac function and does not provide immune suppression with an improved cardiovascular safety profile over fingolimod in rats.

  18. Measurement of CA1P and CA in leaves

    SciTech Connect

    Moore, B.d.; Kobza, J.; Seemann, J.R. )

    1990-05-01

    Carboxyarabinitol-1-phosphate (CA1P) and carboxyarabinitol (CA) were assayed in leaves by isotope dilution. {sup 14}C-labeled standards were synthesized from (2-{sup 14}C) CABP using acid (CA1P) or alkaline (CA) phosphatase. Either was added to boiling 80% EtOH along with liquid N{sub 2}-killed leaves. Each was largely purified by anion exchange chromatography. CA1P samples were subjected to 2D-TLE/TLC. The specific activity of the {sup 14}C-containing spot was measured using alkaline phosphatase. CA samples were run on an HPLC and the specific activity was determined using a UV monitor and a flow-through radioisotope detector. In 3 of the tested species, light/dark amount of CA1P (nmol/mg Chl) were kidney bean, 0.7/67; sugar beet, 0.8/33; and Alocasia, 0/3.4. Light/dark CA levels (nmol/mg Chl) in these respective species were 897/653, 3.2/3.5, and 5.7/4.6. These results support the hypothesis that CA is a product of CA1P metabolism in vivo under high light, but also indicate that CA is not the only intermediate involved in CA1P synthesis under low light/dark conditions.

  19. AtPng1p. The first plant transglutaminase.

    PubMed

    Della Mea, Massimiliano; Caparrós-Ruiz, David; Claparols, Inmaculada; Serafini-Fracassini, Donatella; Rigau, Joan

    2004-08-01

    Studies have revealed in plant chloroplasts, mitochondria, cell walls, and cytoplasm the existence of transglutaminase (TGase) activities, similar to those known in animals and prokaryotes having mainly structural roles, but no protein has been associated to this type of activity in plants. A recent computational analysis has shown in Arabidopsis the presence of a gene, AtPng1p, which encodes a putative N-glycanase. AtPng1p contains the Cys-His-Asp triad present in the TGase catalytic domain. AtPng1p is a single gene expressed ubiquitously in the plant but at low levels in all light-assayed conditions. The recombinant AtPng1p protein could be immuno-detected using animal TGase antibodies. Furthermore, western-blot analysis using antibodies raised against the recombinant AtPng1p protein have lead to its detection in microsomal fraction. The purified protein links polyamines-spermine (Spm) > spermidine (Spd) > putrescine (Put)-and biotin-cadaverine to dimethylcasein in a calcium-dependent manner. Analyses of the gamma-glutamyl-derivatives revealed that the formation of covalent linkages between proteins and polyamines occurs via the transamidation of gamma-glutamyl residues of the substrate, confirming that the AtPng1p gene product acts as a TGase. The Ca(2+)- and GTP-dependent cross-linking activity of the AtPng1p protein can be visualized by the polymerization of bovine serum albumine, obtained, like the commercial TGase, at basic pH and in the presence of dithiotreitol. To our knowledge, this is the first reported plant protein, characterized at molecular level, showing TGase activity, as all its parameters analyzed so far agree with those typically exhibited by the animal TGases.

  20. Unfolded protein response regulates yeast small GTPase Arl1p activation at late Golgi via phosphorylation of Arf GEF Syt1p

    PubMed Central

    Hsu, Jia-Wei; Tang, Pei-Hua; Wang, I-Hao; Liu, Chia-Lun; Chen, Wen-Hui; Tsai, Pei-Chin; Chen, Kuan-Yu; Chen, Kuan-Jung; Yu, Chia-Jung

    2016-01-01

    ADP ribosylation factor (Arf) GTPases are key regulators of membrane traffic at the Golgi complex. In yeast, Arf guanine nucleotide-exchange factor (GEF) Syt1p activates Arf-like protein Arl1p, which was accompanied by accumulation of golgin Imh1p at late Golgi, but whether and how this function of Syt1p is regulated remains unclear. Here, we report that the inositol-requiring kinase 1 (Ire1p)-mediated unfolded protein response (UPR) modulated Arl1p activation at late Golgi. Arl1p activation was dependent on both kinase and endo-RNase activities of Ire1p. Moreover, constitutively active transcription factor Hac1p restored the Golgi localization of Arl1p and Imh1p in IRE1-deleted cells. Elucidating the mechanism of Ire1p–Hac1p axis actions, we found that it regulated phosphorylation of Syt1p, which enhances Arl1p activation, recruitment of Imh1p to the Golgi, and Syt1p interaction with Arl1p. Consistent with these findings, the induction of UPR by tunicamycin treatment increases phosphorylation of Syt1p, resulting in Arl1p activation. Thus, these findings clarify how the UPR influences the roles of Syt1p, Arl1p, and Imh1p in Golgi transport. PMID:26966233

  1. Del 1p36 syndrome: a newly emerging clinical entity.

    PubMed

    Battaglia, Agatino

    2005-08-01

    Monosomy 1p36 is a recently delineated contiguous gene syndrome, which is now considered to be the most common subtelomeric microdeletion syndrome. From the recent literature it appears as if 1p36 deletions account for 0.5-1.2% of idiopathic mental retardation. The deletions can be detected by high resolution cytogenetic studies in a minority of patients, and fluorescence in situ hybridisation (FISH) is required in most. The deletions' parent of origin seems still unclear, although in one large series it was shown to be maternal. 1p36 deletion syndrome is characterized by distinct craniofacial features, associated with developmental delay/mental retardation, hypotonia, muscle hypotrophy, seizures, brain abnormalities, and heart defects. To help child neurologists and other professionals in the recognition of this emerging and common chromosomal syndrome, we have reviewed published articles on patients with this deletion.

  2. [An updated review of 1p36 deletion (monosomy) syndrome].

    PubMed

    Bello, Sabina; Rodríguez-Moreno, Antonio

    The Monosomy 1p36 deletion syndrome is part of the group of diseases known as Rare Diseases. The objective of the present work is to review the characteristics of Monosomy 1p36 deletion syndrome. The monosomy 1p36 deletion syndrome phenotype includes: dysmorphic craniofacial features; large anterior fontanelle, unibrow, deep-set eyes, epicanthus, wide nasal root/bridge, mandible hypoplasia, abnormal location of the pinna, philtrum and pointed chin; neurological alterations: seizures and hydrocephalus (in some cases). Cerebral malformations: ventricular hypertrophy, increased subarachnoid space, morphological alterations of corpus callosum, cortical atrophy, delays in myelinisation, periventricular leukomalacia and periventricular heterotopia. These alterations produce intellectual disability and delays in motor growth, communication skills, language, social and adaptive behaviour. It is Hearing and vision impairments are also observed in subjects with this syndrome, as well as alterations of cardiac, endocrine and urinary systems and alterations at skin and skeletal level.

  3. Physical map of 1p36, placement of breakpoints in monosomy 1p36, and clinical characterization of the syndrome.

    PubMed

    Heilstedt, Heidi A; Ballif, Blake C; Howard, Leslie A; Lewis, Richard A; Stal, Samuel; Kashork, Catherine D; Bacino, Carlos A; Shapira, Stuart K; Shaffer, Lisa G

    2003-05-01

    Monosomy 1p36 is the most common terminal deletion syndrome. This contiguous gene deletion syndrome is presumably caused by haploinsufficiency of a number of genes. We have constructed a contig of overlapping large-insert clones for the most distal 10.5 Mb of 1p36, evaluated the deletion sizes in 61 subjects with monosomy 1p36 from 60 families, and created a natural deletion panel. We found pure terminal deletions, interstitial deletions, derivative chromosomes, and more complex rearrangements. Breakpoints were "binned" into 0.5-Mb regions. Analyses revealed some clustering of breakpoints but no single common breakpoint. Determination of the parental origin showed that 60% of de novo 1p36 terminal deletions arose from the maternally inherited chromosome. Of the 61 subjects, 30 were examined systematically through a protocol at the Texas Children's Hospital General Clinical Research Center. Specifically, we report hearing evaluations, palatal and ophthalmological examinations, echocardiograms, neurological assessments, and thyroid function tests. To our knowledge, this systematic molecular and clinical characterization of monosomy 1p36 is the largest and most comprehensive study of this deletion syndrome to date. Many cytogenetically visible, apparent terminal deletions are more complex than anticipated by cytogenetics, as revealed at the molecular level by our study. Our clinical findings allow for the more accurate recognition of the syndrome and for proper medical evaluation.

  4. Late-stage optimization of a tercyclic class of S1P3-sparing, S1P1 receptor agonists.

    PubMed

    Horan, Joshua C; Kuzmich, Daniel; Liu, Pingrong; DiSalvo, Darren; Lord, John; Mao, Can; Hopkins, Tamara D; Yu, Hui; Harcken, Christian; Betageri, Raj; Hill-Drzewi, Melissa; Patenaude, Lori; Patel, Monica; Fletcher, Kimberly; Terenzzio, Donna; Linehan, Brian; Xia, Heather; Patel, Mita; Studwell, Debbie; Miller, Craig; Hickey, Eugene; Levin, Jeremy I; Smith, Dustin; Kemper, Raymond A; Modis, Louise K; Bannen, Lynne C; Chan, Diva S; Mac, Morrison B; Ng, Stephanie; Wang, Yong; Xu, Wei; Lemieux, René M

    2016-01-15

    Poor solubility and cationic amphiphilic drug-likeness were liabilities identified for a lead series of S1P3-sparing, S1P1 agonists originally developed from a high-throughput screening campaign. This work describes the subsequent optimization of these leads by balancing potency, selectivity, solubility and overall molecular charge. Focused SAR studies revealed favorable structural modifications that, when combined, produced compounds with overall balanced profiles. The low brain exposure observed in rat suggests that these compounds would be best suited for the potential treatment of peripheral autoimmune disorders.

  5. RNA binding protein Pub1p regulates glycerol production and stress tolerance by controlling Gpd1p activity during winemaking.

    PubMed

    Orozco, Helena; Sepúlveda, Ana; Picazo, Cecilia; Matallana, Emilia; Aranda, Agustín

    2016-06-01

    Glycerol is a key yeast metabolite in winemaking because it contributes to improve the organoleptic properties of wine. It is also a cellular protective molecule that enhances the tolerance of yeasts to osmotic stress and promotes longevity. Thus, its production increases by genetic manipulation, which is of biotechnological and basic interest. Glycerol is produced by diverting glycolytic glyceraldehyde-3-phosphate through the action of glycerol-3-phosphate dehydrogenase (coded by genes GPD1 and GPD2). Here, we demonstrate that RNA-binding protein Pub1p regulates glycerol production by controlling Gpd1p activity. Its deletion does not alter GPD1 mRNA levels, but protein levels and enzymatic activity increase, which explains the higher intracellular glycerol concentration and greater tolerance to osmotic stress of the pub1∆ mutant. PUB1 deletion also enhances the activity of nicotinamidase, a longevity-promoting enzyme. Both enzymatic activities are partially located in peroxisomes, and we detected peroxisome formation during wine fermentation. The role of Pub1p in life span control depends on nutrient conditions and is related with the TOR pathway, and a major connection between RNA metabolism and the nutrient signaling response is established.

  6. Highly selective and potent agonists of sphingosine-1-phosphate 1 (S1P1) receptor.

    PubMed

    Vachal, Petr; Toth, Leslie M; Hale, Jeffrey J; Yan, Lin; Mills, Sander G; Chrebet, Gary L; Koehane, Carol A; Hajdu, Richard; Milligan, James A; Rosenbach, Mark J; Mandala, Suzanne

    2006-07-15

    Novel series of sphingosine-1-phosphate (S1P) receptor agonists were developed through a systematic SAR aimed to achieve high selectivity for a single member of the S1P family of receptors, S1P1. The optimized structure represents a highly S1P1-selective and efficacious agonist: S1P1/S1P2, S1P1/S1P3, S1P1/S1P4>10,000-fold, S1P1/S1P5>600-fold, while EC50 (S1P1) <0.2 nM. In vivo experiments are consistent with S1P1 receptor agonism alone being sufficient for achieving desired lymphocyte-lowering effect.

  7. Molecular Oxygen in Oort Cloud Comet 1P/Halley

    NASA Astrophysics Data System (ADS)

    Rubin, M.; Altwegg, K.; van Dishoeck, E. F.; Schwehm, G.

    2015-12-01

    Recently, the ROSINA mass spectrometer suite on board the European Space Agency's Rosetta spacecraft discovered an abundant amount of molecular oxygen, O2, in the coma of Jupiter family comet 67P/Churyumov-Gerasimenko of O2/H2O = 3.80 ± 0.85%. It could be shown that O2 is indeed a parent species and that the derived abundances point to a primordial origin. Crucial questions are whether the O2 abundance is peculiar to comet 67P/Churyumov-Gerasimenko or Jupiter family comets in general, and also whether Oort cloud comets such as comet 1P/Halley contain similar amounts of molecular oxygen. We investigated mass spectra obtained by the Neutral Mass Spectrometer instrument during the flyby by the European Space Agency's Giotto probe of comet 1P/Halley. Our investigation indicates that a production rate of O2 of 3.7 ± 1.7% with respect to water is indeed compatible with the obtained Halley data and therefore that O2 might be a rather common and abundant parent species.

  8. MOLECULAR OXYGEN IN OORT CLOUD COMET 1P/HALLEY

    SciTech Connect

    Rubin, M.; Altwegg, K.; Dishoeck, E. F. van; Schwehm, G.

    2015-12-10

    Recently, the ROSINA mass spectrometer suite on board the European Space Agency's Rosetta spacecraft discovered an abundant amount of molecular oxygen, O{sub 2}, in the coma of Jupiter family comet 67P/Churyumov–Gerasimenko of O{sub 2}/H{sub 2}O = 3.80 ± 0.85%. It could be shown that O{sub 2} is indeed a parent species and that the derived abundances point to a primordial origin. Crucial questions are whether the O{sub 2} abundance is peculiar to comet 67P/Churyumov–Gerasimenko or Jupiter family comets in general, and also whether Oort cloud comets such as comet 1P/Halley contain similar amounts of molecular oxygen. We investigated mass spectra obtained by the Neutral Mass Spectrometer instrument during the flyby by the European Space Agency's Giotto probe of comet 1P/Halley. Our investigation indicates that a production rate of O{sub 2} of 3.7 ± 1.7% with respect to water is indeed compatible with the obtained Halley data and therefore that O{sub 2} might be a rather common and abundant parent species.

  9. Elastic Pitch Beam Tail Rotor for LOH

    DTIC Science & Technology

    1976-07-01

    15 - 5PH stainless steel, Condition H 1025 and Fe : 18, 00 psi, 1105 Fa = 18,000(1- 155,000 Fa = 17,872 psi The margin of safety in fatigue is then Fa...bladed elastomeric hub assembly consists of a 15 - 5PH stainless steel hub, a 2024-T4 aluminum hub cap, and a 15 - 5PH stainless steel hub spacer. Current OH...through the elastomeric pivot mounted in the 15 - 5PH stainless steel bearing housing attached to the inboard fitting of the airfoil section, The

  10. Sphingosine 1-phosphate (S1P)/S1P receptor 1 signaling regulates receptor activator of NF-{kappa}B ligand (RANKL) expression in rheumatoid arthritis

    SciTech Connect

    Takeshita, Harunori; Kitano, Masayasu; Iwasaki, Tsuyoshi; Kitano, Sachie; Tsunemi, Sachi; Sato, Chieri; Sekiguchi, Masahiro; Azuma, Naoto; Miyazawa, Keiji; Hla, Timothy; Sano, Hajime

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer MH7A cells and CD4{sup +} T cells expressed S1P1 and RANKL. Black-Right-Pointing-Pointer S1P increased RANKL expression in MH7A cells and CD4{sup +} T cells. Black-Right-Pointing-Pointer The effect of S1P in MH7A cells was inhibited by specific Gi/Go inhibitors. -- Abstract: Sphingosine 1-phosphate (S1P)/S1P receptor 1 (S1P1) signaling plays an important role in synovial cell proliferation and inflammatory gene expression by rheumatoid arthritis (RA) synoviocytes. The purpose of this study is to clarify the role of S1P/S1P1 signaling in the expression of receptor activator of NF-{kappa}B ligand (RANKL) in RA synoviocytes and CD4{sup +} T cells. We demonstrated MH7A cells, a human RA synovial cell line, and CD4{sup +} T cells expressed S1P1 and RANKL. Surprisingly, S1P increased RANKL expression in MH7A cells and CD4{sup +} T cells in a dose-dependent manner. Moreover, S1P enhanced RANKL expression induced by stimulation with TNF-{alpha} in MH7A cells and CD4{sup +} T cells. These effects of S1P in MH7A cells were inhibited by pretreatment with PTX, a specific Gi/Go inhibitor. These findings suggest that S1P/S1P1 signaling may play an important role in RANKL expression by MH7A cells and CD4{sup +} T cells. S1P/S1P1 signaling of RA synoviocytes is closely connected with synovial hyperplasia, inflammation, and RANKL-induced osteoclastogenesis in RA. Thus, regulation of S1P/S1P1 signaling may become a novel therapeutic target for RA.

  11. Transcriptional Auto-Regulation of RUNX1 P1 Promoter.

    PubMed

    Martinez, Milka; Hinojosa, Marcela; Trombly, Daniel; Morin, Violeta; Stein, Janet; Stein, Gary; Javed, Amjad; Gutierrez, Soraya E

    2016-01-01

    RUNX1 a member of the family of runt related transcription factors (RUNX), is essential for hematopoiesis. The expression of RUNX1 gene is controlled by two promoters; the distal P1 promoter and the proximal P2 promoter. Several isoforms of RUNX1 mRNA are generated through the use of both promoters and alternative splicing. These isoforms not only differs in their temporal expression pattern but also exhibit differences in tissue specificity. The RUNX1 isoforms derived from P2 are expressed in a variety of tissues, but expression of P1-derived isoform is restricted to cells of hematopoietic lineage. However, the control of hematopoietic-cell specific expression is poorly understood. Here we report regulation of P1-derived RUNX1 mRNA by RUNX1 protein. In silico analysis of P1 promoter revealed presence of two evolutionary conserved RUNX motifs, 0.6kb upstream of the transcription start site, and three RUNX motifs within 170bp of the 5'UTR. Transcriptional contribution of these RUNX motifs was studied in myeloid and T-cells. RUNX1 genomic fragment containing all sites show very low basal activity in both cell types. Mutation or deletion of RUNX motifs in the UTR enhances basal activity of the RUNX1 promoter. Chromatin immunoprecipitation revealed that RUNX1 protein is recruited to these sites. Overexpression of RUNX1 in non-hematopoietic cells results in a dose dependent activation of the RUNX1 P1 promoter. We also demonstrate that RUNX1 protein regulates transcription of endogenous RUNX1 mRNA in T-cell. Finally we show that SCL transcription factor is recruited to regions containing RUNX motifs in the promoter and the UTR and regulates activity of the RUNX1 P1 promoter in vitro. Thus, multiple lines of evidence show that RUNX1 protein regulates its own gene transcription.

  12. Sphingosine 1-phosphate (S1P) induces COX-2 expression and PGE2 formation via S1P receptor 2 in renal mesangial cells.

    PubMed

    Völzke, Anja; Koch, Alexander; Meyer Zu Heringdorf, Dagmar; Huwiler, Andrea; Pfeilschifter, Josef

    2014-01-01

    Understanding the mechanisms of sphingosine 1-phosphate (S1P)-induced cyclooxygenase (COX)-2 expression and prostaglandin E2 (PGE2) formation in renal mesangial cells may provide potential therapeutic targets to treat inflammatory glomerular diseases. Thus, we evaluated the S1P-dependent signaling mechanisms which are responsible for enhanced COX-2 expression and PGE2 formation in rat mesangial cells under basal conditions. Furthermore, we investigated whether these mechanisms are operative in the presence of angiotensin II (Ang II) and of the pro-inflammatory cytokine interleukin-1β (IL-1β). Treatment of rat and human mesangial cells with S1P led to concentration-dependent enhanced expression of COX-2. Pharmacological and molecular biology approaches revealed that the S1P-dependent increase of COX-2 mRNA and protein expression was mediated via activation of S1P receptor 2 (S1P2). Further, inhibition of Gi and p42/p44 MAPK signaling, both downstream of S1P2, abolished the S1P-induced COX-2 expression. In addition, S1P/S1P2-dependent upregulation of COX-2 led to significantly elevated PGE2 levels, which were further potentiated in the presence of Ang II and IL-1β. A functional consequence downstream of S1P/S1P2 signaling is mesangial cell migration that is stimulated by S1P. Interestingly, inhibition of COX-2 by celecoxib and SC-236 completely abolished the migratory response. Overall, our results demonstrate that extracellular S1P induces COX-2 expression via activation of S1P2 and subsequent Gi and p42/p44 MAPK-dependent signaling in renal mesangial cells leading to enhanced PGE2 formation and cell migration that essentially requires COX-2. Thus, targeting S1P/S1P2 signaling pathways might be a novel strategy to treat renal inflammatory diseases.

  13. Selective coupling of the S1P3 receptor subtype to S1P-mediated RhoA activation and cardioprotection.

    PubMed

    Yung, Bryan S; Brand, Cameron S; Xiang, Sunny Y; Gray, Charles B B; Means, Christopher K; Rosen, Hugh; Chun, Jerold; Purcell, Nicole H; Brown, Joan Heller; Miyamoto, Shigeki

    2017-02-01

    Sphingosine-1-phosphate (S1P), a bioactive lysophospholipid, is generated and released at sites of tissue injury in the heart and can act on S1P1, S1P2, and S1P3 receptor subtypes to affect cardiovascular responses. We established that S1P causes little phosphoinositide hydrolysis and does not induce hypertrophy indicating that it does not cause receptor coupling to Gq. We previously demonstrated that S1P confers cardioprotection against ischemia/reperfusion by activating RhoA and its downstream effector PKD. The S1P receptor subtypes and G proteins that regulate RhoA activation and downstream responses in the heart have not been determined. Using siRNA or pertussis toxin to inhibit different G proteins in NRVMs we established that S1P regulates RhoA activation through Gα13 but not Gα12, Gαq, or Gαi. Knockdown of the three major S1P receptors using siRNA demonstrated a requirement for S1P3 in RhoA activation and subsequent phosphorylation of PKD, and this was confirmed in studies using isolated hearts from S1P3 knockout (KO) mice. S1P treatment reduced infarct size induced by ischemia/reperfusion in Langendorff perfused wild-type (WT) hearts and this protection was abolished in the S1P3 KO mouse heart. CYM-51736, an S1P3-specific agonist, also decreased infarct size after ischemia/reperfusion to a degree similar to that achieved by S1P. The finding that S1P3 receptor- and Gα13-mediated RhoA activation is responsible for protection against ischemia/reperfusion suggests that selective targeting of S1P3 receptors could provide therapeutic benefits in ischemic heart disease.

  14. Yos1p is a novel subunit of the Yip1p-Yif1p complex and is required for transport between the endoplasmic reticulum and the Golgi complex.

    PubMed

    Heidtman, Matthew; Chen, Catherine Z; Collins, Ruth N; Barlowe, Charles

    2005-04-01

    Yeast Yip1p is a member of a conserved family of transmembrane proteins that interact with Rab GTPases. Previous studies also have indicated a role for Yip1p in the biogenesis of endoplasmic reticulum (ER)-derived COPII transport vesicles. In this report, we describe the identification and characterization of the uncharacterized open reading frame YER074W-A as a novel multicopy suppressor of the thermosensitive yip1-4 strain. We have termed this gene Yip One Suppressor 1 (YOS1). Yos1p is essential for growth and for function of the secretory pathway; depletion or inactivation of Yos1p blocks transport between the ER and the Golgi complex. YOS1 encodes an integral membrane protein of 87 amino acids that is conserved in eukaryotes. Yos1p localizes to ER and Golgi membranes and is efficiently packaged into ER-derived COPII transport vesicles. Yos1p associates with Yip1p and Yif1p, indicating Yos1p is a novel subunit of the Yip1p-Yif1p complex.

  15. Regulation of human cerebro-microvascular endothelial baso-lateral adhesion and barrier function by S1P through dual involvement of S1P1 and S1P2 receptors.

    PubMed

    Wiltshire, Rachael; Nelson, Vicky; Kho, Dan Ting; Angel, Catherine E; O'Carroll, Simon J; Graham, E Scott

    2016-01-27

    Herein we show that S1P rapidly and acutely reduces the focal adhesion strength and barrier tightness of brain endothelial cells. xCELLigence biosensor technology was used to measure focal adhesion, which was reduced by S1P acutely and this response was mediated through both S1P1 and S1P2 receptors. S1P increased secretion of several pro-inflammatory mediators from brain endothelial cells. However, the magnitude of this response was small in comparison to that mediated by TNFα or IL-1β. Furthermore, S1P did not significantly increase cell-surface expression of any key cell adhesion molecules involved in leukocyte recruitment, included ICAM-1 and VCAM-1. Finally, we reveal that S1P acutely and dynamically regulates microvascular endothelial barrier tightness in a manner consistent with regulated rapid opening followed by closing and strengthening of the barrier. We hypothesise that the role of the S1P receptors in this process is not to cause barrier dysfunction, but is related to controlled opening of the endothelial junctions. This was revealed using real-time measurement of barrier integrity using ECIS ZΘ TEER technology and endothelial viability using xCELLigence technology. Finally, we show that these responses do not occur simply though the pharmacology of a single S1P receptor but involves coordinated action of S1P1 and S1P2 receptors.

  16. Regulation of human cerebro-microvascular endothelial baso-lateral adhesion and barrier function by S1P through dual involvement of S1P1 and S1P2 receptors

    PubMed Central

    Wiltshire, Rachael; Nelson, Vicky; Kho, Dan Ting; Angel, Catherine E.; O’Carroll, Simon J.; Graham, E. Scott

    2016-01-01

    Herein we show that S1P rapidly and acutely reduces the focal adhesion strength and barrier tightness of brain endothelial cells. xCELLigence biosensor technology was used to measure focal adhesion, which was reduced by S1P acutely and this response was mediated through both S1P1 and S1P2 receptors. S1P increased secretion of several pro-inflammatory mediators from brain endothelial cells. However, the magnitude of this response was small in comparison to that mediated by TNFα or IL-1β. Furthermore, S1P did not significantly increase cell-surface expression of any key cell adhesion molecules involved in leukocyte recruitment, included ICAM-1 and VCAM-1. Finally, we reveal that S1P acutely and dynamically regulates microvascular endothelial barrier tightness in a manner consistent with regulated rapid opening followed by closing and strengthening of the barrier. We hypothesise that the role of the S1P receptors in this process is not to cause barrier dysfunction, but is related to controlled opening of the endothelial junctions. This was revealed using real-time measurement of barrier integrity using ECIS ZΘ TEER technology and endothelial viability using xCELLigence technology. Finally, we show that these responses do not occur simply though the pharmacology of a single S1P receptor but involves coordinated action of S1P1 and S1P2 receptors. PMID:26813587

  17. Copy number neutral loss of heterozygosity at 17p and homozygous mutations of TP53 are associated with complex chromosomal aberrations in patients newly diagnosed with myelodysplastic syndromes.

    PubMed

    Svobodova, Karla; Zemanova, Zuzana; Lhotska, Halka; Novakova, Milena; Podskalska, Lucie; Belickova, Monika; Brezinova, Jana; Sarova, Iveta; Izakova, Silvia; Lizcova, Libuse; Berkova, Adela; Siskova, Magda; Jonasova, Anna; Cermak, Jaroslav; Michalova, Kyra

    2016-03-01

    Complex karyotypes are seen in approximately 20% of patients with myelodysplastic syndromes (MDS) and are associated with a high risk of transformation to acute myeloid leukemia and poor outcomes in patients. Copy number neutral loss of heterozygosity (CN-LOH, i.e., both copies of a chromosomal pair or their parts originate from one parent) might contribute to increased genomic instability in the bone-marrow cells of patients with MDS. The pathological potential of CN-LOH, which arises as a clonal aberration in a proportion of somatic cells, consists of tumor suppressor gene and oncogene homozygous mutations. The aim of our study was to evaluate the frequency of CN-LOH at 17p in bone-marrow cells of newly diagnosed MDS patients with complex chromosomal aberrations and to assess its correlation with mutations in the TP53 gene (17p13.1). CN-LOH was detected in 40 chromosomal regions in 21 (29%) of 72 patients analyzed. The changes in 27 of the 40 regions identified were sporadic. The most common finding was CN-LOH of the short arm of chromosome 17, which was detected in 13 (18%) of 72 patients. A mutational analysis confirmed the homozygous mutation of TP53 in all CN-LOH 17p patients, among which two frameshift mutations are not registered in the International Agency for Research on Cancer TP53 Database. CN-LOH 17p correlated with aggressive disease (median overall survival 4 months) and was strongly associated with a complex karyotype in the cohort studied, which might cause rapid disease progression in high-risk MDS. No other CN-LOH region previously recorded in MDS or AML patients (1p, 4q, 7q, 11q, 13q, 19q, 21q) was detected in our cohort of patients with complex karyotype examined at the diagnosis of MDS. The LOH region appeared to be balanced (i.e., with no DNA copy number change) when examined with conventional and molecular cytogenetic methods. Therefore, a microarray that detects single-nucleotide polymorphisms is an ideal method with which to identify and

  18. S1P lyase in thymic perivascular spaces promotes egress of mature thymocytes via up-regulation of S1P receptor 1.

    PubMed

    Maeda, Yasuhiro; Yagi, Hideki; Takemoto, Kana; Utsumi, Hiroyuki; Fukunari, Atsushi; Sugahara, Kunio; Masuko, Takashi; Chiba, Kenji

    2014-05-01

    Sphingosine 1-phosphate (S1P) and S1P receptor 1 (S1P1) play an important role in the egress of mature CD4 or CD8 single-positive (SP) thymocytes from the thymus. Fingolimod hydrochloride (FTY720), an S1P1 functional antagonist, induced significant accumulation of CD62L(high)CD69(low) mature SP thymocytes in the thymic medulla. Immunohistochemical staining using anti-S1P1 antibody revealed that S1P1 is predominantly expressed on thymocytes in the thymic medulla and is strongly down-regulated even at 3h after FTY720 administration. 2-Acetyl-4-tetrahydroxybutylimidazole (THI), an S1P lyase inhibitor, also induced accumulation of mature SP thymocytes in the thymic medulla with an enlargement of the perivascular spaces (PVS). At 6h after THI administration, S1P1-expressing thymocytes reduced partially as if to form clusters and hardly existed in the proximity of CD31-expressing blood vessels in the thymic medulla, suggesting S1P lyase expression in the cells constructing thymic medullary PVS. To determine the cells expressing S1P lyase in the thymus, we newly established a mAb (YK19-2) specific for mouse S1P lyase. Immunohistochemical staining with YK19-2 revealed that S1P lyase is predominantly expressed in non-lymphoid thymic stromal cells in the thymic medulla. In the thymic medullary PVS, S1P lyase was expressed in ER-TR7-positive cells (reticular fibroblasts and pericytes) and CD31-positive vascular endothelial cells. Our findings suggest that S1P lyase expressed in the thymic medullary PVS keeps the tissue S1P concentration low around the vessels and promotes thymic egress via up-regulation of S1P1.

  19. The effect of the bioactive sphingolipids S1P and C1P on multipotent stromal cells--new opportunities in regenerative medicine.

    PubMed

    Marycz, Krzysztof; Śmieszek, Agnieszka; Jeleń, Marta; Chrząstek, Klaudia; Grzesiak, Jakub; Meissner, Justyna

    2015-09-01

    Sphingosine-1-phosphate (S1P) and ceramide-1-phosphate (C1P) belong to a family of bioactive sphingolipids that act as important extracellular signaling molecules and chemoattractants. This study investigated the influence of S1P and C1P on the morphology, proliferation activity and osteogenic properties of rat multipotent stromal cells derived from bone marrow (BMSCs) and subcutaneous adipose tissue (ASCs). We show that S1P and C1P can influence mesenchymal stem cells (MSCs), each in a different manner. S1P stimulation promoted the formation of cellular aggregates of BMSCs and ASCs, while C1P had an effect on the regular growth pattern and expanded intercellular connections, thereby increasing the proliferative activity. Although osteogenic differentiation of MSCs was enhanced by the addition of S1P, the effectiveness of osteoblast differentiation was more evident in BMSCs, particularly when biochemical and molecular marker levels were considered. The results of the functional osteogenic differentiation assay, which includes an evaluation of the efficiency of extracellular matrix mineralization (SEM-EDX), revealed the formation of numerous mineral aggregates in BMSC cultures stimulated with S1P. Our data demonstrated that in an appropriate combination, the bioactive sphingolipids S1P and C1P may find wide application in regenerative medicine, particularly in bone regeneration with the use of MSCs.

  20. Sphingosine 1-phosphate receptor 1 (S1P(1)) upregulation and amelioration of experimental autoimmune encephalomyelitis by an S1P(1) antagonist.

    PubMed

    Cahalan, Stuart M; Gonzalez-Cabrera, Pedro J; Nguyen, Nhan; Guerrero, Miguel; Cisar, Elizabeth A George; Leaf, Nora B; Brown, Steven J; Roberts, Edward; Rosen, Hugh

    2013-02-01

    Sphingosine 1-phosphate receptor 1 (S1P(1)) is a G protein-coupled receptor that is critical for proper lymphocyte development and recirculation. Agonists to S1P(1) are currently in use clinically for the treatment of multiple sclerosis, and these drugs may act on both S1P(1) expressed on lymphocytes and S1P(1) expressed within the central nervous system. Agonists to S1P(1) and deficiency in S1P(1) both cause lymphocyte sequestration in the lymph nodes. In the present study, we show that S1P(1) antagonism induces lymphocyte sequestration in the lymph nodes similar to that observed with S1P(1) agonists while upregulating S1P(1) on lymphocytes and endothelial cells. Additionally, we show that S1P(1) antagonism reverses experimental autoimmune encephalomyelitis in mice without acting on S1P(1) expressed within the central nervous system, demonstrating that lymphocyte sequestration via S1P(1) antagonism is sufficient to alleviate autoimmune pathology.

  1. HTLV-1 p30II: selective repressor of gene expression.

    PubMed

    Green, Patrick L

    2004-11-24

    Human T-lymphotropic virus type-1 (HTLV-1) is a complex retrovirus that causes adult T-cell leukemia/lymphoma (ATL) and is implicated in a variety of lymphocyte-mediated disorders. HTLV-1 pX ORF II encodes two proteins, p13II and p30II whose roles are beginning to be defined in the virus life cycle. Previous studies indicate the importance of these viral proteins in the ability of the virus to maintain viral loads and persist in an animal model of HTLV-1 infection. Intriguing new studies indicate that p30II is a multifunctional regulator that differentially modulates CREB and Tax-responsive element-mediated transcription through its interaction with CREB-binding protein (CBP)/p300 and specifically binds and represses tax/rex mRNA nuclear export. A new study characterized the role of p30II in regulation of cellular gene expression using comprehensive human gene arrays. Interestingly, p30II is an overall repressor of cellular gene expression, while selectively favoring the expression of regulatory gene pathways important to T lymphocytes. These new findings suggest that HTLV-1, which is associated with lymphoproliferative diseases, uses p30II to selectively repress cellular and viral gene expression to favor the survival of cellular targets ultimately resulting in leukemogenesis.

  2. Hyperoxia-induced p47phox activation and ROS generation is mediated through S1P transporter Spns2, and S1P/S1P1&2 signaling axis in lung endothelium.

    PubMed

    Harijith, Anantha; Pendyala, Srikanth; Ebenezer, David L; Ha, Alison W; Fu, Panfeng; Wang, Yue-Ting; Ma, Ke; Toth, Peter T; Berdyshev, Evgeny V; Kanteti, Prasad; Natarajan, Viswanathan

    2016-08-01

    Hyperoxia-induced lung injury adversely affects ICU patients and neonates on ventilator assisted breathing. The underlying culprit appears to be reactive oxygen species (ROS)-induced lung damage. The major contributor of hyperoxia-induced ROS is activation of the multiprotein enzyme complex NADPH oxidase. Sphingosine-1-phosphate (S1P) signaling is known to be involved in hyperoxia-mediated ROS generation; however, the mechanism(s) of S1P-induced NADPH oxidase activation is unclear. Here, we investigated various steps in the S1P signaling pathway mediating ROS production in response to hyperoxia in lung endothelium. Of the two closely related sphingosine kinases (SphKs)1 and 2, which synthesize S1P from sphingosine, only Sphk1(-/-) mice conferred protection against hyperoxia-induced lung injury. S1P is metabolized predominantly by S1P lyase and partial deletion of Sgpl1 (Sgpl1(+/-)) in mice accentuated lung injury. Hyperoxia stimulated S1P accumulation in human lung microvascular endothelial cells (HLMVECs), and downregulation of S1P transporter spinster homolog 2 (Spns2) or S1P receptors S1P1&2, but not S1P3, using specific siRNA attenuated hyperoxia-induced p47(phox) translocation to cell periphery and ROS generation in HLMVECs. These results suggest a role for Spns2 and S1P1&2 in hyperoxia-mediated ROS generation. In addition, p47(phox) (phox:phagocyte oxidase) activation and ROS generation was also reduced by PF543, a specific SphK1 inhibitor in HLMVECs. Our data indicate a novel role for Spns2 and S1P1&2 in the activation of p47(phox) and production of ROS involved in hyperoxia-mediated lung injury in neonatal and adult mice.

  3. Sphingosine kinase-1, S1P transporter spinster homolog 2 and S1P2 mRNA expressions are increased in liver with advanced fibrosis in human.

    PubMed

    Sato, Masaya; Ikeda, Hitoshi; Uranbileg, Baasanjav; Kurano, Makoto; Saigusa, Daisuke; Aoki, Junken; Maki, Harufumi; Kudo, Hiroki; Hasegawa, Kiyoshi; Kokudo, Norihiro; Yatomi, Yutaka

    2016-08-26

    The role of sphingosine 1-phosphate (S1P) in liver fibrosis or inflammation was not fully examined in human. Controversy exists which S1P receptors, S1P1 and S1P3 vs S1P2, would be importantly involved in its mechanism. To clarify these matters, 80 patients who received liver resection for hepatocellular carcinoma and 9 patients for metastatic liver tumor were enrolled. S1P metabolism was analyzed in background, non-tumorous liver tissue. mRNA levels of sphingosine kinase 1 (SK1) but not SK2 were increased in livers with fibrosis stages 3-4 compared to those with 0-2 and to normal liver. However, S1P was not increased in advanced fibrotic liver, where mRNA levels of S1P transporter spinster homolog 2 (SPNS2) but not S1P-degrading enzymes were enhanced. Furthermore, mRNA levels of S1P2 but not S1P1 or S1P3 were increased in advanced fibrotic liver. These increased mRNA levels of SK1, SPNS2 and S1P2 in fibrotic liver were correlated with α-smooth muscle actin mRNA levels in liver, and with serum ALT levels. In conclusion, S1P may be actively generated, transported to outside the cells, and bind to its specific receptor in human liver to play a role in fibrosis or inflammation. Altered S1P metabolism in fibrotic liver may be their therapeutic target.

  4. The Photometric lightcurve of Comet 1P/Halley

    NASA Astrophysics Data System (ADS)

    Bair, Allison N.; Schleicher, David G.

    2014-11-01

    Comet 1P/Halley is considered an important object for a number of reasons. Not only is it the first-identified and brightest periodic comet, being the only periodic comet visible to the naked eye at every apparition, but in 1986 Halley became the first comet to be imaged by fly-by spacecraft. The NASA-funded International Halley Watch (IHW) directly supported the spacecraft by providing narrowband filters for groundbased photometric observations, and until the arrival of Hale-Bopp (1995 O1), Halley was the subject of the largest groundbased observational campaign in history. Following considerable controversy regarding its rotation period, it was eventually determined to be in complex rotation -- the first comet to be so identified. While the overall brightness variations of the coma repeated with a period of about 7.4 days, the detailed period and shape of the lightcurve constantly evolved. The determination of the specific characteristics of each of the two components of its non-principal axis rotational state has remained elusive.To resolve this situation we have now incorporated all of the narrowband photometry, taken by 21 telescopes from around the world and submitted to the IHW archive, to create the most complete homogeneous lightcurve possible. Using measurements of three gas species and the dust, the lightcurve was investigated and found to alternate between a double- and triple-peaked shape, with no single feature being present throughout the entire duration of our dataset (316 days). The apparent period as a function of time was extracted and seen to vary in a step-wise manner between 7.27 and 7.60 days. Taken together, these results were used to produce a synthetic lightcurve revealing Halley's behavior even when no data were available. Details of this and other results, to be used to constrain future detailed modeling, will be presented. This research is supported by NASA's Planetary Atmospheres Program.

  5. Downregulation of sphingosine 1-phosphate (S1P) receptor 1 by dexamethasone inhibits S1P-induced mesangial cell migration.

    PubMed

    Koch, Alexander; Jäger, Manuel; Völzke, Anja; Grammatikos, Georgios; Zu Heringdorf, Dagmar Meyer; Huwiler, Andrea; Pfeilschifter, Josef

    2015-06-01

    Sphingosine 1-phosphate (S1P) is generated by sphingosine kinase (SK)-1 and -2 and acts mainly as an extracellular ligand at five specific receptors, denoted S1P1-5. After activation, S1P receptors regulate important processes in the progression of renal diseases, such as mesangial cell migration and survival. Previously, we showed that dexamethasone enhances SK-1 activity and S1P formation, which protected mesangial cells from stress-induced apoptosis. Here we demonstrate that dexamethasone treatment lowered S1P1 mRNA and protein expression levels in rat mesangial cells. This effect was abolished in the presence of the glucocorticoid receptor antagonist RU-486. In addition, in vivo studies showed that dexamethasone downregulated S1P1 expression in glomeruli isolated from mice treated with dexamethasone (10 mg/kg body weight). Functionally, we identified S1P1 as a key player mediating S1P-induced mesangial cell migration. We show that dexamethasone treatment significantly lowered S1P-induced migration of mesangial cells, which was again reversed in the presence of RU-486. In summary, we suggest that dexamethasone inhibits S1P-induced mesangial cell migration via downregulation of S1P1. Overall, these results demonstrate that dexamethasone has functional important effects on sphingolipid metabolism and action in renal mesangial cells.

  6. Epigenetic regulation of pro-inflammatory cytokine secretion by sphingosine 1-phosphate (S1P) in acute lung injury: Role of S1P lyase.

    PubMed

    Ebenezer, David L; Fu, Panfeng; Suryadevara, Vidyani; Zhao, Yutong; Natarajan, Viswanathan

    2017-01-01

    Cellular level of sphingosine-1-phosphate (S1P), the simplest bioactive sphingolipid, is tightly regulated by its synthesis catalyzed by sphingosine kinases (SphKs) 1 & 2 and degradation mediated by S1P phosphatases, lipid phosphate phosphatases, and S1P lyase. The pleotropic actions of S1P are attributed to its unique inside-out (extracellular) signaling via G-protein-coupled S1P1-5 receptors, and intracellular receptor independent signaling. Additionally, S1P generated in the nucleus by nuclear SphK2 modulates HDAC1/2 activity, regulates histone acetylation, and transcription of pro-inflammatory genes. Here, we present data on the role of S1P lyase mediated S1P signaling in regulating LPS-induced inflammation in lung endothelium. Blocking S1P lyase expression or activity attenuated LPS-induced histone acetylation and secretion of pro-inflammatory cytokines. Degradation of S1P by S1P lyase generates Δ2-hexadecenal and ethanolamine phosphate and the long-chain fatty aldehyde produced in the cytoplasmic compartment of the endothelial cell seems to modulate histone acetylation pattern, which is different from the nuclear SphK2/S1P signaling and inhibition of HDAC1/2. These in vitro studies suggest that S1P derived long-chain fatty aldehyde may be an epigenetic regulator of pro-inflammatory genes in sepsis-induced lung inflammation. Trapping fatty aldehydes and other short chain aldehydes such as 4-hydroxynonenal derived from S1P degradation and lipid peroxidation, respectively by cell permeable agents such as phloretin or other aldehyde trapping agents may be useful in treating sepsis-induced lung inflammation via modulation of histone acetylation. .

  7. A Comprehensive Membrane Interactome Mapping of Sho1p Reveals Fps1p as a Novel Key Player in the Regulation of the HOG Pathway in S. cerevisiae

    PubMed Central

    Lam, Mandy Hiu Yi; Snider, Jamie; Rehal, Monique; Wong, Victoria; Aboualizadeh, Farzaneh; Drecun, Luka; Wong, Olivia; Jubran, Bellal; Li, Meirui; Ali, Mehrab; Jessulat, Matthew; Deineko, Viktor; Miller, Rachel; Lee, Mid eum; Park, Hay-Oak; Davidson, Alan; Babu, Mohan; Stagljar, Igor

    2017-01-01

    Sho1p, an integral membrane protein, plays a vital role in the high-osmolarity glycerol (HOG) mitogen-activated protein kinase pathway in the yeast Saccharomyces cerevisiae. Activated under conditions of high osmotic stress, it interacts with other HOG pathway proteins to mediate cell signaling events, ensuring that yeast cells can adapt and remain viable. In an attempt to further understand how the function of Sho1p is regulated through its protein–protein interactions (PPIs), we identified 49 unique Sho1p PPIs through the use of membrane yeast two-hybrid (MYTH), an assay specifically suited to identify PPIs of full-length integral membrane proteins in their native membrane environment. Secondary validation by literature search, or two complementary PPI assays, confirmed 80% of these interactions, resulting in a high-quality Sho1p interactome. This set of putative PPIs included both previously characterized interactors, along with a large subset of interactors that have not been previously identified as binding to Sho1p. The SH3 domain of Sho1p was found to be important for binding to many of these interactors. One particular novel interactor of interest is the glycerol transporter Fps1p, which was shown to require the SH3 domain of Sho1p for binding via its N-terminal soluble regulatory domain. Furthermore, we found that Fps1p is involved in the positive regulation of Sho1p function and plays a role in the phosphorylation of the downstream kinase Hog1p. This study represents the largest membrane interactome analysis of Sho1p to date and complements past studies on the HOG pathway by increasing our understanding of Sho1p regulation. PMID:25644660

  8. Refined FISH characterization of a de novo 1p22-p36.2 paracentric inversion and associated 1p21-22 deletion in a patient with signs of 1p36 microdeletion syndrome.

    PubMed

    Finelli, P; Giardino, D; Russo, S; Gottardi, G; Cogliati, F; Grugni, G; Natacci, F; Larizza, L

    2001-04-01

    We report on a 10-year-old boy presenting with obesity, moderate mental retardation, large anterior fontanelle at birth, mild physical anomalies including mid-face hypoplasia, deep-set eyes, long philtrum, and small mouth. He was found to carry a paracentric inversion inv(1)(p22p36.2) associated with a 10 cM deletion at the proximal breakpoint. By YAC FISH, the boundaries of the deletion were established at IB1028 (1p21) and WI-5166 (1p22) STSs contained in YACs 781E8 and 954F6, respectively. This large region, covering about 10 cM, contains the COL11A1 and AMY2B genes, whose haploinsufficiency does not seem to contribute significantly to the clinical phenotype. On the other hand, the patient's clinical manifestations, also including visual problems and moderate mental retardation, are those typically observed in the 1p36 deletion syndrome. Refined mapping of the telomeric 1p36.2 inversion breakpoint was obtained by FISH of a PAC contig constructed to encompass this subinterval of the 1p36 microdeletion syndrome region. PACs 1024B10 and 884E7 were found to span the breakpoint, suggesting that the clinical signs of the 1p36 microdeletion syndrome might be due to disruption of a sequence lying at 1p36.2.

  9. An 8.9 Mb 19p13 duplication associated with precocious puberty and a sporadic 3.9 Mb 2q23.3q24.1 deletion containing NR4A2 in mentally retarded members of a family with an intrachromosomal 19p-into-19q between-arm insertion.

    PubMed

    Lybaek, Helle; Ørstavik, Karen Helene; Prescott, Trine; Hovland, Randi; Breilid, Harald; Stansberg, Christine; Steen, Vidar Martin; Houge, Gunnar

    2009-07-01

    In a 2 and a half-year-old girl with onset of puberty before the age of 5 months, short stature, hand anomalies and severe mental retardation, an 8.9 Mb interstitial 19p13 duplication containing 215 predicted genes was detected. It was initially assumed that the duplication involved the kisspeptin receptor gene, GPR54, known to stimulate induction of puberty, but more refined duplication mapping excluded this possibility. In an attempt to further understand the genotype-phenotype correlation, global gene expression was measured in skin fibroblasts. The overall expression pattern was quite similar to controls, and only about 25% of the duplicated genes had an expression level that was increased by more than 1.3-fold, with no obvious changes that could explain the precocious puberty. The proband's mother carried a balanced between-arm insertion of the duplicated segment that resembled a pericentric inversion. The same insertion was found in several other family members, including one who had lost a daughter with severe mental retardation and menarche at the age of 10 years. Another close relative was severely mentally retarded, but neither dysmorphic nor microcephalic. His phenotype was initially ascribed to a presumed cryptic chromosome 19 imbalance caused by the 19p-into19q insertion, but subsequent array-CGH detected a 3.9-Mb deletion of 2q23.3q24.1. This novel microdeletion involves seven genes, of which FMNL2, a suggested regulator of Rho-GTPases, and NR4A2, an essential gene for differentiation of dopaminergic neurons, may be critical genes for the proposed 2q23q24 microdeletion syndrome.

  10. Novel selective allosteric and bitopic ligands for the S1P(3) receptor.

    PubMed

    Jo, Euijung; Bhhatarai, Barun; Repetto, Emanuela; Guerrero, Miguel; Riley, Sean; Brown, Steven J; Kohno, Yasushi; Roberts, Edward; Schürer, Stephan C; Rosen, Hugh

    2012-12-21

    Sphingosine 1-phosphate (S1P) is a lysophospholipid signaling molecule that regulates important biological functions, including lymphocyte trafficking and vascular development, by activating G protein-coupled receptors for S1P, namely, S1P(1) through S1P(5). Here, we map the S1P(3) binding pocket with a novel allosteric agonist (CYM-5541), an orthosteric agonist (S1P), and a novel bitopic antagonist (SPM-242). With a combination of site-directed mutagenesis, ligand competition assay, and molecular modeling, we concluded that S1P and CYM-5541 occupy different chemical spaces in the ligand binding pocket of S1P(3). CYM-5541 allowed us to identify an allosteric site where Phe263 is a key gate-keeper residue for its affinity and efficacy. This ligand lacks a polar moiety, and the novel allosteric hydrophobic pocket permits S1P(3) selectivity of CYM-5541 within the highly similar S1P receptor family. However, a novel S1P(3)-selective antagonist, SPM-242, in the S1P(3) pocket occupies the ligand binding spaces of both S1P and CYM-5541, showing its bitopic mode of binding. Therefore, our coordinated approach with biochemical data and molecular modeling, based on our recently published S1P(1) crystal structure data in a highly conserved set of related receptors with a shared ligand, provides a strong basis for the successful optimization of orthosteric, allosteric, and bitopic modulators of S1P(3).

  11. Pub1p C-terminal RRM domain interacts with Tif4631p through a conserved region neighbouring the Pab1p binding site.

    PubMed

    Santiveri, Clara M; Mirassou, Yasmina; Rico-Lastres, Palma; Martínez-Lumbreras, Santiago; Pérez-Cañadillas, José Manuel

    2011-01-01

    Pub1p, a highly abundant poly(A)+ mRNA binding protein in Saccharomyces cerevisiae, influences the stability and translational control of many cellular transcripts, particularly under some types of environmental stresses. We have studied the structure, RNA and protein recognition modes of different Pub1p constructs by NMR spectroscopy. The structure of the C-terminal RRM domain (RRM3) shows a non-canonical N-terminal helix that packs against the canonical RRM fold in an original fashion. This structural trait is conserved in Pub1p metazoan homologues, the TIA-1 family, defining a new class of RRM-type domains that we propose to name TRRM (TIA-1 C-terminal domain-like RRM). Pub1p TRRM and the N-terminal RRM1-RRM2 tandem bind RNA with high selectivity for U-rich sequences, with TRRM showing additional preference for UA-rich ones. RNA-mediated chemical shift changes map to β-sheet and protein loops in the three RRMs. Additionally, NMR titration and biochemical in vitro cross-linking experiments determined that Pub1p TRRM interacts specifically with the N-terminal region (1-402) of yeast eIF4G1 (Tif4631p), very likely through the conserved Box1, a short sequence motif neighbouring the Pab1p binding site in Tif4631p. The interaction involves conserved residues of Pub1p TRRM, which define a protein interface that mirrors the Pab1p-Tif4631p binding mode. Neither protein nor RNA recognition involves the novel N-terminal helix, whose functional role remains unclear. By integrating these new results with the current knowledge about Pub1p, we proposed different mechanisms of Pub1p recruitment to the mRNPs and Pub1p-mediated mRNA stabilization in which the Pub1p/Tif4631p interaction would play an important role.

  12. Smad3 deficiency leads to mandibular condyle degradation via the sphingosine 1-phosphate (S1P)/S1P3 signaling axis.

    PubMed

    Mori, Hiroki; Izawa, Takashi; Tanaka, Eiji

    2015-10-01

    Temporomandibular joint osteoarthritis is a degenerative disease that is characterized by permanent cartilage destruction. Transforming growth factor (TGF)-β is one of the most abundant cytokines in the bone matrix and is shown to regulate the migration of osteoprogenitor cells. It is hypothesized that TGF-β/Smad3 signaling affects cartilage homeostasis by influencing sphingosine 1-phosphate (S1P)/S1P receptor signaling and chondrocyte migration. We therefore investigated the molecular mechanisms by which crosstalk may occur between TGF-β/Smad3 and S1P/S1P receptor signaling to maintain condylar cartilage and to prevent temporomandibular joint osteoarthritis. Abnormalities in the condylar subchondral bone, including dynamic changes in bone mineral density and microstructure, were observed in Smad3(-/-) mice by microcomputed tomography. Cell-free regions and proteoglycan loss characterized the cartilage degradation present, and increased numbers of apoptotic chondrocytes and matrix metalloproteinase 13(+) chondrocytes were also detected. Furthermore, expression of S1P receptor 3 (S1P3), but not S1P1 or S1P2, was significantly down-regulated in the condylar cartilage of Smad3(-/-) mice. By using RNA interference technology and pharmacologic tools, S1P was found to transactivate Smad3 in an S1P3/TGF-β type II receptor-dependent manner, and S1P3 was found to be required for TGF-β-induced migration of chondrocyte cells and downstream signal transduction via Rac1, RhoA, and Cdc42. Taken together, these results indicate that the Smad3/S1P3 signaling pathway plays an important role in the pathogenesis of temporomandibular joint osteoarthritis.

  13. Bni1p implicated in cytoskeletal control is a putative target of Rho1p small GTP binding protein in Saccharomyces cerevisiae.

    PubMed Central

    Kohno, H; Tanaka, K; Mino, A; Umikawa, M; Imamura, H; Fujiwara, T; Fujita, Y; Hotta, K; Qadota, H; Watanabe, T; Ohya, Y; Takai, Y

    1996-01-01

    The RHO1 gene encodes a homolog of mammalian RhoA small GTP binding protein in the yeast Saccharomyces cerevisiae. Rho1p is localized at the growth sites, including the bud tip and the cytokinesis site, and is required for bud formation. We have recently shown that Pkc1p, a yeast homolog of mammalian protein kinase C, and glucan synthase are targets of Rho1p. Using the two-hybrid screening system, we cloned a gene encoding a protein which interacted with the GTP-bound form of Rho1p. This gene was identified as BNI1, known to be implicated in cytokinesis or establishment of cell polarity in S.cerevisiae. Bni1p shares homologous domains (FH1 and FH2 domains) with proteins involved in cytokinesis or establishment of cell polarity, including formin of mouse, capu and dia of Drosophila and FigA of Aspergillus. A temperature-sensitive mutation in which the RHO1 gene was replaced by the mammalian RhoA gene showed a synthetically lethal interaction with the bni1 mutation and the RhoA bni1 mutant accumulated cells with a deficiency in cytokinesis. Furthermore, this synthetic lethality was caused by the incapability of RhoA to activate Pkc1p, but not glucan synthase. These results suggest that Rho1p regulates cytoskeletal reorganization at least through Bni1p and Pkc1p. Images PMID:8947028

  14. Full pharmacological efficacy of a novel S1P1 agonist that does not require S1P-like head-group interactions

    PubMed Central

    Gonzalez-Cabrera, Pedro J.; Jo, Euijung; Sanna, M. Germana; Brown, Steven; Leaf, Nora; Marsolais, David; Schaeffer, Marie-Therese; Chapman, Jacqueline; Cameron, Michael; Guerrero, Miguel; Roberts, Edward; Rosen, Hugh

    2008-01-01

    Strong evidence exists for interactions of zwitterionic phosphate and amine groups in Sphingosine-1 phosphate (S1P) to conserved R and E residues present at the extracellular face of transmembrane-3 (TM3) of S1P receptors. The contribution of R120 and E121 for high affinity ligand-receptor interactions is essential, as single-point R120A or E121A S1P1 mutants neither bind S1P nor transduce S1P function. Because S1P receptors are therapeutically interesting, identifying potent selective agonists with different binding modes and in vivo efficacy is of pharmacological importance. Here we describe a modestly water-soluble highly-selective S1P1 agonist (CYM-5442) that does not require R120 or E121 residues for activating S1P1-dependent p42/p44 MAPK phosphorylation, which defines a new hydrophobic pocket in S1P1. CYM-5442 is a full agonist in vitro for S1P1 internalization, phosphorylation and ubiquitination. Importantly, CYM-5442 was a full agonist for induction and maintenance of S1P1-dependent lymphopenia, decreasing B-lymphocytes by 65% and T-lymphocytes by 85% of vehicle. Induction of CYM-5442 lymphopenia was dose and time-dependent, requiring serum concentrations in the 50 nM range. In vitro measures of S1P1 activation by CYM-5442 were non-competitively inhibited by a specific S1P1 antagonist (W146), competitive for S1P, FTY720-P and SEW2871. In addition, lymphopenia by CYM-5442 was reversed by W146 administration or upon pharmacokinetic agonist clearance. Pharmacokinetics in mice also indicated that CYM-5442 partitions significantly in central nervous tissue. These data show that CYM-5442 activates S1P1-dependent pathways in vitro and to levels of full efficacy in vivo through a hydrophobic pocket, separable from the orthosteric site of S1P binding that is headgroup dependent. PMID:18708635

  15. Ligand-binding pocket shape differences between S1P1 and S1P3 determine efficiency of chemical probe identification by uHTS

    PubMed Central

    Schürer, Stephan C.; Brown, Steven J.; Cabrera, Pedro Gonzales; Schaeffer, Marie-Therese; Chapman, Jacqueline; Jo, Euijung; Chase, Peter; Spicer, Tim; Hodder, Peter; Rosen, Hugh

    2008-01-01

    We have studied the Sphingosine 1-phosphate (S1P) receptor system to better understand why certain molecular targets within a closely related family are much more tractable when identifying compelling chemical leads. Five medically important G protein-coupled receptors for S1P regulate heart rate, coronary artery caliber, endothelial barrier integrity, and lymphocyte trafficking. Selective S1P receptor agonist probes would be of great utility to study receptor subtype-specific function. Through systematic screening of the same libraries, we identified novel selective agonists chemotypes for each of the S1P1 and S1P3 receptors. uHTS for S1P1 was more effective than for S1P3, with many selective, low nanomolar hits of proven mechanism emerging for. Receptor structure modeling and ligand docking reveal differences between the receptor binding pockets, which are the basis for sub-type selectivity. Novel selective agonists interact primarily in the hydrophobic pocket of the receptor in the absence of head-group interactions. Chemistry-space and shape-based analysis of the screening libraries in combination with the binding models explain the observed differential hit rates and enhanced efficiency for lead discovery for S1P1 vs. S1P3 in this closely related receptor family. PMID:18590333

  16. Sphingosine 1-phosphate (S1P) carrier-dependent regulation of endothelial barrier: high density lipoprotein (HDL)-S1P prolongs endothelial barrier enhancement as compared with albumin-S1P via effects on levels, trafficking, and signaling of S1P1.

    PubMed

    Wilkerson, Brent A; Grass, G Daniel; Wing, Shane B; Argraves, W Scott; Argraves, Kelley M

    2012-12-28

    Sphingosine 1-phosphate (S1P) is a blood-borne lysosphingolipid that acts to promote endothelial cell (EC) barrier function. In plasma, S1P is associated with both high density lipoproteins (HDL) and albumin, but it is not known whether the carriers impart different effects on S1P signaling. Here we establish that HDL-S1P sustains EC barrier longer than albumin-S1P. We showed that the sustained barrier effects of HDL-S1P are dependent on signaling by the S1P receptor, S1P1, and involve persistent activation of Akt and endothelial NOS (eNOS), as well as activity of the downstream NO target, soluble guanylate cyclase (sGC). Total S1P1 protein levels were found to be higher in response to HDL-S1P treatment as compared with albumin-S1P, and this effect was not associated with increased S1P1 mRNA or dependent on de novo protein synthesis. Several pieces of evidence indicate that long term EC barrier enhancement activity of HDL-S1P is due to specific effects on S1P1 trafficking. First, the rate of S1P1 degradation, which is proteasome-mediated, was slower in HDL-S1P-treated cells as compared with cells treated with albumin-S1P. Second, the long term barrier-promoting effects of HDL-S1P were abrogated by treatment with the recycling blocker, monensin. Finally, cell surface levels of S1P1 and levels of S1P1 in caveolin-enriched microdomains were higher after treatment with HDL-S1P as compared with albumin-S1P. Together, the findings reveal S1P carrier-specific effects on S1P1 and point to HDL as the physiological mediator of sustained S1P1-PI3K-Akt-eNOS-sGC-dependent EC barrier function.

  17. A Prokaryotic S1P Lyase Degrades Extracellular S1P In Vitro and In Vivo: Implication for Treating Hyperproliferative Disorders

    PubMed Central

    Huwiler, Andrea; Bourquin, Florence; Kotelevets, Nataliya; Pastukhov, Oleksandr; Capitani, Guido; Grütter, Markus G.; Zangemeister-Wittke, Uwe

    2011-01-01

    Sphingosine-1-phosphate (S1P) regulates a broad spectrum of fundamental cellular processes like proliferation, death, migration and cytokine production. Therefore, elevated levels of S1P may be causal to various pathologic conditions including cancer, fibrosis, inflammation, autoimmune diseases and aberrant angiogenesis. Here we report that S1P lyase from the prokaryote Symbiobacterium thermophilum (StSPL) degrades extracellular S1P in vitro and in blood. Moreover, we investigated its effect on cellular responses typical of fibrosis, cancer and aberrant angiogenesis using renal mesangial cells, endothelial cells, breast (MCF-7) and colon (HCT 116) carcinoma cells as disease models. In all cell types, wild-type StSPL, but not an inactive mutant, disrupted MAPK phosphorylation stimulated by exogenous S1P. Functionally, disruption of S1P receptor signaling by S1P depletion inhibited proliferation and expression of connective tissue growth factor in mesangial cells, proliferation, migration and VEGF expression in carcinoma cells, and proliferation and migration of endothelial cells. Upon intravenous injection of StSPL in mice, plasma S1P levels rapidly declined by 70% within 1 h and then recovered to normal 6 h after injection. Using the chicken chorioallantoic membrane model we further demonstrate that also under in vivo conditions StSPL, but not the inactive mutant, inhibited tumor cell-induced angiogenesis as an S1P-dependent process. Our data demonstrate that recombinant StSPL is active under extracellular conditions and holds promise as a new enzyme therapeutic for diseases associated with increased levels of S1P and S1P receptor signaling. PMID:21829623

  18. Sphingosine 1-phosphate (S1P) receptor agonists mediate pro-fibrotic responses in normal human lung fibroblasts via S1P2 and S1P3 receptors and Smad-independent signaling.

    PubMed

    Sobel, Katrin; Menyhart, Katalin; Killer, Nina; Renault, Bérengère; Bauer, Yasmina; Studer, Rolf; Steiner, Beat; Bolli, Martin H; Nayler, Oliver; Gatfield, John

    2013-05-24

    Synthetic sphingosine 1-phosphate receptor 1 modulators constitute a new class of drugs for the treatment of autoimmune diseases. Sphingosine 1-phosphate (S1P) signaling, however, is also involved in the development of fibrosis. Using normal human lung fibroblasts, we investigated the induction of fibrotic responses by the S1P receptor (S1PR) agonists S1P, FTY720-P, ponesimod, and SEW2871 and compared them with the responses induced by the known fibrotic mediator TGF-β1. In contrast to TGF-β1, S1PR agonists did not induce expression of the myofibroblast marker α-smooth muscle actin. However, TGF-β1, S1P, and FTY720-P caused robust stimulation of extracellular matrix (ECM) synthesis and increased pro-fibrotic marker gene expression including connective tissue growth factor. Ponesimod showed limited and SEW2871 showed no pro-fibrotic potential in these readouts. Analysis of pro-fibrotic signaling pathways showed that in contrast to TGF-β1, S1PR agonists did not activate Smad2/3 signaling but rather activated PI3K/Akt and ERK1/2 signaling to induce ECM synthesis. The strong induction of ECM synthesis by the nonselective agonists S1P and FTY720-P was due to the stimulation of S1P2 and S1P3 receptors, whereas the weaker induction of ECM synthesis at high concentrations of ponesimod was due to a low potency activation of S1P3 receptors. Finally, in normal human lung fibroblast-derived myofibroblasts that were generated by TGF-β1 pretreatment, S1P and FTY720-P were effective stimulators of ECM synthesis, whereas ponesimod was inactive, because of the down-regulation of S1P3R expression in myofibroblasts. These data demonstrate that S1PR agonists are pro-fibrotic via S1P2R and S1P3R stimulation using Smad-independent pathways.

  19. The yeast dynamin-related GTPase Vps1p functions in the organization of the actin cytoskeleton via interaction with Sla1p.

    PubMed

    Yu, Xianwen; Cai, Mingjie

    2004-08-01

    Recent studies have suggested that the function of the large GTPase dynamin in endocytosis in mammalian cells may comprise a modulation of actin cytoskeleton. The role of dynamin in actin cytoskeleton organization in the yeast Saccharomyces cerevisiae has remained undefined. In this report, we found that one of the yeast dynamin-related proteins, Vps1p, is required for normal actin cytoskeleton organization. At both permissive and non-permissive temperatures, the vps1 mutants exhibited various degrees of phenotypes commonly associated with actin cytoskeleton defects: depolarized and aggregated actin structures, hypersensitivity to the actin cytoskeleton toxin latrunculin-A, randomized bud site selection and chitin deposition, and impaired efficiency in the internalization of membrane receptors. Over-expression of the GTPase mutants of vps1 also led to actin abnormalities. Consistent with these actin-related defects, Vps1p was found to interact physically, and partially co-localize, with the actin-regulatory protein Sla1p. The normal cellular localization of Sla1p required Vps1p and could be altered by over-expression of a region of Vps1p that was involved in the interaction with Sla1p. The same region also promoted mis-sorting of the vacuolar protein carboxypeptidase Y upon over-expression. These findings suggest that the functions of the dynamin-related protein Vps1p in actin cytoskeleton dynamics and vacuolar protein sorting are probably related to each other.

  20. Identification and subcellular localization of a novel mammalian dynamin-related protein homologous to yeast Vps1p and Dnm1p.

    PubMed

    Shin, H W; Shinotsuka, C; Torii, S; Murakami, K; Nakayama, K

    1997-09-01

    The dynamin family of GTP-binding proteins are implicated in vesicular transport. These include mammalian dynamins I, II, III, and yeast Vps1p and Dnm1p. Dynamin is involved in the formation of clathrin-coated vesicles at the plasma membrane. On the other hand, Vps1p and Dnm1p appear to be involved in transport from the late Golgi compartment to vacuoles and in an endocytic process, respectively. In this study, we identified a novel human protein, named Dnm1p/Vps1p-like protein (DVLP). It resembled more closely Dnm1p and Vps1p than dynamins not only in the primary structure but also in the domain organization. DVLP mRNA was expressed ubiquitously, suggesting that this protein plays a fundamental role in cellular function. Immunofluorescence analysis of cells expressing epitope-tagged DVLP revealed that it showed a diffused perinuclear staining pattern that was not superimposed on that of the marker protein for the Golgi apparatus, trans-Golgi network, lysosomes, endosomes, or endoplasmic reticulum. These data suggest that DVLP is not involved in the formation of known coated vesicles.

  1. HDL-S1P: cardiovascular functions, disease-associated alterations, and therapeutic applications.

    PubMed

    Levkau, Bodo

    2015-01-01

    Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid contained in High-density lipoproteins (HDL) and has drawn considerable attention in the lipoprotein field as numerous studies have demonstrated its contribution to several functions inherent to HDL. Some of them are partly and some entirely due to the S1P contained in HDL (HDL-S1P). Despite the presence of over 1000 different lipids in HDL, S1P stands out as it possesses its own cell surface receptors through which it exercises key physiological functions. Most of the S1P in human plasma is associated with HDL, and the amount of HDL-S1P influences the quality and quantity of HDL-dependent functions. The main binding partner of S1P in HDL is apolipoprotein M but others may also exist particularly under conditions of acute S1P elevations. HDL not only exercise functions through their S1P content but have also an impact on genuine S1P signaling by influencing S1P bioactivity and receptor presentation. HDL-S1P content is altered in human diseases such as atherosclerosis, coronary artery disease, myocardial infarction, renal insufficiency and diabetes mellitus. Low HDL-S1P has also been linked to impaired HDL functions associated with these disorders. Although the pathophysiological and molecular reasons for such disease-associated shifts in HDL-S1P are little understood, there have been successful approaches to circumvent their adverse implications by pharmacologically increasing HDL-S1P as means to improve HDL function. This mini-review will cover the current understanding of the contribution of HDL-S1P to physiological HDL function, its alteration in disease and ways for its restoration to correct HDL dysfunction.

  2. Oncogenic S1P signalling in EBV-associated nasopharyngeal carcinoma activates AKT and promotes cell migration through S1P receptor 3.

    PubMed

    Lee, Hui Min; Lo, Kwok-Wai; Wei, Wenbin; Tsao, Sai Wah; Chung, Grace Tin Yun; Ibrahim, Maha Hafez; Dawson, Christopher W; Murray, Paul G; Paterson, Ian C; Yap, Lee Fah

    2017-02-27

    Undifferentiated nasopharyngeal carcinoma (NPC) is a cancer with high metastatic potential that is consistently associated with Epstein-Barr virus (EBV) infection. In this study, we have investigated the functional contribution of sphingosine-1-phosphate (S1P) signalling to the pathogenesis of NPC. We show that EBV infection or ectopic expression of the EBV-encoded latent genes (EBNA1, LMP1 and LMP2A) can up-regulate sphingosine kinase 1 (SPHK1), the key enzyme that produces S1P, in NPC cell lines. Exogenous addition of S1P promotes the migration of NPC cells through the activation of AKT; shRNA knockdown of SPHK1 resulted in a reduction in the levels of activated AKT and inhibition of cell migration. We also show that S1P receptor 3 (S1PR3) mRNA is over-expressed in EBV-positive NPC patient-derived xenografts and a subset of primary NPC tissues, and that knockdown of S1PR3 suppressed the activation of AKT and the S1P-induced migration of NPC cells. Taken together, our data point to a central role for EBV in mediating the oncogenic effects of S1P in NPC and identify S1P signalling as a potential therapeutic target in this disease.

  3. A shunt pathway limits the CaaX processing of Hsp40 Ydj1p and regulates Ydj1p-dependent phenotypes

    PubMed Central

    Hildebrandt, Emily R; Cheng, Michael; Zhao, Peng; Kim, June H; Wells, Lance; Schmidt, Walter K

    2016-01-01

    The modifications occurring to CaaX proteins have largely been established using few reporter molecules (e.g. Ras, yeast a-factor mating pheromone). These proteins undergo three coordinated COOH-terminal events: isoprenylation of the cysteine, proteolytic removal of aaX, and COOH-terminal methylation. Here, we investigated the coupling of these modifications in the context of the yeast Ydj1p chaperone. We provide genetic, biochemical, and biophysical evidence that the Ydj1p CaaX motif is isoprenylated but not cleaved and carboxylmethylated. Moreover, we demonstrate that Ydj1p-dependent thermotolerance and Ydj1p localization are perturbed when alternative CaaX motifs are transplanted onto Ydj1p. The abnormal phenotypes revert to normal when post-isoprenylation events are genetically interrupted. Our findings indicate that proper Ydj1p function requires an isoprenylatable CaaX motif that is resistant to post-isoprenylation events. These results expand on the complexity of protein isoprenylation and highlight the impact of post-isoprenylation events in regulating the function of Ydj1p and perhaps other CaaX proteins. DOI: http://dx.doi.org/10.7554/eLife.15899.001 PMID:27525482

  4. S1P3 confers differential S1P migration by autoreactive and non-autoreactive immature B cells and is required for normal B cell development

    PubMed Central

    Donovan, Erin E.; Pelanda, Roberta; Torres, Raul M.

    2010-01-01

    SUMMARY During B cell development, immature B cell fate is determined by whether the B cell antigen receptor is engaged in the bone marrow. Immature B cells that are non-autoreactive continue maturation and emigrate from the marrow whereas autoreactive immature B cells remain and are tolerized. However, the microenvironment where these events occur and the chemoattractants responsible for immature B cell trafficking within and out of the bone marrow remain largely undefined. Sphingosine 1-phosphate (S1P) is a chemoattractant that directs lymphocyte trafficking and thymocyte egress and in this study we investigated whether S1P contributed to B cell development, egress and positioning within the bone marrow. Our findings show that immature B cells are chemotactic towards S1P but that this response is dependent on antigen receptor specificity: non-autoreactive, but not autoreactive, immature B cells migrate towards S1P and are shown to require S1P3 receptor for this response. Despite this response, S1P3 is shown not to facilitate immature B cell egress but is required for normal B cell development including the positioning of transitional B cells within bone marrow sinusoids. These data indicate that S1P3 signaling directs immature B cells to a bone marrow microenvironment important for both tolerance induction and maturation. PMID:20039302

  5. S1P differentially regulates migration of human ovarian cancer and human ovarian surface epithelial cells

    PubMed Central

    Wang, Dongmei; Zhao, Zhenwen; Caperell-Grant, Andrea; Yang, Gong; Mok, Samuel C.; Liu, Jinsong; Bigsby, Robert M.; Xu, Yan

    2009-01-01

    Epithelial ovarian cancer (EOC) arises from the epithelial layer covering the surface of ovaries and intra-peritoneal metastasis is commonly observed at diagnosis. Sphingosine-1-phosphate (S1P), a bioactive lipid signaling molecule, is potentially involved in EOC tumorigenesis. We have found that S1P is elevated in human EOC ascites. We show that physiologically relevant concentrations of S1P stimulate migration and invasion of EOC cells, but inhibit migration of human ovarian surface epithelial (HOSE) cells. In addition, S1P inhibits lysophosphatidic acid (LPA)-induced cell migration in HOSE, but not in EOC cells. We have provided the first line of evidence that the expression levels of S1P receptor subtypes are not the only determinants for how cells respond to S1P. Even though S1P1 is expressed and functional in HOSE cells, the inhibitory effect mediated by S1P2 is dominant in those cells. The cellular pre-existing stress fibers are also important determinants for the migratory response to S1P. Differential S1P-induced morphology changes are noted in EOC and HOSE cells. Pre-existing stress fibers in HOSE cells are further enhanced by S1P treatment, resulting in the negative migratory response to S1P. By contrast, EOC cells lost stress fibers and S1P treatment induces filopodium-like structures at cell edges, which correlates with increased cell motility. In addition, inhibition of the protein kinase C pathway is likely to be involved in the inhibitory effect of S1P on LPA-induced cell migration in HOSE cells. These findings are important for the development of new therapeutics targeting S1P and LPA in EOC. PMID:18645009

  6. Interaction of integrin β4 with S1P receptors in S1P- and HGF-induced endothelial barrier enhancement.

    PubMed

    Ni, Xiuqin; Epshtein, Yulia; Chen, Weiguo; Zhou, Tingting; Xie, Lishi; Garcia, Joe G N; Jacobson, Jeffrey R

    2014-06-01

    We previously reported sphingosine 1-phosphate (S1P) and hepatocyte growth factor (HGF) augment endothelial cell (EC) barrier function and attenuate murine acute lung inury (ALI). While the mechanisms underlying these effects are not fully understood, S1P and HGF both transactivate the S1P receptor, S1PR1 and integrin β4 (ITGB4) at membrane caveolin-enriched microdomains (CEMs). In the current study, we investigated the roles of S1PR2 and S1PR3 in S1P/HGF-mediated EC signaling and their associations with ITGB4. Our studies confirmed ITGB4 and S1PR2/3 are recruited to CEMs in human lung EC in response to either S1P (1 µM, 5 min) or HGF (25 ng/ml, 5 min). Co-immunoprecipitation experiments identified an S1P/HGF-mediated interaction of ITGB4 with both S1PR2 and S1PR3. We then employed an in situ proximity ligation assay (PLA) to confirm a direct ITGB4-S1PR3 association induced by S1P/HGF although a direct association was not detectable between S1PR2 and ITGB4. S1PR1 knockdown (siRNA), however, abrogated S1P/HGF-induced ITGB4-S1PR2 associations while there was no effect on ITGB4-S1PR3 associations. Moreover, PLA confirmed a direct association between S1PR1 and S1PR2 induced by S1P and HGF. Finally, silencing of S1PR2 significantly attenuated S1P/HGF-induced EC barrier enhancement as measured by transendothelial resistance while silencing of S1PR3 significantly augmented S1P/HGF-induced barrier enhancement. These results confirm an important role for S1PR2 and S1PR3 in S1P/HGF-mediated EC barrier responses that are associated with their complex formation with ITGB4. Our findings elucidate novel mechanisms of EC barrier regulation that may ultimately lead to new therapeutic targets for disorders characterized by increased vascular permeability including ALI.

  7. Phytosphingosine 1-phosphate: a high affinity ligand for the S1P(4)/Edg-6 receptor.

    PubMed

    Candelore, Mari Rios; Wright, Michael J; Tota, Laurie M; Milligan, James; Shei, Gan-ju; Bergstrom, James D; Mandala, Suzanne M

    2002-09-27

    It has been reported recently that the phosphorylated form of the immunomodulator FTY720 activates sphingosine 1-phosphate G protein-coupled receptors. Therefore, understanding the biology of this new class of receptors will be important in clarifying the immunological function of bioactive lysosphingolipid ligands. The S1P(4) receptor has generated interest due to its lymphoid tissue distribution. While the S1P(4) receptor binds the prototypical ligand, S1P, a survey of other lysosphingolipids demonstrated that 4D-hydroxysphinganine 1-phosphate, more commonly known as phytosphingosine 1-phosphate (PhS1P), binds to S1P(4) with higher affinity. Using radiolabeled S1P (S133P), the affinity of PhS1P for the S1P(4) receptor is 1.6nM, while that of S1P is nearly 50-fold lower (119+/-20nM). Radiolabeled PhS1P proved to be superior to S133P in routine binding assays due to improved signal-to-noise ratio. The present study demonstrates the utility of a novel radiolabeled probe, PhS133P, for in vitro studies of the S1P(4) receptor pharmacology.

  8. S1P lyase in skeletal muscle regeneration and satellite cell activation: exposing the hidden lyase.

    PubMed

    Saba, Julie D; de la Garza-Rodea, Anabel S

    2013-01-01

    Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid whose actions are essential for many physiological processes including angiogenesis, lymphocyte trafficking and development. In addition, S1P serves as a muscle trophic factor that enables efficient muscle regeneration. This is due in part to S1P's ability to activate quiescent muscle stem cells called satellite cells (SCs) that are needed for muscle repair. However, the molecular mechanism by which S1P activates SCs has not been well understood. Further, strategies for harnessing S1P signaling to recruit SCs for therapeutic benefit have been lacking. S1P is irreversibly catabolized by S1P lyase (SPL), a highly conserved enzyme that catalyzes the cleavage of S1P at carbon bond C(2-3), resulting in formation of hexadecenal and ethanolamine-phosphate. SPL enhances apoptosis through substrate- and product-dependent events, thereby regulating cellular responses to chemotherapy, radiation and ischemia. SPL is undetectable in resting murine skeletal muscle. However, we recently found that SPL is dynamically upregulated in skeletal muscle after injury. SPL upregulation occurred in the context of a tightly orchestrated genetic program that resulted in a transient S1P signal in response to muscle injury. S1P activated quiescent SCs via a sphingosine-1-phosphate receptor 2 (S1P2)/signal transducer and activator of transcription 3 (STAT3)-dependent pathway, thereby facilitating skeletal muscle regeneration. Mdx mice, which serve as a model for muscular dystrophy (MD), exhibited skeletal muscle SPL upregulation and S1P deficiency. Pharmacological SPL inhibition raised skeletal muscle S1P levels, enhanced SC recruitment and improved mdx skeletal muscle regeneration. These findings reveal how S1P can activate SCs and indicate that SPL suppression may provide a therapeutic strategy for myopathies. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.

  9. Bioactive lipids S1P and C1P are prometastatic factors in human rhabdomyosarcoma, and their tissue levels increase in response to radio/chemotherapy.

    PubMed

    Schneider, Gabriela; Bryndza, Ewa; Abdel-Latif, Ahmed; Ratajczak, Janina; Maj, Magdalena; Tarnowski, Maciej; Klyachkin, Yuri M; Houghton, Peter; Morris, Andrew J; Vater, Axel; Klussmann, Sven; Kucia, Magdalena; Ratajczak, Mariusz Z

    2013-07-01

    Evidence suggests that bioactive lipids may regulate pathophysiologic functions such as cancer cell metastasis. Therefore, we determined that the bioactive lipid chemoattractants sphingosine-1-phosphate (S1P) and ceramide-1-phosphate (C1P) strongly enhanced the in vitro motility and adhesion of human rhabdomyosarcoma (RMS) cells. Importantly, this effect was observed at physiologic concentrations for both bioactive lipids, which are present in biologic fluids, and were much stronger than the effects observed in response to known RMS prometastatic factors such as stromal derived factors-1 (SDF-1/CXCL12) or hepatocyte growth factor/scatter factor (HGF/SF). We also present novel evidence that the levels of S1P and C1P were increased in several organs after γ-irradiation or chemotherapy, which indicates an unwanted prometastatic environment related to treatment. Critically, we found that the metastasis of RMS cells in response to S1P can be effectively inhibited in vivo with the S1P-specific binder NOX-S93 that is based on a high-affinity Spiegelmer. These data indicate that bioactive lipids play a vital role in dissemination of RMS and contribute to the unwanted side effects of radio/chemotherapy by creating a prometastatic microenvironment.

  10. Multiple functions of the vacuolar sorting protein Ccz1p in Saccharomyces cerevisiae

    SciTech Connect

    Hoffman-Sommer, Marta; Migdalski, Andrzej; Rytka, Joanna; Kucharczyk, Roza . E-mail: roza@ibb.waw.pl

    2005-04-01

    The CCZ1 (YBR131w) gene encodes a protein required for fusion of various transport intermediates with the vacuole. Ccz1p, in a complex with Mon1p, is a close partner of Ypt7p in the processes of fusion of endosomes to vacuoles and homotypic vacuole fusion. In this work, we exploited the Ca{sup 2+}-sensitivity of the ccz1{delta} mutant to identify genes specifically interacting with CCZ1, basing on functional multicopy suppression of calcium toxicity. The presented results indicate that Ccz1p functions in the cell either in association with Mon1p and Ypt7p in fusion at the vacuolar membrane, or-separately-with Arl1p at early steps of vacuolar transport. We also show that suppression of calcium toxicity by the calcium pumps Pmr1p and Pmc1p is restricted only to the subset of mutants defective in vacuole morphology. The mechanisms of Ca{sup 2+}-pump-mediated suppression also differ from each other, since the action of Pmr1p, but not Pmc1p, appears to require Arl1p function.

  11. A girl with 1p36 deletion syndrome and congenital fiber type disproportion myopathy.

    PubMed

    Okamoto, Nobuhiko; Toribe, Yasuhisa; Nakajima, Tohru; Okinaga, Takeshi; Kurosawa, Kenji; Nonaka, Ikuya; Shimokawa, Osamu; Matsumoto, Noamichi

    2002-01-01

    Chromosome 1p36 deletion syndrome is characterized by hypotonia, moderate to severe developmental and growth retardation, and characteristic craniofacial dysmorphism. Muscle hypotonia and delayed motor development are almost constant features of the syndrome. We report a 4-year-old Japanese girl with 1p36 deletion syndrome whose muscle pathology showed congenital fiber type disproportion (CFTD) myopathy. This is the first case report of 1p36 deletion associated with CFTD. This association may indicate that one of the CFTD loci is located at 1p36. Ski proto-oncogene -/- mice have phenotypes that resemble some of the features observed in patients with 1p36 deletion syndrome. Because fluorescent in situ hybridization analysis revealed that the human SKI gene is deleted in our patient, some genes in 1p36, including SKI proto-oncogene, may be involved in muscle hypotonia and delayed motor development in this syndrome.

  12. Discovery of a novel series of potent S1P1 agonists.

    PubMed

    Crosignani, Stefano; Bombrun, Agnes; Covini, David; Maio, Maurizio; Marin, Delphine; Quattropani, Anna; Swinnen, Dominique; Simpson, Don; Sauer, Wolfgang; Françon, Bernard; Martin, Thierry; Cambet, Yves; Nichols, Anthony; Martinou, Isabelle; Burgat-Charvillon, Fabienne; Rivron, Delphine; Donini, Cristina; Schott, Olivier; Eligert, Valerie; Novo-Perez, Laurence; Vitte, Pierre-Alain; Arrighi, Jean-François

    2010-03-01

    The discovery of a novel series of S1P1 agonists is described. Starting from a micromolar HTS positive, iterative optimization gave rise to several single-digit nanomolar S1P1 agonists. The compounds were able to induce internalization of the S1P1 receptor, and a selected compound was shown to be able to induce lymphopenia in mice after oral dosing.

  13. Biophysical Characterization of the Iron in Mitochondria from Atm1p-depleted Saccharomyces cerevisiae

    PubMed Central

    Miao, Ren; Kim, Hansoo; Koppolu, Uma Mahendra Kumar; Ellis, E. Ann; Scott, Robert A.; Lindahl, Paul A.

    2009-01-01

    Atm1p is an ABC transporter localized in the mitochondrial inner membrane; it functions to export an unknown species into the cytosol and is involved in cellular iron metabolism. Depletion or deletion of Atm1p causes Fe accumulation in mitochondria and a defect in cytosolic Fe/S cluster assembly, but reportedly not a defect in mitochondrial Fe/S cluster assembly. In this study the nature of the accumulated Fe was examined using Mössbauer spectroscopy, EPR, electronic absorption spectroscopy, X-ray absorption spectroscopy, and electron microscopy. The Fe that accumulated in aerobically grown cells was in the form of Fe(III) phosphate nanoparticles similar to that which accumulates in yeast frataxin Yfh1p-deleted or yeast ferredoxin Yah1p-depleted cells. Relative to WT mitochondria, Fe/S cluster and heme levels in Atm1p-depleted mitochondria from aerobic cells were significantly diminished. Atm1p-depletion also caused a build-up of nonheme Fe(II) ions in the mitochondria and an increase in oxidative damage. Atm1p-depleted mitochondria isolated from anaerobically grown cells exhibited WT levels of Fe/S clusters and hemes, and they did not hyper-accumulate Fe. Atm1p-depleted cells lacked Leu1p activity, regardless of whether they were grown aerobically or anaerobically. These results indicate that Atm1p does not participate in mitochondrial Fe/S cluster assembly, and that the species exported by Atm1p is required for cytosolic Fe/S cluster assembly. The Fe/S cluster defect and the Fe-accumulation phenotype, resulting from the depletion of Atm1p in aerobic cells (but not in anaerobic cells), may be secondary effects that are observed only when cells are exposed to oxygen during growth. Reactive oxygen species generated under these conditions might degrade iron-sulfur clusters and lower heme levels in the organelle. PMID:19761223

  14. Role of Pex21p for Piggyback Import of Gpd1p and Pnc1p into Peroxisomes of Saccharomyces cerevisiae*

    PubMed Central

    Effelsberg, Daniel; Cruz-Zaragoza, Luis Daniel; Tonillo, Jason; Schliebs, Wolfgang; Erdmann, Ralf

    2015-01-01

    Proteins designated for peroxisomal protein import harbor one of two common peroxisomal targeting signals (PTS). In the yeast Saccharomyces cerevisiae, the oleate-induced PTS2-dependent import of the thiolase Fox3p into peroxisomes is conducted by the soluble import receptor Pex7p in cooperation with the auxiliary Pex18p, one of two supposedly redundant PTS2 co-receptors. Here, we report on a novel function for the co-receptor Pex21p, which cannot be fulfilled by Pex18p. The data establish Pex21p as a general co-receptor in PTS2-dependent protein import, whereas Pex18p is especially important for oleate-induced import of PTS2 proteins. The glycerol-producing PTS2 protein glycerol-3-phosphate dehydrogenase Gpd1p shows a tripartite localization in peroxisomes, in the cytosol, and in the nucleus under osmotic stress conditions. We show the following: (i) Pex21p is required for peroxisomal import of Gpd1p as well as a key enzyme of the NAD+ salvage pathway, Pnc1p; (ii) Pnc1p, a nicotinamidase without functional PTS2, is co-imported into peroxisomes by piggyback transport via Gpd1p. Moreover, the specific transport of these two enzymes into peroxisomes suggests a novel regulatory role for peroxisomes under various stress conditions. PMID:26276932

  15. A role of the sphingosine-1-phosphate (S1P)-S1P receptor 2 pathway in epithelial defense against cancer (EDAC).

    PubMed

    Yamamoto, Sayaka; Yako, Yuta; Fujioka, Yoichiro; Kajita, Mihoko; Kameyama, Takeshi; Kon, Shunsuke; Ishikawa, Susumu; Ohba, Yusuke; Ohno, Yusuke; Kihara, Akio; Fujita, Yasuyuki

    2016-02-01

    At the initial step of carcinogenesis, transformation occurs in single cells within epithelia, where the newly emerging transformed cells are surrounded by normal epithelial cells. A recent study revealed that normal epithelial cells have an ability to sense and actively eliminate the neighboring transformed cells, a process named epithelial defense against cancer (EDAC). However, the molecular mechanism of this tumor-suppressive activity is largely unknown. In this study, we investigated a role for the sphingosine-1-phosphate (S1P)-S1P receptor 2 (S1PR2) pathway in EDAC. First, we show that addition of the S1PR2 inhibitor significantly suppresses apical extrusion of RasV12-transformed cells that are surrounded by normal cells. In addition, knockdown of S1PR2 in normal cells induces the same effect, indicating that S1PR2 in the surrounding normal cells plays a positive role in the apical elimination of the transformed cells. Of importance, not endogenous S1P but exogenous S1P is involved in this process. By using FRET analyses, we demonstrate that S1PR2 mediates Rho activation in normal cells neighboring RasV12-transformed cells, thereby promoting accumulation of filamin, a crucial regulator of EDAC. Collectively these data indicate that S1P is a key extrinsic factor that affects the outcome of cell competition between normal and transformed epithelial cells.

  16. Young-Simpson syndrome (YSS), a variant of del(1)(p36) syndrome?

    PubMed

    Robinson, Deanne Mraz; Meagher, Cecilia C; Orlowski, Craig C; Lagoe, Erin Caine; Fong, Chin-To

    2008-06-15

    The Young-Simpson syndrome (YSS) and 1p36 deletion syndrome are both characterized by facial and heart abnormalities, congenital hypothyroidism, and severe growth and developmental retardation. However, the YSS is characterized by the presence of blepharophimosis and epicanthus inversus, findings not described in monosomy 1p36 patients. We describe a girl with YSS, who presented with the typical facial findings, global retardation, congenital hypothyroidism, and congenital dilated cardiomyopathy. Comparative genomic hybridization chromosomal microarray analysis showed a 1p36.3 deletion, a finding not previously reported in other YSS cases. We propose that YSS is a variant of the 1p36 deletion syndrome.

  17. Flow-regulated endothelial S1P receptor-1 signaling sustains vascular development

    PubMed Central

    Jung, Bongnam; Obinata, Hideru; Galvani, Sylvain; Mendelson, Karen; Ding, Bisen; Skoura, Athanasia; Kinzel, Bernd; Brinkmann, Volker; Rafii, Shahin; Evans, Todd; Hla, Timothy

    2012-01-01

    SUMMARY During angiogenesis, nascent vascular sprouts fuse to form vascular networks enabling efficient circulation. Mechanisms that stabilize the vascular plexus are not well understood. Sphingosine 1-phosphate (S1P) is a blood-borne lipid mediator implicated in the regulation of vascular and immune systems. Here we describe a mechanism by which the G protein-coupled S1P receptor-1 (S1P1) stabilizes the primary vascular network. A gradient of S1P1 expression from the mature regions of the vascular network to the growing vascular front was observed. In the absence of endothelial S1P1, adherens junctions are destabilized, barrier function is breached, and flow is perturbed resulting in abnormal vascular hypersprouting. Interestingly, S1P1 responds to S1P as well as laminar shear stress to transduce flow-mediated signaling in endothelial cells both in vitro and in vivo. These data demonstrate that blood flow and circulating S1P activate endothelial S1P1 to stabilize blood vessels in development and homeostasis. PMID:22975328

  18. Steps in reductive activation of the disulfide-generating enzyme Ero1p

    PubMed Central

    Heldman, Nimrod; Vonshak, Ohad; Sevier, Carolyn S; Vitu, Elvira; Mehlman, Tevie; Fass, Deborah

    2010-01-01

    Ero1p is the primary catalyst of disulfide bond formation in the yeast endoplasmic reticulum (ER). Ero1p contains a pair of essential disulfide bonds that participate directly in the electron transfer pathway from substrate thiol groups to oxygen. Remarkably, elimination of certain other Ero1p disulfides by mutation enhances enzyme activity. In particular, the C150A/C295A Ero1p mutant exhibits increased thiol oxidation in vitro and in vivo and interferes with redox homeostasis in yeast cells by hyperoxidizing the ER. Inhibitory disulfides of Ero1p are thus important for enzyme regulation. To visualize the differences between de-regulated and wild-type Ero1p, we determined the crystal structure of Ero1p C150A/C295A. The structure revealed local changes compared to the wild-type enzyme around the sites of mutation, but no conformational transitions within 25 Å of the active site were observed. To determine how the C150—C295 disulfide nonetheless participates in redox regulation of Ero1p, we analyzed using mass spectrometry the changes in Ero1p disulfide connectivity as a function of time after encounter with reducing substrates. We found that the C150—C295 disulfide sets a physiologically appropriate threshold for enzyme activation by guarding a key neighboring disulfide from reduction. This study illustrates the diverse and interconnected roles that disulfides can play in redox regulation of protein activity. PMID:20669236

  19. FRG1P-mediated aggregation of proteins involved in pre-mRNA processing.

    PubMed

    van Koningsbruggen, Silvana; Straasheijm, Kirsten R; Sterrenburg, Ellen; de Graaf, Natascha; Dauwerse, Hans G; Frants, Rune R; van der Maarel, Silvère M

    2007-02-01

    FRG1 is considered a candidate gene for facioscapulohumeral muscular dystrophy (FSHD) based on its location at chromosome 4qter and its upregulation in FSHD muscle. The FRG1 protein (FRG1P) localizes to nucleoli, Cajal bodies (and speckles), and has been suggested to be a component of the human spliceosome but its exact function is unknown. Recently, transgenic mice overexpressing high levels of FRG1P in skeletal muscle were described to present with muscular dystrophy. Moreover, upregulation of FRG1P was demonstrated to correlate with missplicing of specific pre-mRNAs. In this study, we have combined colocalization studies with yeast two-hybrid screens to identify proteins that associate with FRG1P. We demonstrate that artificially induced nucleolar aggregates of VSV-FRG1P specifically sequester proteins involved in pre-mRNA processing. In addition, we have identified SMN, PABPN1, and FAM71B, a novel speckle and Cajal body protein, as binding partners of FRG1P. All these proteins are, or seem to be, involved in RNA biogenesis. Our data confirm the presence of FRG1P in protein complexes containing human spliceosomes and support a potential role of FRG1P in either splicing or another step in nuclear RNA biogenesis. Intriguingly, among FRG1P-associated proteins are SMN and PABPN1, both being involved in neuromuscular disorders, possibly through RNA biogenesis-related processes.

  20. Distinct roles of galactose-1P in galactose-mediated growth arrest of yeast deficient in galactose-1P uridylyltransferase (GALT) and UDP-galactose 4'-epimerase (GALE).

    PubMed

    Mumma, Jane Odhiambo; Chhay, Juliet S; Ross, Kerry L; Eaton, Jana S; Newell-Litwa, Karen A; Fridovich-Keil, Judith L

    2008-02-01

    Galactose is metabolized in humans and other species by the three-enzyme Leloir pathway comprised of galactokinase (GALK), galactose 1-P uridylyltransferase (GALT), and UDP-galactose 4'-epimerase (GALE). Impairment of GALT or GALE in humans results in the potentially lethal disorder galactosemia, and loss of either enzyme in yeast results in galactose-dependent growth arrest of cultures despite the availability of an alternate carbon source. In contrast, loss of GALK in humans is not life-threatening, and in yeast has no impact on the growth of cultures challenged with galactose. Further, the growth of both GALT-null and GALE-null yeast challenged with galactose is rescued by loss of GALK, thereby implicating the GALK reaction product, gal-1P, for a role in the galactose-sensitivity of both strains. However, the nature of that relationship has remained unclear. Here we have developed and applied a doxycycline-repressible allele of galactokinase to define the quantitative relationship between galactokinase activity, gal-1P accumulation, and growth arrest of galactose-challenged GALT or GALE-deficient yeast. Our results demonstrate a clear threshold relationship between gal-1P accumulation and galactose-mediated growth arrest in both GALT-null and GALE-null yeast, however, the threshold for the two strains is distinct. Further, we tested the galactose-sensitivity of yeast double-null for GALT and GALE, and found that although loss of GALT barely changed accumulation of gal-1P, it significantly lowered the accumulation of UDP-gal, and also dramatically rescued growth of the GALE-null cells. Together, these data suggest that while gal-1P alone may account for the galactose-sensitivity of GALT-null cells, other factors, likely to include UDP-gal accumulation, must contribute to the galactose-sensitivity of GALE-null cells.

  1. Pathophysiological Consequences of a Break in S1P1-Dependent Homeostasis of Vascular Permeability Revealed by S1P1 Competitive Antagonism

    PubMed Central

    Bigaud, Marc; Dincer, Zuhal; Bollbuck, Birgit; Dawson, Janet; Beckmann, Nicolau; Beerli, Christian; Fishli-Cavelti, Gina; Nahler, Michaela; Angst, Daniela; Janser, Philipp; Otto, Heike; Rosner, Elisabeth; Hersperger, Rene; Bruns, Christian; Quancard, Jean

    2016-01-01

    Rational Homeostasis of vascular barriers depends upon sphingosine 1-phosphate (S1P) signaling via the S1P1 receptor. Accordingly, S1P1 competitive antagonism is known to reduce vascular barrier integrity with still unclear pathophysiological consequences. This was explored in the present study using NIBR-0213, a potent and selective S1P1 competitive antagonist. Results NIBR-0213 was tolerated at the efficacious oral dose of 30 mg/kg BID in the rat adjuvant-induced arthritis (AiA) model, with no sign of labored breathing. However, it induced dose-dependent acute vascular pulmonary leakage and pleural effusion that fully resolved within 3–4 days, as evidenced by MRI monitoring. At the supra-maximal oral dose of 300 mg/kg QD, NIBR-0213 impaired lung function (with increased breathing rate and reduced tidal volume) within the first 24 hrs. Two weeks of NIBR-0213 oral dosing at 30, 100 and 300 mg/kg QD induced moderate pulmonary changes, characterized by alveolar wall thickening, macrophage accumulation, fibrosis, micro-hemorrhage, edema and necrosis. In addition to this picture of chronic inflammation, perivascular edema and myofiber degeneration observed in the heart were also indicative of vascular leakage and its consequences. Conclusions Overall, these observations suggest that, in the rat, the lung is the main target organ for the S1P1 competitive antagonism-induced acute vascular leakage, which appears first as transient and asymptomatic but could lead, upon chronic dosing, to lung remodeling with functional impairments. Hence, this not only raises the question of organ specificity in the homeostasis of vascular barriers, but also provides insight into the pre-clinical evaluation of a potential safety window for S1P1 competitive antagonists as drug candidates. PMID:28005953

  2. Synthesis and evaluation of CS-2100, a potent, orally active and S1P(3)- sparing S1P(1) agonist.

    PubMed

    Nakamura, Tsuyoshi; Asano, Masayoshi; Sekiguchi, Yukiko; Mizuno, Yumiko; Tamaki, Kazuhiko; Nara, Futoshi; Kawase, Yumi; Yabe, Yoshiyuki; Nakai, Daisuke; Kamiyama, Emi; Urasaki-Kaneno, Yoko; Shimozato, Takaichi; Doi-Komuro, Hiromi; Kagari, Takashi; Tomisato, Wataru; Inoue, Ryotaku; Nagasaki, Miyuki; Yuita, Hiroshi; Oguchi-Oshima, Keiko; Kaneko, Reina; Nishi, Takahide

    2012-05-01

    Modulators of sphingosine phosphate receptor-1 (S1P(1)) have recently been focused as a suppressant of autoimmunity. We have discovered a 4-ethylthiophene-based S1P(1) agonist 1-({4-Ethyl-5-[5-(4-phenoxyphenyl)-1,2,4-oxadiazol-3-yl]-2-thienyl}methyl)azetidine-3-carboxylic acid (CS-2100, 8) showing potent S1P(1) agonist activity against S1P(3) and an excellent in vivo potency. We report herein the synthesis of CS-2100 (8) and pharmacological effects such as S1P(1) and S1P(3) agonist activity in vitro, peripheral blood lymphocyte lowering effects and the suppressive effects on adjuvant-induced arthritis and experimental autoimmune encephalomyelitis (EAE) in animal models. The pharmacokinetic data were also reported. CS-2100 (8) had >5000-fold greater agonist activity for human S1P(1) (EC(50); 4.0 nM) relative to S1P(3) (EC(50); >20,000 nM). Following administration of single oral doses of 0.1 and 1 mg/kg of CS-2100 (8) in rats, lymphocyte counts decreased significantly, with a nadir at 8 and/or 12 h post-dose and recovery to vehicle control levels by 24-48 h post-dose. CS-2100 (8) is efficacious in the adjuvant-induced arthritis model in rats (ID(50); 0.44 mg/kg). In the EAE model compared to the vehicle-treated group, significant decreases in the cumulative EAE scores were observed for 0.3 and 1 mg/kg CS-2100 (8) groups in mice. While CS-2100 (8) showed potent efficacy in various animal disease models, it was also revealed that the central 1,2,4-oxadiazole ring of CS-2100 (8) was decomposed by enterobacteria in intestine of rats and monkeys, implicating the latent concern about an external susceptibility in its metabolic process in the upcoming clinical studies.

  3. Abdominal paraganglioma in a young woman with 1p36 deletion syndrome.

    PubMed

    Murakoshi, Miki; Takasawa, Kei; Nishioka, Masato; Asakawa, Masahiro; Kashimada, Kenichi; Yoshimoto, Takanobu; Yamamoto, Toshiyuki; Takekoshi, Kazuhiro; Ogawa, Yoshihiro; Shimohira, Masayuki

    2017-02-01

    1p36 deletion syndrome is the most common terminal deletion syndrome, and the genomic regions that contribute to specific 1p36 deletion syndrome-related phenotypes were recently identified. Deletions in the 1p36 region have been documented in various tumor tissues, which indicates correlation between loss of heterozygosity of 1p36 and tumor development, and the existence of tumor suppressors in this region. Therefore, it was suspected that patients with 1p36 deletion syndrome have a higher risk of tumor development; however, only a few child cases of neuroblastoma with 1p36 deletion syndrome have been reported. We report the first case of 1p36 deletion syndrome with paraganglioma (PGL) and include genetic investigation. The 24-year-old woman with 1p36 deletion syndrome had severe intellectual disability, dilated cardiomyopathy, and distinct dysmorphic features, and presented with persistent vomiting accompanied by hypertension (178/115 mmHg). Abdominal CT revealed a 40 × 50 mm retroperitoneal mass and substantial elevations of plasma and urine norepinephrine (15.4 nmol/L and 1022 µmol/mol creatinine, respectively); abnormal uptake of (123) I-MIBG in the tumor led to PGL diagnosis. The patient was not able to have surgery because of substantial surgical risks; however, a combination of α- and β-blockade was effective for blood pressure control. Array CGH revealed a deletion over 4.5 Mb, from the 1p telomere but excluding the SDHB region. Comprehensive mutational analysis of PGL-associated genes (RET, VHL, TMEM127, MAX, and SDHA/B/C/D) was negative. These results indicate that the germline 1p36 deletion might be "1st hit" of tumor development, and PGL might be a novel complication of 1p36 deletion syndrome. © 2016 Wiley Periodicals, Inc.

  4. To stay or to leave: Stem cells and progenitor cells navigating the S1P gradient.

    PubMed

    Liu, Jingjing; Hsu, Andrew; Lee, Jen-Fu; Cramer, Daniel E; Lee, Menq-Jer

    2011-01-26

    Most hematopoietic stem progenitor cells (HSPCs) reside in bone marrow (BM), but a small amount of HSPCs have been found to circulate between BM and tissues through blood and lymph. Several lines of evidence suggest that sphingosine-1-phosphate (S1P) gradient triggers HSPC egression to blood circulation after mobilization from BM stem cell niches. Stem cells also visit certain tissues. After a temporary 36 h short stay in local tissues, HSPCs go to lymph in response to S1P gradient between lymph and tissue and eventually enter the blood circulation. S1P also has a role in the guidance of the primitive HSPCs homing to BM in vivo, as S1P analogue FTY720 treatment can improve HSPC BM homing and engraftment. In stress conditions, various stem cells or progenitor cells can be attracted to local injured tissues and participate in local tissue cell differentiation and tissue rebuilding through modulation the expression level of S1P(1), S1P(2) or S1P(3) receptors. Hence, S1P is important for stem cells circulation in blood system to accomplish its role in body surveillance and injury recovery.

  5. To fingolimod and beyond: The rich pipeline of drug candidates that target S1P signaling.

    PubMed

    Chew, Wee Siong; Wang, Wei; Herr, Deron R

    2016-11-01

    Sphingosine 1-phosphate (S1P) is an extracellular lipid signaling molecule that acts as a selective, high-affinity ligand for a family of five G protein-coupled receptors. This signaling system was first identified twenty years ago, and has since been shown to regulate a diverse range of physiological processes and disease states, such as cardiovascular development, immune function, hypoxic responses, and cancer. The therapeutic potential of targeting this system took center stage when it was demonstrated that the immune modulator, fingolimod (FTY720/Gilenya), exerts it lymphopenic effect by acting on S1P receptors, primarily on S1P receptor 1 (S1P1). In 2010, fingolimod became the first oral medication approved for the treatment of multiple sclerosis (MS). Since then, second-generation S1P receptor modulators have been under development in an effort to provide improved safety and efficacy profiles for MS, and to broaden their use to other autoimmune indications. Beyond the development of S1P1-modulators, there has been considerable effort in targeting other components of the S1P signaling pathway for the treatment of other diseases, such as cardiovascular disease, sepsis, and cancer. This manuscript provides an overview of the clinical and preclinical development of drugs targeting S1P signaling.

  6. Inp1p is a peroxisomal membrane protein required for peroxisome inheritance in Saccharomyces cerevisiae.

    PubMed

    Fagarasanu, Monica; Fagarasanu, Andrei; Tam, Yuen Yi C; Aitchison, John D; Rachubinski, Richard A

    2005-06-06

    Cells have evolved molecular mechanisms for the efficient transmission of organelles during cell division. Little is known about how peroxisomes are inherited. Inp1p is a peripheral membrane protein of peroxisomes of Saccharomyces cerevisiae that affects both the morphology of peroxisomes and their partitioning during cell division. In vivo 4-dimensional video microscopy showed an inability of mother cells to retain a subset of peroxisomes in dividing cells lacking the INP1 gene, whereas cells overexpressing INP1 exhibited immobilized peroxisomes that failed to be partitioned to the bud. Overproduced Inp1p localized to both peroxisomes and the cell cortex, supporting an interaction of Inp1p with specific structures lining the cell periphery. The levels of Inp1p vary with the cell cycle. Inp1p binds Pex25p, Pex30p, and Vps1p, which have been implicated in controlling peroxisome division. Our findings are consistent with Inp1p acting as a factor that retains peroxisomes in cells and controls peroxisome division. Inp1p is the first peroxisomal protein directly implicated in peroxisome inheritance.

  7. Basis for regulated RNA cleavage by functional analysis of RNase L and Ire1p.

    PubMed Central

    Dong, B; Niwa, M; Walter, P; Silverman, R H

    2001-01-01

    RNase L and Ire1p are members of a superfamily of regulated endoribonucleases that play essential roles in mediating diverse types of cellular stress responses. 2'-5' oligoadenylates, produced in response to interferon treatment and viral double-stranded RNA, are necessary to activate RNase L. In contrast, unfolded proteins in the endoplasmic reticulum activate Ire1p, a transmembrane serine/threonine kinase and endoribonuclease. To probe their similarities and differences, molecular properties of wild-type and mutant forms of human RNase L and yeast Ire1p were compared. Surprisingly, RNase L and Ire1p showed mutually exclusive RNA substrate specificity and partially overlapping but not identical requirements for phylogenetically conserved amino acid residues in their nuclease domains. A functional model for RNase L was generated based on the comparative analysis with Ire1p that assigns novel roles for ankyrin repeats and kinase-like domains. PMID:11333017

  8. Evidence for the h_b(1P) meson in the decay Upsilon(3S) --> pi0 h_b(1P)

    SciTech Connect

    Lees, J.P.

    2011-08-12

    Using a sample of 122 million {Upsilon}(3S) events recorded with the BABAR detector at the PEP-II asymmetric-energy e{sup +}e{sup -} collider at SLAC, we search for the h{sub b}(1P) spin-singlet partner of the P-wave {chi}{sub b}(1P) states in the sequential decay {Upsilon}(3S) {yields} {pi}{sup 0}h{sub b}(1P), h{sub b}(1P) {yields} {gamma}{eta}{sub b}(1S). We observe an excess of events above background in the distribution of the recoil mass against the {pi}{sup 0} at mass 9902 {+-} 4(stat.) {+-} 1(syst.) MeV/c{sup 2}. The width of the observed signal is consistent with experimental resolution, and its significance is 3.0 {sigma}, including systematic uncertainties. We obtain the value (3.7 {+-} 1.1 (stat.) {+-} 0.7 (syst.)) x 10{sup -4} for the product branching fraction {Beta}({Upsilon}(3S) {yields} {pi}{sup 0}h{sub b}) x {Beta}(h{sub b} {yields} {gamma}{eta}{sub b}).

  9. Mild craniosynostosis with 1p36.3 trisomy and 1p36.3 deletion syndrome caused by familial translocation t(Y;1).

    PubMed

    Hiraki, Yoko; Fujita, Hiroko; Yamamori, Shunji; Ohashi, Hirofumi; Eguchi, Maki; Harada, Naoki; Mizuguchi, Takeshi; Matsumoto, Naomichi

    2006-08-15

    We report on a 20-year-old man and a 16-year-old woman with a chromosomal imbalance derived from a balanced translocation, t(Y;1)(q12;p36.3) of the father. The man had a partial trisomy for 1p36.3-pter [46,X,der(Y)t(Y:1)(q12;p36.3)] and mild craniosynostosis of metopic and sagittal sutures as well as a borderline mental impairment, while the woman with a deletion for 1p36.3-pter [46,XX,der(1)t(Y;1)(q12;p36.3)] showed dysmorphic face with large anterior fontanel and severe developmental delay. Fluorescence in situ hybridization (FISH) showed that his trisomy spanned the 5.3-Mb region from 1p telomere harboring the critical region for craniosynostosis. To our knowledge, the man is the first case of a pure type of simple 1p36.3 trisomy as the effect of heterochromatic Yq12-qter deletion likely does not affect phenotype.

  10. The yeast Sks1p kinase signaling network regulates pseudohyphal growth and glucose response.

    PubMed

    Johnson, Cole; Kweon, Hye Kyong; Sheidy, Daniel; Shively, Christian A; Mellacheruvu, Dattatreya; Nesvizhskii, Alexey I; Andrews, Philip C; Kumar, Anuj

    2014-03-01

    The yeast Saccharomyces cerevisiae undergoes a dramatic growth transition from its unicellular form to a filamentous state, marked by the formation of pseudohyphal filaments of elongated and connected cells. Yeast pseudohyphal growth is regulated by signaling pathways responsive to reductions in the availability of nitrogen and glucose, but the molecular link between pseudohyphal filamentation and glucose signaling is not fully understood. Here, we identify the glucose-responsive Sks1p kinase as a signaling protein required for pseudohyphal growth induced by nitrogen limitation and coupled nitrogen/glucose limitation. To identify the Sks1p signaling network, we applied mass spectrometry-based quantitative phosphoproteomics, profiling over 900 phosphosites for phosphorylation changes dependent upon Sks1p kinase activity. From this analysis, we report a set of novel phosphorylation sites and highlight Sks1p-dependent phosphorylation in Bud6p, Itr1p, Lrg1p, Npr3p, and Pda1p. In particular, we analyzed the Y309 and S313 phosphosites in the pyruvate dehydrogenase subunit Pda1p; these residues are required for pseudohyphal growth, and Y309A mutants exhibit phenotypes indicative of impaired aerobic respiration and decreased mitochondrial number. Epistasis studies place SKS1 downstream of the G-protein coupled receptor GPR1 and the G-protein RAS2 but upstream of or at the level of cAMP-dependent PKA. The pseudohyphal growth and glucose signaling transcription factors Flo8p, Mss11p, and Rgt1p are required to achieve wild-type SKS1 transcript levels. SKS1 is conserved, and deletion of the SKS1 ortholog SHA3 in the pathogenic fungus Candida albicans results in abnormal colony morphology. Collectively, these results identify Sks1p as an important regulator of filamentation and glucose signaling, with additional relevance towards understanding stress-responsive signaling in C. albicans.

  11. The Saccharomyces cerevisiae protein Stm1p facilitates ribosome preservation during quiescence

    SciTech Connect

    Van Dyke, Natalya; Chanchorn, Ekkawit; Van Dyke, Michael W.

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer Stm1p confers increased resistance to the macrolide starvation-mimic rapamycin. Black-Right-Pointing-Pointer Stm1p maintains 80S ribosome integrity during stationary phase-induced quiescence. Black-Right-Pointing-Pointer Stm1p facilitates polysome formation following quiescence exit. Black-Right-Pointing-Pointer Stm1p facilitates protein synthesis following quiescence exit. Black-Right-Pointing-Pointer Stm1p is a ribosome preservation factor under conditions of nutrient deprivation. -- Abstract: Once cells exhaust nutrients from their environment, they enter an alternative resting state known as quiescence, whereby proliferation ceases and essential nutrients are obtained through internal stores and through the catabolism of existing macromolecules and organelles. One example of this is ribophagy, the degradation of ribosomes through the process of autophagy. However, some ribosomes need to be preserved for an anticipated recovery from nutrient deprivation. We found that the ribosome-associated protein Stm1p greatly increases the quantity of 80S ribosomes present in quiescent yeast cells and that these ribosomes facilitate increased protein synthesis rates once nutrients are restored. These findings suggest that Stm1p can act as a ribosome preservation factor under conditions of nutrient deprivation and restoration.

  12. Lack of evidence for monosomy 1p36 in patients with Prader-Willi-like phenotype.

    PubMed

    Rodríguez, V R; Mazzucato, L F; Pina-Neto, J M

    2008-08-01

    Monosomy 1p36 is the most common subtelomeric microdeletion syndrome with an incidence rate estimated to be 1 in 5000 births. A hypothesis of a similarity between patients with 1p36 deletion and those with Prader-Willi syndrome and the existence of two different phenotypes for 1p36 microdeletion has been suggested. The main objective of the present study was to determine the existence of 1p36 microdeletion in a sample of patients with mental retardation, obesity and hyperphagia who tested negative by the methylation test for Prader-Willi syndrome. Sixteen patients (7 females, 9 males), 16-26 years old, were evaluated with high-resolution cytogenetic analysis at 550-850 band levels and with 11 polymorphic microsatellite markers located in the 1p36 region. All patients had normal cytogenetic and molecular results. The results obtained by high-resolution cytogenetic methodology were confirmed by the molecular analyses. We did not detect a 1p36 microdeletion in 16 subjects with the Prader-Willi-like phenotype, which reinforces that no correlation seems to exist between Prader-Willi-like phenotype and the 1p36 microdeletion syndrome.

  13. Severe lysosomal storage disease of liver in del(1)(p36): a new presentation.

    PubMed

    Haimi, Motti; Iancu, Theodore C; Shaffer, Lisa G; Lerner, Aaron

    2011-01-01

    1p36 deletion is the most common terminal deletion syndrome with an estimated occurrence of 1:5000 live births. The deletion is of variable size. It usually involves less than 10 Mb in the 1pter-1p36.23 interval. Variability of the phenotype is partially related to the extent of the deletion. Some children with a 1p36 deletion were reported with obesity and hyperphagia, raising the question of possible phenotypic overlap with Prader-Willi syndrome. Correlation between presence of obesity and the size of the deletion has only been documented in one case. We report a 11-year-old girl with 1p36 deletion and the classical dysmorphological features. In late infancy, she developed an uncontrolled voracious appetite, overweight, truncal obesity and elevated serum transaminases. Liver biopsy disclosed severe steatosis. The hepatocytes contained accumulation of lipofuscins. Lipolysosomes were abnormally numerous and extremely enlarged. These features have not been previously reported in 1p36 deletion. Oligonucleotide-based microarray analysis showed a subtelomeric 2.2 Mb deletion at 1p36.33p36.32. This suggests that this chromosome segment is a critical region for obesity and hyperphagia. The accumulation in the liver with abnormal ultrastructure may be an additional feature of this form of syndromal obesity. 1p36 deletion syndrome should be considered in patients with obesity, hyperphagia and liver fat accumulation.

  14. Molecular characterization of a monosomy 1p36 presenting as an Aicardi syndrome phenocopy.

    PubMed

    Bursztejn, Anne-Claire; Bronner, Myriam; Peudenier, Sylviane; Grégoire, Marie-José; Jonveaux, Philippe; Nemos, Christophe

    2009-11-01

    Monosomy 1p36 is the most frequent terminal deletion known in Humans. Typical craniofacial features, developmental delay/mental retardation, seizures and sensorineural defects characterize 1p36 deletion syndrome. Aicardi syndrome (AIS) is a rare genetic disorder characterized by chorioretinal lacunae, corpus callosum agenesis and infantile spasms responsible for mental retardation. By screening DNA from diagnosed AIS patients with oligonucleotide array-based comparative genomic hybridization (aCGH), we report a 1p36 monosomy in this study. There were no other deletions or duplications. Regarding clinical criteria, the patient did not have the typical facial appearance commonly described for 1p36 monosomy patients. We showed that this 1p36 monosomy corresponded to combined interstitial and terminal de novo deletions of the chromosome 1 leading to an 11.73 Mb deletion confirmed with qPCR. By microsatellite markers and FISH analyses, we have concluded that this deletion occurred on maternal chromosome 1 during oogenesis. We did find some clinical features shared by the 1p36 monosomy and AIS: infantile spasms, corpus callosum dysgenesis, ophthalmological abnormalities, and skeletal malformations. To date, no relationship between these two phenotypes has been established. We conclude that the monosomy 1p36 should be considered in the differential diagnosis of AIS.

  15. Identification of a pepducin acting as S1P3 receptor antagonist.

    PubMed

    Severino, Beatrice; Incisivo, Giuseppina Maria; Fiorino, Ferdinando; Bertolino, Antonio; Frecentese, Francesco; Barbato, Francesco; Manganelli, Serena; Maggioni, Giada; Capasso, Domenica; Caliendo, Giuseppe; Santagada, Vincenzo; Sorrentino, Raffaella; Roviezzo, Fiorentina; Perissutti, Elisa

    2013-11-01

    Sphingosine-1-phosphate (S1P) is a bioactive lipid with key functions in the immune, inflammatory, and cardiovascular systems. S1P exerts its action through the interaction with a family of five known G protein-coupled receptors, named S1P(1-5). Among them, S1P(3) has been implicated in the pathological processes of a number of diseases, including sepsis and cancer. KRX-725 (compound 1) is a pepducin that mimics the effects of S1P by triggering specifically S1P(3). Here, aiming to identify novel S1P(3) antagonists, we carried out an alanine scanning analysis to address the contribution of the side chains of each amino acid residue to the peptide function. Then, deleted peptides from both the C- and N-terminus were prepared in order to determine the minimal sequence for activity and to identify the structural requirements for agonistic and, possibly, antagonistic behaviors. The pharmacological results of the Ala-scan derived compounds (2-10) suggested a high tolerance of the pepducin 1 to amino acid substitutions. Importantly, the deleted peptide 16 has the ability to inhibit, in a dose-dependent manner, both pepducin 1-induced vasorelaxation and fibroblast proliferation. Finally, a computational analysis was performed on the prepared compounds, showing that the supposed antagonists 16 and 17 appeared to be aligned with each other but not with the others. These results suggested a correlation between specific conformations and activities.

  16. Sec1p Binds to Snare Complexes and Concentrates at Sites of Secretion

    PubMed Central

    Carr, Chavela M.; Grote, Eric; Munson, Mary; Hughson, Frederick M.; Novick, Peter J.

    1999-01-01

    Proteins of the Sec1 family have been shown to interact with target-membrane t-SNAREs that are homologous to the neuronal protein syntaxin. We demonstrate that yeast Sec1p coprecipitates not only the syntaxin homologue Ssop, but also the other two exocytic SNAREs (Sec9p and Sncp) in amounts and in proportions characteristic of SNARE complexes in yeast lysates. The interaction between Sec1p and Ssop is limited by the abundance of SNARE complexes present in sec mutants that are defective in either SNARE complex assembly or disassembly. Furthermore, the localization of green fluorescent protein (GFP)-tagged Sec1p coincides with sites of vesicle docking and fusion where SNARE complexes are believed to assemble and function. The proposal that SNARE complexes act as receptors for Sec1p is supported by the mislocalization of GFP-Sec1p in a mutant defective for SNARE complex assembly and by the robust localization of GFP-Sec1p in a mutant that fails to disassemble SNARE complexes. The results presented here place yeast Sec1p at the core of the exocytic fusion machinery, bound to SNARE complexes and localized to sites of secretion. PMID:10427089

  17. Evidence that the Yeast Desaturase Ole1p Exists as a Dimer In Vivo

    SciTech Connect

    Lou, Y.; Shanklin, J.

    2010-06-18

    Desaturase enzymes are composed of two classes, the structurally well characterized soluble class found predominantly in the plastids of higher plants and the more widely distributed but poorly structurally defined integral membrane class. Despite their distinct evolutionary origins, the two classes both require an iron cofactor and molecular oxygen for activity and are inhibited by azide and cyanide, suggesting strong mechanistic similarities. The fact that the soluble desaturase is active as a homodimer prompted us test the hypothesis that an archetypal integral membrane desaturase from Saccharomyces cerevisiae, the {Delta}{sup o}-acyl-Co-A desaturase Ole1p, also exhibits a dimeric organization. Ole1p was chosen because it is one of the best characterized integral membrane desaturase and because it retains activity when fused with epitope tags. FLAG-Ole1p was detected by Western blotting of immunoprecipitates in which anti-Myc antibodies were used for capture from yeast extracts co-expressing Ole1p-Myc and Ole1p-FLAG. Interaction was confirmed by two independent bimolecular complementation assays (i.e. the split ubiquitin system and the split luciferase system). Co-expression of active and inactive Ole1p subunits resulted in an {approx}75% suppression of the accumulation of palmitoleic acid, demonstrating that the physiologically active form of Ole1p in vivo is the dimer in which both protomers must be functional.

  18. Preliminary observations on genetic alterations in pilocytic astrocytomas associated with neurofibromatosis 1.

    PubMed Central

    Tada, Kenji; Kochi, Masato; Saya, Hideyuki; Kuratsu, Jun-ichi; Shiraishi, Shoji; Kamiryo, Takanori; Shinojima, Naoki; Ushio, Yukitaka

    2003-01-01

    Neurofibromatosis 1 (NF1) is an autosomal dominant disorder that predisposes sufferers to various forms of neoplasia. Among affected individuals, 15%-20% develop astrocytomas, especially pilocytic astrocytomas (PA), which are benign and classified as grade I by the World Health Organization. They are generally well circumscribed, and their progression is slow. NF1-associated PAs (NF1-PAs) occasionally behave as aggressive tumors. To elucidate underlying genetic events in clinically progressive NF1-PAs, we performed molecular genetic analysis on 12 PAs, including 3 NF1-PAs, for pS3, p16, and epidermal growth factor receptor genes, as well as loss of heterozygosity (LOH) on chromosome 1p, 10, 17, and 19q. None of the obvious genetic alterations typically seen in higher grade astrocytomas were found in 9 sporadic PAs. However, in 2 of 3 NF1-PAs, microsatellite analysis showed LOH10, including the PTEN (phosphatase and tensin homolog deleted on chromosome 10) gene locus, despite the diagnosis of pilocytic astrocytoma;one of these also manifested homozygous deletion of the p16 gene. The other NF1-PA harbored only LOH of the NF1 gene locus (17q). Our preliminary results support the hypothesis that some NF1-PAs differ genetically from sporadic PAs. PMID:14565158

  19. Translocation involving 1p and 17q is a recurrent genetic alteration of human neuroblastoma cells

    SciTech Connect

    Savelyeva, L.; Corvi, R.; Schwab, M. )

    1994-08-01

    Human neuroblastoma cells often are monosomic for the distal portion of 1p (1p36). The authors report that the deleted 1p material in cells of neuroblastoma lines is preferentially replaced by material from chromosome 17, resulting from an unbalanced 1;17 translocation. Chromosome 17 often acquires instability, followed by the integration of fragments into various marker chromosomes. As a consequence, 17q material can increase over 17p material. The nonrandom frequency of 1;17 translocations appears to indicate an as-yet-undefined contribution to neuroblastoma development. 35 refs., 4 figs., 1 tab.

  20. Novel airway findings in a patient with 1p36 deletion syndrome.

    PubMed

    Ferril, Geoffrey R; Barham, Henry P; Prager, Jeremy D

    2014-01-01

    1p36 deletion syndrome comprises a phenotypic presentation that includes central nervous system, cardiac, and craniofacial anomalies. There has been no report of associated airway anomalies with this syndrome. We present here a case report and literature review. Prenatally, amniocentesis for chromosomal analysis was performed on our patient, with results consistent with 1p36 deletion syndrome. Respiratory distress and unsuccessful attempts at intubation prompted transfer to Children's Hospital of Colorado. Microlaryngoscopy was subsequently performed, revealing a persistent buccopharyngeal membrane and unidentifiable larynx. Emergent tracheostomy was then performed to secure the airway. Airway anomalies may be associated with 1p36 deletion syndrome.

  1. A patient with monosomy 1p36, atypical features and phenotypic similarities with Cantu syndrome.

    PubMed

    Tan, Tiong Yang; Bankier, Agnes; Slater, Howard R; Northrop, Emma L; Zacharin, Margaret; Savarirayan, Ravi

    2005-12-15

    We report on a 16-year-old boy with a distal 1p36 deletion with some clinical features consistent with Cantu syndrome (OMIM#239850). He also has hypercholesterolemia, type II diabetes, recurrent bony fractures, and non-alcoholic steatohepatitis, not previously described in either condition. The 1p36 deletion was detected in a screen of all chromosome subtelomeres using multiplex ligation-dependent probe amplification and was verified using FISH with a region-specific BAC clone. We suggest that patients suspected of having Cantu syndrome, especially those with unusual or more severe manifestations be analyzed for distal 1p36 deletions.

  2. Regulation of S1P receptors and sphingosine kinases expression in acute pulmonary endothelial cell injury

    PubMed Central

    Liu, Huiying; Zhang, Zili; Li, Puyuan; Yuan, Xin; Zheng, Jing; Liu, Jinwen

    2016-01-01

    Background Acute lung injury and acute respiratory distress syndrome (ALI/ARDS) is a severe clinical syndrome with mortality rate as high as 30–40%. There is no treatment yet to improve pulmonary endothelial barrier function in patients with severe pulmonary edema. Developing therapies to protect endothelial barrier integrity and stabilizing gas exchange is getting more and more attention. Sphingosine-1-phosphate (S1P) is able to enhance the resistance of endothelial cell barrier. S1P at physiological concentrations plays an important role in maintaining endothelial barrier function. Proliferation, regeneration and anti-inflammatory activity that mesenchymal stem cells (MSCs) exhibit make it possible to regulate the homeostatic control of S1P. Methods By building a pulmonary endothelial cell model of acute injury, we investigated the regulation of S1P receptors and sphingosine kinases expression by MSCs during the treatment of acute lung injury using RT-PCR, and investigated the HPAECs Micro-electronics impedance using Real Time Cellular Analysis. Results It was found that the down-regulation of TNF-α expression was more significant when MSC was used in combination with S1P. The combination effection mainly worked on S1PR2, S1PR3 and SphK2. The results show that when MSCs were used in combination with S1P, the selectivity of S1P receptors was increased and the homeostatic control of S1P concentration was improved through regulation of expression of S1P metabolic enzymes. Discussions The study found that, as a potential treatment, MSCs could work on multiple S1P related genes simultaneously. When it was used in combination with S1P, the expression regulation result of related genes was not simply the superposition of each other, but more significant outcome was obtained. This study establishes the experimental basis for further exploring the efficacy of improving endothelial barrier function in acute lung injury, using MSCs in combination with S1P and their

  3. Loss of CDC5 function in Saccharomyces cerevisiae leads to defects in Swe1p regulation and Bfa1p/Bub2p-independent cytokinesis.

    PubMed Central

    Park, Chong Jin; Song, Sukgil; Lee, Philip R; Shou, Wenying; Deshaies, Raymond J; Lee, Kyung S

    2003-01-01

    In many organisms, polo kinases appear to play multiple roles during M-phase progression. To provide new insights into the function of budding yeast polo kinase Cdc5p, we generated novel temperature-sensitive cdc5 mutants by mutagenizing the C-terminal domain. Here we show that, at a semipermissive temperature, the cdc5-3 mutant exhibited a synergistic bud elongation and growth defect with loss of HSL1, a component important for normal G(2)/M transition. Loss of SWE1, which phosphorylates and inactivates the budding yeast Cdk1 homolog Cdc28p, suppressed the cdc5-3 hsl1Delta defect, suggesting that Cdc5p functions at a point upstream of Swe1p. In addition, the cdc5-4 and cdc5-7 mutants exhibited chained cell morphologies with shared cytoplasms between the connected cell bodies, indicating a cytokinetic defect. Close examination of these mutants revealed delayed septin assembly at the incipient bud site and loosely organized septin rings at the mother-bud neck. Components in the mitotic exit network (MEN) play important roles in normal cytokinesis. However, loss of BFA1 or BUB2, negative regulators of the MEN, failed to remedy the cytokinetic defect of these mutants, indicating that Cdc5p promotes cytokinesis independently of Bfa1p and Bub2p. Thus, Cdc5p contributes to the activation of the Swe1p-dependent Cdc28p/Clb pathway, normal septin function, and cytokinesis. PMID:12586693

  4. A Role for Myosin-I in Actin Assembly through Interactions with Vrp1p, Bee1p, and the Arp2/3 Complex

    PubMed Central

    Evangelista, Marie; Klebl, Bert M.; Tong, Amy H.Y.; Webb, Bradley A.; Leeuw, Thomas; Leberer, Ekkehard; Whiteway, Malcolm; Thomas, David Y.; Boone, Charles

    2000-01-01

    Type I myosins are highly conserved actin-based molecular motors that localize to the actin-rich cortex and participate in motility functions such as endocytosis, polarized morphogenesis, and cell migration. The COOH-terminal tail of yeast myosin-I proteins, Myo3p and Myo5p, contains an Src homology domain 3 (SH3) followed by an acidic domain. The myosin-I SH3 domain interacted with both Bee1p and Vrp1p, yeast homologues of human WASP and WIP, adapter proteins that link actin assembly and signaling molecules. The myosin-I acidic domain interacted with Arp2/3 complex subunits, Arc40p and Arc19p, and showed both sequence similarity and genetic redundancy with the COOH-terminal acidic domain of Bee1p (Las17p), which controls Arp2/3-mediated actin nucleation. These findings suggest that myosin-I proteins may participate in a diverse set of motility functions through a role in actin assembly. PMID:10648568

  5. Observation of the h(c)(1P) Using e+ e- collisions above the DD threshold.

    PubMed

    Pedlar, T K; Cronin-Hennessy, D; Hietala, J; Dobbs, S; Metreveli, Z; Seth, K K; Tomaradze, A; Xiao, T; Martin, L; Powell, A; Wilkinson, G; Mendez, H; Ge, J Y; Miller, D H; Shipsey, I P J; Xin, B; Adams, G S; Hu, D; Moziak, B; Napolitano, J; Ecklund, K M; Insler, J; Muramatsu, H; Park, C S; Pearson, L J; Thorndike, E H; Ricciardi, S; Thomas, C; Artuso, M; Blusk, S; Mountain, R; Skwarnicki, T; Stone, S; Zhang, L M; Bonvicini, G; Cinabro, D; Lincoln, A; Smith, M J; Zhou, P; Zhu, J; Naik, P; Rademacker, J; Asner, D M; Edwards, K W; Randrianarivony, K; Tatishvili, G; Briere, R A; Vogel, H; Onyisi, P U E; Rosner, J L; Alexander, J P; Cassel, D G; Das, S; Ehrlich, R; Gibbons, L; Gray, S W; Hartill, D L; Heltsley, B K; Kreinick, D L; Kuznetsov, V E; Patterson, J R; Peterson, D; Riley, D; Ryd, A; Sadoff, A J; Shi, X; Sun, W M; Yelton, J; Rubin, P; Lowrey, N; Mehrabyan, S; Selen, M; Wiss, J; Libby, J; Kornicer, M; Mitchell, R E; Shepherd, M R; Tarbert, C M; Besson, D

    2011-07-22

    Using 586  pb(-1) of e+ e- collision data at E(c.m.) = 4170  MeV, produced at the Cornell Electron Storage Ring collider and collected with the CLEO-c detector, we observe the process e+ e- → π+ π- h(c)(1P). We measure its cross section to be 15.6±2.3±1.9±3.0  pb, where the third error is due to the external uncertainty on the branching fraction of ψ(2S) → π0 h(c)(1P), which we use for normalization. We also find evidence for e+ e- → ηh(c)(1P) at 4170 MeV at the 3σ level and see hints of a rise in the e+ e- → π+ π- h(c)(1P) cross section at 4260 MeV.

  6. Mini-Review: Monosomy 1p36 syndrome: reviewing the correlation between deletion sizes and phenotypes.

    PubMed

    Rocha, C F; Vasques, R B; Santos, S R; Paiva, C L A

    2016-02-22

    The major clinical features of monosomy 1p36 deletion are developmental delay and hypotonia associated with short stature and craniofacial dysmorphisms. The objective of this study was to review the cases of 1p36 deletion that was reported between 1999 and 2014, in order to identify a possible correlation between the size of the 1p36-deleted segment and the clinical phenotype of the disease. Scientific articles published in the (National Center for Biotechnology Information; NCBI http://www.ncbi.nlm.nih.gov/pubmed) and Scientific Electronic Library Online (www.scielo.com.br) databases were searched using key word combinations, such as "1p36 deletion", "monosomy 1p36 deletion", and "1p36 deletion syndrome". Articles in English or Spanish reporting the correlation between deletion sizes and the respective clinical phenotypes were retrieved, while letters, reviews, guidelines, and studies with mouse models were excluded. Among the 746 retrieved articles, only 17 (12 case reports and 5 series of cases), comprising 29 patients (9 males and 20 females, aged 0 months (neonate) to 22 years) bearing the 1p36 deletions and whose clinical phenotypes were described, met the inclusion criteria. The genotype-phenotype correlation in monosomy 1p36 is a challenge because of the variability in the size of the deleted segment, as well as in the clinical manifestations of similar size deletions. Therefore, the severity of the clinical features was not always associated with the deletion size, possibly because of the other influences, such as stochastic factors, epigenetic events, or reduced penetration of the deleted genes.

  7. Choroid plexus hyperplasia and monosomy 1p36: report of new findings.

    PubMed

    Puvabanditsin, Surasak; Garrow, Eugene; Patel, Neisha; D'Elia, Alexis; Zaafran, Ahmed; Phattraprayoon, Nanthida; Davis, Suzanne Elizabeth

    2008-08-01

    Monosomy 1p36 is a newly delineated multiple congenital anomalies/mental retardation syndrome characterized by mental retardation, growth delay, epilepsy, congenital heart defects, characteristic facial appearance, and precocious puberty. It is now considered to be one of the most common subtelomeric micro-deletion syndromes. This article reports new findings of choroid plexus hyperplasia and dextrocardia with situs solitus in a patient who had deletion of chromosome 1p26.33 with a brief review of the literature.

  8. [Turner syndrome and monosomy 1p36 deletion syndrome misdiagnosed as thyropenia: report of one case].

    PubMed

    Meng, Xubiao; Li, Zhiming; Liu, Tingting; Wen, Zhiming

    2013-12-01

    A 21-year-old woman with a short stature presented with primary amenorrhoea and a 45X karyotype, and comparative genomic hybridization revealed 1p36 deletion and abnormal genes in multiple chromosomes to support the diagnosis of Turner syndrome and monosomy 1p36 deletion syndrome. The main clinical features of this condition include microsomia, poor sexual development, menoschesis, gigantorectum, absence of internal genitalia, sometimes with thyropenia and low intelligence. This disease can be easily diagnosed for its heterogeneous clinical manifestations.

  9. FXR1P is a GSK3β substrate regulating mood and emotion processing

    PubMed Central

    Del’Guidice, Thomas; Latapy, Camille; Rampino, Antonio; Khlghatyan, Jivan; Lemasson, Morgane; Gelao, Barbara; Quarto, Tiziana; Rizzo, Giuseppe; Barbeau, Annie; Lamarre, Claude; Bertolino, Alessandro; Blasi, Giuseppe; Beaulieu, Jean-Martin

    2015-01-01

    Inhibition of glycogen synthase kinase 3β (GSK3β) is a shared action believed to be involved in the regulation of behavior by psychoactive drugs such as antipsychotics and mood stabilizers. However, little is known about the identity of the substrates through which GSK3β affects behavior. We identified fragile X mental retardation-related protein 1 (FXR1P), a RNA binding protein associated to genetic risk for schizophrenia, as a substrate for GSK3β. Phosphorylation of FXR1P by GSK3β is facilitated by prior phosphorylation by ERK2 and leads to its down-regulation. In contrast, behaviorally effective chronic mood stabilizer treatments in mice inhibit GSK3β and increase FXR1P levels. In line with this, overexpression of FXR1P in the mouse prefrontal cortex also leads to comparable mood-related responses. Furthermore, functional genetic polymorphisms affecting either FXR1P or GSK3β gene expression interact to regulate emotional brain responsiveness and stability in humans. These observations uncovered a GSK3β/FXR1P signaling pathway that contributes to regulating mood and emotion processing. Regulation of FXR1P by GSK3β also provides a mechanistic framework that may explain how inhibition of GSK3β can contribute to the regulation of mood by psychoactive drugs in mental illnesses such as bipolar disorder. Moreover, this pathway could potentially be implicated in other biological functions, such as inflammation and cell proliferation, in which FXR1P and GSK3 are known to play a role. PMID:26240334

  10. Second generation S1P pathway modulators: research strategies and clinical developments.

    PubMed

    Bigaud, Marc; Guerini, Danilo; Billich, Andreas; Bassilana, Frederic; Brinkmann, Volker

    2014-05-01

    Multiple Sclerosis (MS) is a chronic autoimmune disorder affecting the central nervous system (CNS) through demyelination and neurodegeneration. Until recently, major therapeutic treatments have relied on agents requiring injection delivery. In September 2010, fingolimod/FTY720 (Gilenya, Novartis) was approved as the first oral treatment for relapsing forms of MS. Fingolimod causes down-modulation of S1P1 receptors on lymphocytes which prevents the invasion of autoaggressive T cells into the CNS. In astrocytes, down-modulation of S1P1 by the drug reduces astrogliosis, a hallmark of MS, thereby allowing restoration of productive astrocyte communication with other neural cells and the blood brain barrier. Animal data further suggest that the drug directly supports the recovery of nerve conduction and remyelination. In human MS, such mechanisms may explain the significant decrease in the number of inflammatory markers on brain magnetic resonance imaging in recent clinical trials, and the reduction of brain atrophy by the drug. Fingolimod binds to 4 of the 5 known S1P receptor subtypes, and significant efforts were made over the past 5 years to develop next generation S1P receptor modulators and determine the minimal receptor selectivity needed for maximal therapeutic efficacy in MS patients. Other approaches considered were competitive antagonists of the S1P1 receptor, inhibitors of the S1P lyase to prevent S1P degradation, and anti-S1P antibodies. Below we discuss the current status of the field, and the functional properties of the most advanced compounds. This article is part of a Special Issue entitled New Frontiers in Sphingolipid Biology.

  11. The c.*229C > T gene polymorphism in 3'UTR region of the topoisomerase IIβ binding protein 1 gene and LOH in BRCA1/2 regions and their effect on the risk and progression of human laryngeal carcinoma.

    PubMed

    Starska, Katarzyna; Forma, Ewa; Nowacka-Zawisza, Maria; Lewy-Trenda, Iwona; Ciesielski, Piotr; Pietruszewska, Wioletta; Skóra, Michał; Bryś, Magdalena

    2016-04-01

    Topoisomerase IIβ binding protein 1 (TopBP1), a multiple-BRCT-domain, protein plays crucial roles in chromosome replication, DNA damage repair, apoptosis, and cell cycle checkpoint signalling. The aim of this study was to identify five SNPs at loci potentially located in the 3'UTR region of the TopBP1 gene (rs185903567, rs116645643, rs115160714, rs116195487, rs112843513), their relationship with the risk of squamous cell laryngeal cancer (SCLC), tumor invasiveness, and prognosis. Genotyping was performed in 323 genetically unrelated individuals with SCLC and 418 randomly selected healthy volunteers. Allele-specific TopBP1 mRNA and protein expressions were determined by using real-time PCR and Western blotting techniques, respectively. LOH in BRCA1/BRCA2 was determined by using microsatellite markers. Compared to homozygous common allele carriers, heterozygosity for the T variant was associated with increased risk of SCLC (adjusted odds ratio [OR] = 9.83, 95 % confidence interval [CI]: 3.12-22.16, p dominant < 0.0001). The presence of risk allele at rs115160714 TopBP1 determined a higher incidence of nodal metastases (OR = 7.98, 95 % CI: 3.94-16.00, p = 0.001) and higher tumor grade (OR = 6.48, 95 % CI: 0.86-48.01, p = 0.03). The heterozygotes displayed diffuse tumor growth with no distinct borderline (OR = 3.10, 95 % Cl: 0.92-10.62, p = 0.049) and higher depth of invasion (OR = 2.66, 95 % Cl: 0.78-9.03, p = 0.04). Relationships were also identified between TopBP1 mRNA/protein expression and overall survival (p < 0.0001). The incidence of LOH in BRCA1/BRCA2 was significantly related to higher tumor grade and TFG (p < 0.05). The results of this study suggest that rs115160714 TopBP1 may be a genetic marker of etiology and progression in laryngeal cancer.

  12. Identification of proximal 1p36 deletions using array-CGH: a possible new syndrome.

    PubMed

    Kang, S-H L; Scheffer, A; Ou, Z; Li, J; Scaglia, F; Belmont, J; Lalani, S R; Roeder, E; Enciso, V; Braddock, S; Buchholz, J; Vacha, S; Chinault, A C; Cheung, S W; Bacino, C A

    2007-10-01

    Monosomy 1p36 is the most common terminal deletion syndrome with an estimated occurrence of 1:5000 live births. Typically, the deletions span <10 Mb of 1pter-1p36.23 and result in mental retardation, developmental delay, sensorineural hearing loss, seizures, cardiomyopathy and cardiovascular malformations, and distinct facies including large anterior fontanel, deep-set eyes, straight eyebrows, flat nasal bridge, asymmetric ears, and pointed chin. We report five patients with 'atypical' proximal interstitial deletions from 1p36.23-1p36.11 using array-comparative genomic hybridization. Four patients carry large overlapping deletions of approximately 9.38-14.69 Mb in size, and one patient carries a small 2.97 Mb deletion. Interestingly, these patients manifest many clinical characteristics that are different from those seen in 'classical' monosomy 1p36 syndrome. The clinical presentation in our patients included: pre- and post-natal growth deficiency (mostly post-natal), feeding difficulties, seizures, developmental delay, cardiovascular malformations, microcephaly, limb anomalies, and dysmorphic features including frontal and parietal bossing, abnormally shaped and posteriorly rotated ears, hypertelorism, arched eyebrows, and prominent and broad nose. Most children also displayed hirsutism. Based on the analysis of the clinical and molecular data from our patients and those reported in the literature, we suggest that this chromosomal abnormality may constitute yet another deletion syndrome distinct from the classical distal 1p36 deletion syndrome.

  13. Rtp1p is a karyopherin-like protein required for RNA polymerase II biogenesis.

    PubMed

    Gómez-Navarro, Natalia; Peiró-Chova, Lorena; Rodriguez-Navarro, Susana; Polaina, Julio; Estruch, Francisco

    2013-05-01

    The assembly and nuclear transport of RNA polymerase II (RNA pol II) are processes that require the participation of many auxiliary factors. In a yeast genetic screen, we identified a previously uncharacterized gene, YMR185w (renamed RTP1), which encodes a protein required for the nuclear import of RNA pol II. Using protein affinity purification coupled to mass spectrometry, we identified interactions between Rtp1p and members of the R2TP complex. Rtp1p also interacts, to a different extent, with several RNA pol II subunits. The pattern of interactions is compatible with a role for Rtp1p as an assembly factor that participates in the formation of the Rpb2/Rpb3 subassembly complex and its binding to the Rpb1p-containing subcomplex. Besides, Rtp1p has a molecular architecture characteristic of karyopherins, composed of HEAT repeats, and is able to interact with phenylalanine-glycine-containing nucleoporins. Our results define Rtp1p as a new component of the RNA pol II biogenesis machinery that plays roles in subunit assembly and likely in transport through the nuclear pore complex.

  14. Yeast Uri1p promotes translation initiation and may provide a link to cotranslational quality control.

    PubMed

    Deplazes, Anna; Möckli, Natalie; Luke, Brian; Auerbach, Daniel; Peter, Matthias

    2009-05-20

    Translation initiation in eukaryotes is accomplished by a large set of translation initiation factors, some of which are regulated by signals monitoring intracellular and environmental conditions. Here, we show that Uri1p is required for efficient translation initiation in budding yeast. Indeed, uri1Delta cells are slow growing, sensitive to translation inhibitors and they exhibit an increased 80S peak in polysome profiles. Moreover, GCN4 translation is derepressed in uri1Delta cells, strongly supporting an initiation defect. Genetic and biochemical experiments indicate that Uri1p interacts with the translation initiation factor eIF1A and promotes ternary complex (TC) recruitment to the 40S subunit. Interestingly, we found that Uri1p is also part of a chaperone-network, including the prefoldin Pfd6p and several other proteins involved in cotranslational quality control such as the ribosome-associated Hsp70 chaperone Ssb1p, the Hsp40 Sis1p and the translation elongation factor eEF1A. Together with genetic data, these interactions indicate that Uri1p may coordinate translation initiation and cotranslational quality control.

  15. Bre1p-mediated histone H2B ubiquitylation regulates apoptosis in Saccharomyces cerevisiae.

    PubMed

    Walter, David; Matter, Anja; Fahrenkrog, Birthe

    2010-06-01

    BRE1 encodes an E3 ubiquitin protein ligase that is required for the ubiquitylation of histone H2B at lysine 123 (K123). Ubiquitylation of this histone residue is involved in a variety of cellular processes including gene activation and gene silencing. Abolishing histone H2B ubiquitylation also confers X-ray sensitivity and abrogates checkpoint activation after DNA damage. Here we show that Saccharomyces cerevisiae Bre1p exhibits anti-apoptotic activity in yeast and that this is linked to histone H2B ubiquitylation. We found that enhanced levels of Bre1p protect from hydrogen-peroxide-induced cell death, whereas deletion of BRE1 enhances cell death. Moreover, cells lacking Bre1p show reduced lifespan during chronological ageing, a physiological apoptotic condition in yeast. Importantly, the resistance against apoptosis is conferred by histone H2B ubiquitylation mediated by the E3 ligase activity of Bre1p. Furthermore, we found that the death of Deltabre1 cells depends on the yeast caspase Yca1p, because Deltabre1 cells exhibit increased caspase activity when compared with wild-type cells, and deletion of YCA1 leads to reduced apoptosis sensitivity of cells lacking Bre1p.

  16. Case report of individual with cutaneous immunodeficiency and novel 1p36 duplication

    PubMed Central

    Hatter, Alyn D; Soler, David C; Curtis, Christine; Cooper, Kevin D; McCormick, Thomas S

    2016-01-01

    Introduction Crusted or Norwegian scabies is an infectious skin dermatopathology usually associated with an underlying immunodeficiency condition. It is caused when the mite Sarcoptes scabiei infects the skin, and the immune system is unable to control its spread, leading to a massive hyperinfestation with a simultaneous inflammatory and hyperkeratotic reaction. This is the first report of a novel 1p36 duplication associated with a recurrent infection of crusted scabies. Case report We describe a 34-year-old patient with a cutaneous immunodeficiency characterized by recurrent crusted scabies infestation, diffuse tinea, and recurrent staphylococcal cellulitis, who we suspected had an undiagnosed syndrome. The patient also suffered from mental retardation, renal failure, and premature senescence. A cytogenetic fluorescence in situ hybridization analysis revealed a 9.34 Mb duplication within the short (p) arm of chromosome 1, precisely from 1p36.11 to 1p36.21, with an adjacent 193 kb copy gain entirely within 1p36.11. In addition, chromosome 4 had a 906 kb gain in 4p16.1 and chromosome 9 had a 81 kb copy gain in 9p24.3. Over 100 genes localized within these duplicated regions. Gene expression array revealed 82 genes whose expression changed >1.5-fold compared to a healthy age-matched skin control, but among them only the lipolytic enzyme arylacetamide deacetylase-like 3 was found within the duplicated 1p36 region of chromosome 1. Discussion Although genetic duplications in the 1p36 region have been previously described, our report describes a novel duplicative variant within the 1p36 region. The patient did not have a past history of immunosuppression but was afflicted by a recurrent case of crusted scabies, raising the possibility that the recurrent infection was associated with the 1p36 genetic duplication. Conclusion To our knowledge, the specific duplicated sequence between 1p36.11 and p36.21 found in our patient has never been previously reported. We reviewed and

  17. Mild developmental delay and obesity in two patients with mosaic 1p36 deletion syndrome.

    PubMed

    Shimada, Shino; Maegaki, Yoshihiro; Osawa, Makiko; Yamamoto, Toshiyuki

    2014-02-01

    We identified mosaic 1p36 deletions in two patients with developmental delay, distinctive features, and obesity, who can walk alone and communicate with others. Thus, their neurological defects are milder than those in typical patients with 1p36 deletion syndrome because most patients with 1p36 deletion cannot acquire expressive language. Chromosomal microarray testing revealed 3.0 and 4.5 Mb aberrations in the subtelomeric region of the short arm of chromosome 1. Mean signal ratios of the identified aberrations were -0.4 and -0.5, indicating mosaicism, which was confirmed by fluorescence in situ hybridization analysis with a mosaic ratio of 70% and 77%, respectively. Previous studies demonstrated that deletion of the distal 2-3 Mb region would be responsible for hyperphagia and obesity seen in patients. On the other hand, the severity of the neurological defect often correlates with the size of the terminal deletion of 1p36, and patients with larger deletions of 1p36 would usually show severely impaired developmental milestones and be immobile and aphasic. In such cases, hyperphagia and obesity could be clinically masked. In this study, two patients with mosaic deletions of 1p36 showed obesity as a consequence of hyperphagia. This study suggests that patients with 1p36 deletion would be at risk for hyperphagia and obesity when they have both risk factors, that is, (1) deletions including the 2-3 Mb critical region and (2) milder phenotypes that allow them to reach food on their own and to overeat.

  18. Epilepsy and neurological findings in 11 individuals with 1p36 deletion syndrome.

    PubMed

    Kurosawa, Kenji; Kawame, Hiroshi; Okamoto, Nobuhiko; Ochiai, Yukikatsu; Akatsuka, Akira; Kobayashi, Masahisa; Shimohira, Masayuki; Mizuno, Seiji; Wada, Kazuko; Fukushima, Yoshimitsu; Kawawaki, Hisashi; Yamamoto, Toshiyuki; Masuno, Mitsuo; Imaizumi, Kiyoshi; Kuroki, Yoshikazu

    2005-08-01

    The 1p36 deletion syndrome is a newly delineated multiple congenital anomalies/mental retardation syndrome characterized by mental retardation, growth delay, epilepsy, congenital heart defects, characteristic facial appearance, and precocious puberty. We analyzed 11 patients by fluorescence in situ hybridization (FISH) using commercially available bacterial artificial chromosome and P1-derived artificial chromosome genomic clones to define the chromosomal deletion responsible for the 1p36 deletion syndrome. Cytogenetic investigation revealed two cases with a terminal deletion of 1p36. Nine patients had an apparently normal karyotype with standard G-bands by trypsin using Giemsa (GTG), but FISH screening with the highly polymorphic genetic marker D1Z2, which is mapped to 1p36.3 and contains an unusual reiterated 40-bp variable number tandem repeat, revealed a submicroscopic deletion. All patients had severe to profound mental retardation. Based on the University of California Santa Cruz Genome Browser, we constructed a deletion map and analyzed the relationship between neurological findings and chromosomal deletions for the 11 cases. Six cases had intractable epilepsy and three had no seizures. The common deletion interval was about 1 million base pairs (Mbp) located between RP11-82D16 and RP4-785P20 (Rho guanine exchange factor (GEF) 16). The severity of clinical symptoms correlates with the size of the deletion. This is demonstrated by the 3 patients with at least 8Mbp deletions that display profound mental retardation and congenital heart defects. Although haploinsufficiency of the potassium channel beta-subunit (KCNAB2) is thought to be responsible for intractable seizures in the 1p36 deletion syndrome, this was not the case for 3 of the 11 patients in this study. Further investigation of the 1p36 region is necessary to allow identification of genes responsible for the 1p36 deletion syndrome.

  19. SIRT1 mediates Sphk1/S1P-induced proliferation and migration of endothelial cells.

    PubMed

    Gao, Zhan; Wang, Hua; Xiao, Feng-Jun; Shi, Xue-Feng; Zhang, Yi-Kun; Xu, Qin Qin; Zhang, Xiao-Yan; Ha, Xiao-Qin; Wang, Li-Sheng

    2016-05-01

    Angiogenesis is one of the most important components of embryonic organ formation and vessel growth after birth. Sphingosine kinase 1 (Sphk1) and S1P has been confirmed to participate in various cell signaling pathways and physiological processes including neovascularisation. However, the mechanisms that Sphk1/S1P regulates neovascularisation remain unclear. In this study, we elucidated that Sphk1/S1P upregulates sirtuin 1 (SIRT1), a NAD+ dependent deacetylases protease which exerts multiple cellular functions, to regulate the proliferation and migration of endothelial cells. By using CCK8 and Transwell assays, we demonstrated that Sphk1 and SIRT1 knockdown could significantly decrease proliferation and migration of HUVEC cells. Sphk1 inhibition results in SIRT1 downregulation which could be reversed by exogenous S1P in HUVEC cells. Treatment of HUVECs with S1P reverses the impaired proliferation and migration caused by SIRT1 knockdown. Furthermore, Sphk1 knockdown inhibits the phosphorylation of P38 MAPK, ERK and AKT. Treatment of HUVECs with PD98059, SB203580 and Wortmannin, which are the inhibitors of ERK, P38 MAPK and AKT respectively, resulted in decreased SIRT1 expression and reduced migration of HUVEC cells. Thus, we conclude that Sphk1/S1P induces SIRT1 upregulation through multiple pathways including P38 MAPK, ERK and AKT signals. This is the first report to disclose the existence and roles of Sphk1/S1P/SIRT1 axis in regulation of endothelial cell proliferation and migration, which may provide a theoretical basis for angiogenesis.

  20. Effects of S1P on skeletal muscle repair/regeneration during eccentric contraction.

    PubMed

    Sassoli, Chiara; Formigli, Lucia; Bini, Francesca; Tani, Alessia; Squecco, Roberta; Battistini, Chiara; Zecchi-Orlandini, Sandra; Francini, Fabio; Meacci, Elisabetta

    2011-11-01

    Skeletal muscle regeneration is severely compromised in the case of extended damage. The current challenge is to find factors capable of limiting muscle degeneration and/or potentiating the inherent regenerative program mediated by a specific type of myoblastic cells, the satellite cells. Recent studies from our groups and others have shown that the bioactive lipid, sphingosine 1-phosphate (S1P), promotes myoblast differentiation and exerts a trophic action on denervated skeletal muscle fibres. In the present study, we examined the effects of S1P on eccentric contraction (EC)-injured extensor digitorum longus muscle fibres and resident satellite cells. After EC, skeletal muscle showed evidence of structural and biochemical damage along with significant electrophysiological changes, i.e. reduced plasma membrane resistance and resting membrane potential and altered Na(+) and Ca(2+) current amplitude and kinetics. Treatment with exogenous S1P attenuated the EC-induced tissue damage, protecting skeletal muscle fibre from apoptosis, preserving satellite cell viability and affecting extracellular matrix remodelling, through the up-regulation of matrix metalloproteinase 9 (MMP-9) expression. S1P also promoted satellite cell renewal and differentiation in the damaged muscle. Notably, EC was associated with the activation of sphingosine kinase 1 (SphK1) and with increased endogenous S1P synthesis, further stressing the relevance of S1P in skeletal muscle protection and repair/regeneration. In line with this, the treatment with a selective SphK1 inhibitor during EC, caused an exacerbation of the muscle damage and attenuated MMP-9 expression. Together, these findings are in favour for a role of S1P in skeletal muscle healing and offer new clues for the identification of novel therapeutic approaches to counteract skeletal muscle damage and disease.

  1. The major facilitator superfamily transporter Knq1p modulates boron homeostasis in Kluyveromyces lactis.

    PubMed

    Svrbicka, Alexandra; Toth Hervay, Nora; Gbelska, Yvetta

    2016-03-01

    Boron is an essential micronutrient for living cells, yet its excess causes toxicity. To date, the mechanisms of boron toxicity are poorly understood. Recently, the ScATR1 gene has been identified encoding the main boron efflux pump in Saccharomyces cerevisiae. In this study, we analyzed the ScATR1 ortholog in Kluyveromyces lactis--the KNQ1 gene, to understand whether it participates in boron stress tolerance. We found that the KNQ1 gene, encoding a permease belonging to the major facilitator superfamily, is required for K. lactis boron tolerance. Deletion of the KNQ1 gene led to boron sensitivity and its overexpression increased K. lactis boron tolerance. The KNQ1 expression was induced by boron and the intracellular boron concentration was controlled by Knq1p. The KNQ1 promoter contains two putative binding motifs for the AP-1-like transcription factor KlYap1p playing a central role in oxidative stress defense. Our results indicate that the induction of the KNQ1 expression requires the presence of KlYap1p and that Knq1p like its ortholog ScAtr1p in S. cerevisiae functions as a boron efflux pump providing boron resistance in K. lactis.

  2. RAC1P29S is a spontaneously activating cancer-associated GTPase

    PubMed Central

    Davis, Matthew J.; Ha, Byung Hak; Holman, Edna C.; Halaban, Ruth; Schlessinger, Joseph; Boggon, Titus J.

    2013-01-01

    RAC1 is a small, Ras-related GTPase that was recently reported to harbor a recurrent UV-induced signature mutation in melanoma, resulting in substitution of P29 to serine (RAC1P29S), ranking this the third most frequently occurring gain-of-function mutation in melanoma. Although the Ras family GTPases are mutated in about 30% of all cancers, mutations in the Rho family GTPases have rarely been observed. In this study, we demonstrate that unlike oncogenic Ras proteins, which are primarily activated by mutations that eliminate GTPase activity, the activated melanoma RAC1P29S protein maintains intrinsic GTP hydrolysis and is spontaneously activated by substantially increased inherent GDP/GTP nucleotide exchange. Determination and comparison of crystal structures for activated RAC1 GTPases suggest that RAC1F28L—a known spontaneously activated RAC1 mutant—and RAC1P29S are self-activated in distinct fashions. Moreover, the mechanism of RAC1P29S and RAC1F28L activation differs from the common oncogenic mutations found in Ras-like GTPases that abrogate GTP hydrolysis. The melanoma RAC1P29S gain-of-function point mutation therefore represents a previously undescribed class of cancer-related GTPase activity. PMID:23284172

  3. A prion of yeast metacaspase homolog (Mca1p) detected by a genetic screen.

    PubMed

    Nemecek, Julie; Nakayashiki, Toru; Wickner, Reed B

    2009-02-10

    Saccharomyces cerevisiae can be infected with four amyloid-based prions: [URE3], [PSI(+)], [PIN(+)], and [SWI(+)], due to self-propagating aggregation of Ure2p, Sup35p, Rnq1p and Swi1p, respectively. We searched for new prions of yeast by fusing random segments of yeast DNA to SUP35MC, encoding the Sup35 protein lacking its own prion domain, selecting clones in which Sup35MC function was impaired. Three different clones contained parts of the Q/N-rich amino-terminal domain of Mca1p/Yca1p with the Sup35 part of the fusion protein partially inactive. This inactivity was dominant, segregated 4:0 in meiosis, and was efficiently transferred by cytoplasmic mixing. The inactivity was cured by overexpression of Hsp104, but the prion could arise again in the cured strain (reversible curing). Overproduction of the Mca1 N-terminal domain induced the de novo appearance of the prion form of the fusion. The prion state, which we name [MCA], was transmitted to the chromosomally encoded Mca1p based on genetic, cytological and biochemical tests.

  4. A Mouse Cytoplasmic Exoribonuclease (mXRN1p) with Preference for G4 Tetraplex Substrates

    PubMed Central

    Bashkirov, Vladimir I.; Scherthan, Harry; Solinger, Jachen A.; Buerstedde, Jean-Marie; Heyer, Wolf-Dietrich

    1997-01-01

    Exoribonucleases are important enzymes for the turnover of cellular RNA species. We have isolated the first mammalian cDNA from mouse demonstrated to encode a 5′–3′ exoribonuclease. The structural conservation of the predicted protein and complementation data in Saccharomyces cerevisiae suggest a role in cytoplasmic mRNA turnover and pre-rRNA processing similar to that of the major cytoplasmic exoribonuclease Xrn1p in yeast. Therefore, a key component of the mRNA decay system in S. cerevisiae has been conserved in evolution from yeasts to mammals. The purified mouse protein (mXRN1p) exhibited a novel substrate preference for G4 RNA tetraplex–containing substrates demonstrated in binding and hydrolysis experiments. mXRN1p is the first RNA turnover function that has been localized in the cytoplasm of mammalian cells. mXRN1p was distributed in small granules and was highly enriched in discrete, prominent foci. The specificity of mXRN1p suggests that RNAs containing G4 tetraplex structures may occur in vivo and may have a role in RNA turnover. PMID:9049243

  5. Roles of sphingosine-1-phosphate (S1P) receptors in malignant behavior of glioma cells. Differential effects of S1P{sub 2} on cell migration and invasiveness

    SciTech Connect

    Young, Nicholas; Van Brocklyn, James R. . E-mail: james.vanbrocklyn@osumc.edu

    2007-05-01

    Sphingosine-1-phosphate (S1P) is a bioactive lipid that signals through a family of five G-protein-coupled receptors, termed S1P{sub 1-5}. S1P stimulates growth and invasiveness of glioma cells, and high expression levels of the enzyme that forms S1P, sphingosine kinase-1, correlate with short survival of glioma patients. In this study we examined the mechanism of S1P stimulation of glioma cell proliferation and invasion by either overexpressing or knocking down, by RNA interference, S1P receptor expression in glioma cell lines. S1P{sub 1}, S1P{sub 2} and S1P{sub 3} all contribute positively to S1P-stimulated glioma cell proliferation, with S1P{sub 1} being the major contributor. Stimulation of glioma cell proliferation by these receptors correlated with activation of ERK MAP kinase. S1P{sub 5} blocks glioma cell proliferation, and inhibits ERK activation. S1P{sub 1} and S1P{sub 3} enhance glioma cell migration and invasion. S1P{sub 2} inhibits migration through Rho activation, Rho kinase signaling and stress fiber formation, but unexpectedly, enhances glioma cell invasiveness by stimulating cell adhesion. S1P{sub 2} also potently enhances expression of the matricellular protein CCN1/Cyr61, which has been implicated in tumor cell adhesion, and invasion as well as tumor angiogenesis. A neutralizing antibody to CCN1 blocked S1P{sub 2}-stimulated glioma invasion. Thus, while S1P{sub 2} decreases glioma cell motility, it may enhance invasion through induction of proteins that modulate glioma cell interaction with the extracellular matrix.

  6. Lag1p and Lac1p Are Essential for the Acyl-CoA–dependent Ceramide Synthase Reaction in Saccharomyces cerevisae

    PubMed Central

    Schorling, Stefan; Vallée, Béatrice; Barz, Wolfgang P.; Riezman, Howard; Oesterhelt, Dieter

    2001-01-01

    Lag1p and Lac1p are two homologous transmembrane proteins of the endoplasmic reticulum in Saccharomyces cerevisiae. Homologous genes have been found in a wide variety of eukaryotes. In yeast, both genes, LAC1 and LAG1, are required for efficient endoplasmic reticulum-to-Golgi transport of glycosylphosphatidylinositol-anchored proteins. In this study, we show that lag1Δlac1Δ cells have reduced sphingolipid levels due to a block of the fumonisin B1-sensitive and acyl-CoA–dependent ceramide synthase reaction. The sphingolipid synthesis defect in lag1Δlac1Δ cells can be partially corrected by overexpression of YPC1 or YDC1, encoding ceramidases that have been reported to have acyl-CoA–independent ceramide synthesis activity. Quadruple mutant cells (lag1Δlac1Δypc1Δydc1Δ) do not make any sphingolipids, but are still viable probably because they produce novel lipids. Moreover, lag1Δlac1Δ cells are resistant to aureobasidin A, an inhibitor of the inositolphosphorylceramide synthase, suggesting that aureobasidin A may be toxic because it leads to increased ceramide levels. Based on these data, LAG1 and LAC1 are the first genes to be identified that are required for the fumonisin B1-sensitive and acyl-CoA–dependent ceramide synthase reaction. PMID:11694577

  7. Measurement of the χ b (3 P) mass and of the relative rate of χ b1(1 P) and χ b2(1 P) production

    NASA Astrophysics Data System (ADS)

    Aaij, R.; Adeva, B.; Adinolfi, M.; Affolder, A.; Ajaltouni, Z.; Akar, S.; Albrecht, J.; Alessio, F.; Alexander, M.; Ali, S.; Alkhazov, G.; Alvarez Cartelle, P.; Alves, A. A.; Amato, S.; Amerio, S.; Amhis, Y.; An, L.; Anderlini, L.; Anderson, J.; Andreassen, R.; Andreotti, M.; Andrews, J. E.; Appleby, R. B.; Aquines Gutierrez, O.; Archilli, F.; Artamonov, A.; Artuso, M.; Aslanides, E.; Auriemma, G.; Baalouch, M.; Bachmann, S.; Back, J. J.; Badalov, A.; Baesso, C.; Baldini, W.; Barlow, R. J.; Barschel, C.; Barsuk, S.; Barter, W.; Batozskaya, V.; Battista, V.; Bay, A.; Beaucourt, L.; Beddow, J.; Bedeschi, F.; Bediaga, I.; Belogurov, S.; Belous, K.; Belyaev, I.; Ben-Haim, E.; Bencivenni, G.; Benson, S.; Benton, J.; Berezhnoy, A.; Bernet, R.; Bettler, M.-O.; van Beuzekom, M.; Bien, A.; Bifani, S.; Bird, T.; Bizzeti, A.; Bjørnstad, P. M.; Blake, T.; Blanc, F.; Blouw, J.; Blusk, S.; Bocci, V.; Bondar, A.; Bondar, N.; Bonivento, W.; Borghi, S.; Borgia, A.; Borsato, M.; Bowcock, T. J. V.; Bowen, E.; Bozzi, C.; Brambach, T.; van den Brand, J.; Bressieux, J.; Brett, D.; Britsch, M.; Britton, T.; Brodzicka, J.; Brook, N. H.; Brown, H.; Bursche, A.; Busetto, G.; Buytaert, J.; Cadeddu, S.; Calabrese, R.; Calvi, M.; Calvo Gomez, M.; Campana, P.; Campora Perez, D.; Carbone, A.; Carboni, G.; Cardinale, R.; Cardini, A.; Carson, L.; Carvalho Akiba, K.; Casse, G.; Cassina, L.; Castillo Garcia, L.; Cattaneo, M.; Cauet, Ch.; Cenci, R.; Charles, M.; Charpentier, Ph.; Chefdeville, M.; Chen, S.; Cheung, S.-F.; Chiapolini, N.; Chrzaszcz, M.; Ciba, K.; Cid Vidal, X.; Ciezarek, G.; Clarke, P. E. L.; Clemencic, M.; Cliff, H. V.; Closier, J.; Coco, V.; Cogan, J.; Cogneras, E.; Cojocariu, L.; Collins, P.; Comerma-Montells, A.; Contu, A.; Cook, A.; Coombes, M.; Coquereau, S.; Corti, G.; Corvo, M.; Counts, I.; Couturier, B.; Cowan, G. A.; Craik, D. C.; Cruz Torres, M.; Cunliffe, S.; Currie, R.; D'Ambrosio, C.; Dalseno, J.; David, P.; David, P. N. Y.; Davis, A.; De Bruyn, K.; De Capua, S.; De Cian, M.; De Miranda, J. M.; De Paula, L.; De Silva, W.; De Simone, P.; Decamp, D.; Deckenhoff, M.; Del Buono, L.; Déléage, N.; Derkach, D.; Deschamps, O.; Dettori, F.; Di Canto, A.; Dijkstra, H.; Donleavy, S.; Dordei, F.; Dorigo, M.; Dosil Suárez, A.; Dossett, D.; Dovbnya, A.; Dreimanis, K.; Dujany, G.; Dupertuis, F.; Durante, P.; Dzhelyadin, R.; Dziurda, A.; Dzyuba, A.; Easo, S.; Egede, U.; Egorychev, V.; Eidelman, S.; Eisenhardt, S.; Eitschberger, U.; Ekelhof, R.; Eklund, L.; El Rifai, I.; Elsasser, Ch.; Ely, S.; Esen, S.; Evans, H.-M.; Evans, T.; Falabella, A.; Färber, C.; Farinelli, C.; Farley, N.; Farry, S.; Fay, R. F.; Ferguson, D.; Fernandez Albor, V.; Ferreira Rodrigues, F.; Ferro-Luzzi, M.; Filippov, S.; Fiore, M.; Fiorini, M.; Firlej, M.; Fitzpatrick, C.; Fiutowski, T.; Fontana, M.; Fontanelli, F.; Forty, R.; Francisco, O.; Frank, M.; Frei, C.; Frosini, M.; Fu, J.; Furfaro, E.; Gallas Torreira, A.; Galli, D.; Gallorini, S.; Gambetta, S.; Gandelman, M.; Gandini, P.; Gao, Y.; García Pardiñas, J.; Garofoli, J.; Garra Tico, J.; Garrido, L.; Gaspar, C.; Gauld, R.; Gavardi, L.; Gavrilov, G.; Geraci, A.; Gersabeck, E.; Gersabeck, M.; Gershon, T.; Ghez, Ph.; Gianelle, A.; Gianì, S.; Gibson, V.; Giubega, L.; Gligorov, V. V.; Göbel, C.; Golubkov, D.; Golutvin, A.; Gomes, A.; Gotti, C.; Grabalosa Gándara, M.; Graciani Diaz, R.; Granado Cardoso, L. A.; Graugés, E.; Graziani, G.; Grecu, A.; Greening, E.; Gregson, S.; Griffith, P.; Grillo, L.; Grünberg, O.; Gui, B.; Gushchin, E.; Guz, Yu.; Gys, T.; Hadjivasiliou, C.; Haefeli, G.; Haen, C.; Haines, S. C.; Hall, S.; Hamilton, B.; Hampson, T.; Han, X.; Hansmann-Menzemer, S.; Harnew, N.; Harnew, S. T.; Harrison, J.; He, J.; Head, T.; Heijne, V.; Hennessy, K.; Henrard, P.; Henry, L.; Hernando Morata, J. A.; van Herwijnen, E.; Heß, M.; Hicheur, A.; Hill, D.; Hoballah, M.; Hombach, C.; Hulsbergen, W.; Hunt, P.; Hussain, N.; Hutchcroft, D.; Hynds, D.; Idzik, M.; Ilten, P.; Jacobsson, R.; Jaeger, A.; Jalocha, J.; Jans, E.; Jaton, P.; Jawahery, A.; Jing, F.; John, M.; Johnson, D.; Jones, C. R.; Joram, C.; Jost, B.; Jurik, N.; Kandybei, S.; Kanso, W.; Karacson, M.; Karbach, T. M.; Karodia, S.; Kelsey, M.; Kenyon, I. R.; Ketel, T.; Khanji, B.; Khurewathanakul, C.; Klaver, S.; Klimaszewski, K.; Kochebina, O.; Kolpin, M.; Komarov, I.; Koopman, R. F.; Koppenburg, P.; Korolev, M.; Kozlinskiy, A.; Kravchuk, L.; Kreplin, K.; Kreps, M.; Krocker, G.; Krokovny, P.; Kruse, F.; Kucewicz, W.; Kucharczyk, M.; Kudryavtsev, V.; Kurek, K.; Kvaratskheliya, T.; La Thi, V. N.; Lacarrere, D.; Lafferty, G.; Lai, A.; Lambert, D.; Lambert, R. W.; Lanfranchi, G.; Langenbruch, C.; Langhans, B.; Latham, T.; Lazzeroni, C.; Le Gac, R.; van Leerdam, J.; Lees, J.-P.; Lefèvre, R.; Leflat, A.; Lefrançois, J.; Leo, S.; Leroy, O.; Lesiak, T.; Lespinasse, M.; Leverington, B.; Li, Y.; Likhomanenko, T.; Liles, M.; Lindner, R.; Linn, C.; Lionetto, F.; Liu, B.; Lohn, S.; Longstaff, I.; Lopes, J. H.; Lopez-March, N.; Lowdon, P.; Lu, H.; Lucchesi, D.; Luo, H.; Lupato, A.; Luppi, E.; Lupton, O.; Machefert, F.; Machikhiliyan, I. V.; Maciuc, F.; Maev, O.; Malde, S.; Malinin, A.; Manca, G.; Mancinelli, G.; Mapelli, A.; Maratas, J.; Marchand, J. F.; Marconi, U.; Marin Benito, C.; Marino, P.; Märki, R.; Marks, J.; Martellotti, G.; Martens, A.; Martín Sánchez, A.; Martinelli, M.; Martinez Santos, D.; Martinez Vidal, F.; Martins Tostes, D.; Massafferri, A.; Matev, R.; Mathe, Z.; Matteuzzi, C.; Mazurov, A.; McCann, M.; McCarthy, J.; McNab, A.; McNulty, R.; McSkelly, B.; Meadows, B.; Meier, F.; Meissner, M.; Merk, M.; Milanes, D. A.; Minard, M.-N.; Moggi, N.; Molina Rodriguez, J.; Monteil, S.; Morandin, M.; Morawski, P.; Mordà, A.; Morello, M. J.; Moron, J.; Morris, A.-B.; Mountain, R.; Muheim, F.; Müller, K.; Mussini, M.; Muster, B.; Naik, P.; Nakada, T.; Nandakumar, R.; Nasteva, I.; Needham, M.; Neri, N.; Neubert, S.; Neufeld, N.; Neuner, M.; Nguyen, A. D.; Nguyen, T. D.; Nguyen-Mau, C.; Nicol, M.; Niess, V.; Niet, R.; Nikitin, N.; Nikodem, T.; Novoselov, A.; O'Hanlon, D. P.; Oblakowska-Mucha, A.; Obraztsov, V.; Oggero, S.; Ogilvy, S.; Okhrimenko, O.; Oldeman, R.; Onderwater, C. J. G.; Orlandea, M.; Otalora Goicochea, J. M.; Owen, P.; Oyanguren, A.; Pal, B. K.; Palano, A.; Palombo, F.; Palutan, M.; Panman, J.; Papanestis, A.; Pappagallo, M.; Pappalardo, L. L.; Parkes, C.; Parkinson, C. J.; Passaleva, G.; Patel, G. D.; Patel, M.; Patrignani, C.; Pearce, A.; Pellegrino, A.; Pepe Altarelli, M.; Perazzini, S.; Perret, P.; Perrin-Terrin, M.; Pescatore, L.; Pesen, E.; Petridis, K.; Petrolini, A.; Picatoste Olloqui, E.; Pietrzyk, B.; Pilař, T.; Pinci, D.; Pistone, A.; Playfer, S.; Plo Casasus, M.; Polci, F.; Poluektov, A.; Polycarpo, E.; Popov, A.; Popov, D.; Popovici, B.; Potterat, C.; Price, E.; Prisciandaro, J.; Pritchard, A.; Prouve, C.; Pugatch, V.; Puig Navarro, A.; Punzi, G.; Qian, W.; Rachwal, B.; Rademacker, J. H.; Rakotomiaramanana, B.; Rama, M.; Rangel, M. S.; Raniuk, I.; Rauschmayr, N.; Raven, G.; Reichert, S.; Reid, M. M.; dos Reis, A. C.; Ricciardi, S.; Richards, S.; Rihl, M.; Rinnert, K.; Rives Molina, V.; Roa Romero, D. A.; Robbe, P.; Rodrigues, A. B.; Rodrigues, E.; Rodriguez Perez, P.; Roiser, S.; Romanovsky, V.; Romero Vidal, A.; Rotondo, M.; Rouvinet, J.; Ruf, T.; Ruiz, H.; Ruiz Valls, P.; Saborido Silva, J. J.; Sagidova, N.; Sail, P.; Saitta, B.; Salustino Guimaraes, V.; Sanchez Mayordomo, C.; Sanmartin Sedes, B.; Santacesaria, R.; Santamarina Rios, C.; Santovetti, E.; Sarti, A.; Satriano, C.; Satta, A.; Saunders, D. M.; Savrina, D.; Schiller, M.; Schindler, H.; Schlupp, M.; Schmelling, M.; Schmidt, B.; Schneider, O.; Schopper, A.; Schune, M.-H.; Schwemmer, R.; Sciascia, B.; Sciubba, A.; Semennikov, A.; Sepp, I.; Serra, N.; Serrano, J.; Sestini, L.; Seyfert, P.; Shapkin, M.; Shapoval, I.; Shcheglov, Y.; Shears, T.; Shekhtman, L.; Shevchenko, V.; Shires, A.; Silva Coutinho, R.; Simi, G.; Sirendi, M.; Skidmore, N.; Skwarnicki, T.; Smith, N. A.; Smith, E.; Smith, E.; Smith, J.; Smith, M.; Snoek, H.; Sokoloff, M. D.; Soler, F. J. P.; Soomro, F.; Souza, D.; Souza De Paula, B.; Spaan, B.; Sparkes, A.; Spradlin, P.; Sridharan, S.; Stagni, F.; Stahl, M.; Stahl, S.; Steinkamp, O.; Stenyakin, O.; Stevenson, S.; Stoica, S.; Stone, S.; Storaci, B.; Stracka, S.; Straticiuc, M.; Straumann, U.; Stroili, R.; Subbiah, V. K.; Sun, L.; Sutcliffe, W.; Swientek, K.; Swientek, S.; Syropoulos, V.; Szczekowski, M.; Szczypka, P.; Szumlak, T.; T'Jampens, S.; Teklishyn, M.; Tellarini, G.; Teubert, F.; Thomas, C.; Thomas, E.; van Tilburg, J.; Tisserand, V.; Tobin, M.; Tolk, S.; Tomassetti, L.; Tonelli, D.; Topp-Joergensen, S.; Torr, N.; Tournefier, E.; Tourneur, S.; Tran, M. T.; Tresch, M.; Trisovic, A.; Tsaregorodtsev, A.; Tsopelas, P.; Tuning, N.; Ubeda Garcia, M.; Ukleja, A.; Ustyuzhanin, A.; Uwer, U.; Vagnoni, V.; Valenti, G.; Vallier, A.; Vazquez Gomez, R.; Vazquez Regueiro, P.; Vázquez Sierra, C.; Vecchi, S.; Velthuis, J. J.; Veltri, M.; Veneziano, G.; Vesterinen, M.; Viaud, B.; Vieira, D.; Vieites Diaz, M.; Vilasis-Cardona, X.; Vollhardt, A.; Volyanskyy, D.; Voong, D.; Vorobyev, A.; Vorobyev, V.; Voß, C.; de Vries, J. A.; Waldi, R.; Wallace, C.; Wallace, R.; Walsh, J.; Wandernoth, S.; Wang, J.; Ward, D. R.; Watson, N. K.; Websdale, D.; Whitehead, M.; Wicht, J.; Wiedner, D.; Wilkinson, G.; Williams, M. P.; Williams, M.; Wilson, F. F.; Wimberley, J.; Wishahi, J.; Wislicki, W.; Witek, M.; Wormser, G.; Wotton, S. A.; Wright, S.; Wu, S.; Wyllie, K.; Xie, Y.; Xing, Z.; Xu, Z.; Yang, Z.; Yuan, X.; Yushchenko, O.; Zangoli, M.; Zavertyaev, M.; Zhang, L.; Zhang, W. C.; Zhang, Y.; Zhelezov, A.; Zhokhov, A.; Zhong, L.; Zvyagin, A.

    2014-10-01

    The production of χ b mesons in proton-proton collisions is studied using a data sample collected by the LHCb detector, at centre-of-mass energies of =7 and 8 TeV and corresponding to an integrated luminosity of 3.0 fb-1. The χ b mesons are identified through their decays to ϒ(1 S) γ and ϒ(2 S) γ using photons that converted to e + e - pairs in the detector. The relative prompt production rate of χ b1(1 P) and χ b2(1 P) mesons is measured as a function of the ϒ(1 S) transverse momentum in the χ b rapidity range 2.0 < y <4.5. A precise measurement of the χ b (3 P) mass is also performed. Assuming a mass splitting between the χ b1(3 P) and the χ b2(3 P) states of 10.5 MeV/c2, the measured mass of the χ b1(3 P) meson is

  8. Polymicrogyria and infantile spasms in a patient with 1p36 deletion syndrome.

    PubMed

    Saito, Yoshiaki; Kubota, Masaya; Kurosawa, Kenji; Ichihashi, Izumi; Kaneko, Yuu; Hattori, Ayako; Komaki, Hirofumi; Nakagawa, Eiji; Sugai, Kenji; Sasaki, Masayuki

    2011-05-01

    A 3-months-old boy presented with partial seizures that soon evolved into infantile spasms. Magnetic resonance imaging revealed bilateral perisylvian polymicrogyria with right-sided predominance. ACTH therapy successfully controlled epilepsy and electroencephalograms were normalized. Conventional G-banded chromosomal analysis was performed due to his distinctive features and a derivative chromosome 1 derived from parental balanced translocation with a karyoptype of 46,XY,der(1)t(1;4)(p36.23;q35) was detected. Fluorescent in situ hybridization analysis confirmed the deleted region of 1p36 as large as 8.6Mb. This is the first delineation of concurrent complications of infantile spasms and polymicrogyria in patient with 1p36 deletion. 1p36 deletion syndrome should be broadly recognized as a differential diagnosis of regional polymicrogyria and/or infantile spasms.

  9. Morbid obesity in a child with monosomy 1p36 syndrome.

    PubMed

    Zagalo, Ana; Dias, Patricia; Pereira, Carla; Sampaio, Maria de Lurdes

    2012-03-20

    The monosomy 1p36 syndrome is a cause of syndromic obesity. It is characterised by psychomotor delay, hypotonia and typical craniofacial dysmorphism. Other features commonly associated are behavioural anomalies including hyperphagia and self-injuring, seizures, congenital heart disease and hypothyroidism. The authors report the case of a 9-year and 5-month-boy referred to the paediatric endocrinology clinics for morbid obesity. Clinical findings were generalised obesity with a body mass index >95th centile, acanthosis nigricans of the neck, arms with self inflicted lesions, deep-set eyes, straight eyebrows, broad nasal bridge and pointed chin. He was unable to walk and had no expressive language. Cytogenetic analysis identified 1p36.33-pter deletion (~139 Mb terminal deletion in chromosome 1 short arm) and Y chromosome duplication. The blood analysis showed insulin resistance and dyslipidaemia. The authors emphasise the need to consider monosomy 1p36 as a cause of severe psychomotor delay and obesity.

  10. Morbid obesity in a child with monosomy 1p36 syndrome

    PubMed Central

    Zagalo, Ana; Dias, Patricia; Pereira, Carla; Sampaio, Maria de Lurdes

    2012-01-01

    The monosomy 1p36 syndrome is a cause of syndromic obesity. It is characterised by psychomotor delay, hypotonia and typical craniofacial dysmorphism. Other features commonly associated are behavioural anomalies including hyperphagia and self-injuring, seizures, congenital heart disease and hypothyroidism. The authors report the case of a 9-year and 5-month-boy referred to the paediatric endocrinology clinics for morbid obesity. Clinical findings were generalised obesity with a body mass index >95th centile, acanthosis nigricans of the neck, arms with self inflicted lesions, deep-set eyes, straight eyebrows, broad nasal bridge and pointed chin. He was unable to walk and had no expressive language. Cytogenetic analysis identified 1p36.33-pter deletion (~139 Mb terminal deletion in chromosome 1 short arm) and Y chromosome duplication. The blood analysis showed insulin resistance and dyslipidaemia. The authors emphasise the need to consider monosomy 1p36 as a cause of severe psychomotor delay and obesity. PMID:22605691

  11. Hunt for the 1/sup 1/P/sub 1/ bound state of charmonium

    SciTech Connect

    Porter, F.C.

    1982-02-01

    Using the Crystal Ball detector at SPEAR, we have looked for evidence of the isospin-violating decay psi' ..-->.. ..pi../sup 01/P/sub 1/, where /sup 1/P/sub 1/ is the predicted spin-singlet p-wave bound state of charmonium. For a /sup 1/P/sub 1/ state at the predicted mass (approx. 3520 MeV), we obtain the 95% confidence level limits: BR(psi' ..-->.. ..pi../sup 01/P/sub 1/) < 0.55%, BR(psi' ..-->.. ..pi../sup 01/P/sub 1/)BR(/sup 1/P/sub 1/ ..-->.. ..gamma..n/sub c/ < 0.14%. These limits are compared with simple theoretical predictions.

  12. Electron-impact excitation of the n 1P levels of helium - Theory and experiment

    NASA Technical Reports Server (NTRS)

    Cartwright, David C.; Csanak, George; Trajmar, Sandor; Register, D. F.

    1992-01-01

    New experimental electron-energy-loss data have been used to extract differential and integral cross sections for excitation of the 2 1P level, and for the overlapping (3 1P, 3 1D, 3 3D) levels of helium, at 30-, 50-, and 100-eV incident electron energies. First-order many-body theory (FOMBT) has been used to calculate the differential and integral cross sections for excitation of the n 1P (n = 2,...,6) levels of helium by electron impact, for incident electron energies from threshold to 500 eV. Detailed comparisons between these two new sets of data are made as well as comparisons with appropriate published experimental and theoretical results. A simple scaling relationship is derived from the FOMBT results for n = 2,...,6 that provides differential and integral cross sections for all symmetry final levels of helium with n = 6 or greater.

  13. Combined PDF and Rietveld studies of ADORable zeolites and the disordered intermediate IPC-1P.

    PubMed

    Morris, Samuel A; Wheatley, Paul S; Položij, Miroslav; Nachtigall, Petr; Eliášová, Pavla; Čejka, Jiří; Lucas, Tim C; Hriljac, Joseph A; Pinar, Ana B; Morris, Russell E

    2016-09-28

    The disordered intermediate of the ADORable zeolite UTL has been structurally confirmed using the pair distribution function (PDF) technique. The intermediate, IPC-1P, is a disordered layered compound formed by the hydrolysis of UTL in 0.1 M hydrochloric acid solution. Its structure is unsolvable by traditional X-ray diffraction techniques. The PDF technique was first benchmarked against high-quality synchrotron Rietveld refinements of IPC-2 (OKO) and IPC-4 (PCR) - two end products of IPC-1P condensation that share very similar structural features. An IPC-1P starting model derived from density functional theory was used for the PDF refinement, which yielded a final fit of Rw = 18% and a geometrically reasonable structure. This confirms the layers do stay intact throughout the ADOR process and shows PDF is a viable technique for layered zeolite structure determination.

  14. Dynamical Relativistic Effects in Quasielastic 1p -Shell Proton Knockout from O{sup 16}

    SciTech Connect

    Gao, J.; Anderson, B. D.; Aniol, K. A.; Auerbach, L.; Baker, F. T.; Berthot, J.; Bertin, P.-Y.; Boeglin, W. U.

    2000-04-10

    We have measured the cross section for quasielastic 1p -shell proton knockout in the {sup 16}O( e, e{sup '}p) reaction at {omega}=0.439 GeV and Q{sup 2}=0.8 (GeV/c){sup 2} for missing momentum P{sub miss}{<=}355 MeV /c . We have extracted the response functions R{sub L+TT} , R{sub T} , R{sub LT} , and the left-right asymmetry, A{sub LT} , for the 1p{sub 1/2} and the 1p{sub 3/2} states. The data are well described by relativistic distorted wave impulse approximation calculations. At large P{sub miss} , the structure observed in A{sub LT} indicates the existence of dynamical relativistic effects. (c) 2000 The American Physical Society.

  15. Left-ventricular non-compaction in a patient with monosomy 1p36.

    PubMed

    Thienpont, Bernard; Mertens, Luc; Buyse, Gunnar; Vermeesch, Joris R; Devriendt, Koen

    2007-01-01

    We report on a new-born girl with left ventricular non-compaction (LVNC), dysmorphism and epilepsy. Array-CGH at 1 Mb resolution revealed a deletion of the terminal 4.6 to 5.9 Mb of the short arm of chromosome 1. Cardiac abnormalities such as dilated cardiomyopathy and structural cardiac defects are common findings in patients with monosomy 1p36. This is however the first report describing LVNC in association with the 1p36 deletion syndrome, broadening the spectrum of cardiac anomalies found in association with this syndrome.

  16. Structural insights into how Yrb2p accelerates the assembly of the Xpo1p nuclear export complex.

    PubMed

    Koyama, Masako; Shirai, Natsuki; Matsuura, Yoshiyuki

    2014-11-06

    Proteins and ribonucleoproteins containing a nuclear export signal (NES) assemble with the exportin Xpo1p (yeast CRM1) and Gsp1p-GTP (yeast Ran-GTP) in the nucleus and exit through the nuclear pore complex. In the cytoplasm, Yrb1p (yeast RanBP1) displaces NES from Xpo1p. Efficient export of NES-cargoes requires Yrb2p (yeast RanBP3), a primarily nuclear protein containing nucleoporin-like phenylalanine-glycine (FG) repeats and a low-affinity Gsp1p-binding domain (RanBD). Here, we show that Yrb2p strikingly accelerates the association of Gsp1p-GTP and NES to Xpo1p. We have solved the crystal structure of the Xpo1p-Yrb2p-Gsp1p-GTP complex, a key assembly intermediate that can bind cargo rapidly. Although the NES-binding cleft of Xpo1p is closed in this intermediate, our data suggest that preloading of Gsp1p-GTP onto Xpo1p by Yrb2p, conformational flexibility of Xpo1p, and the low affinity of RanBD enable active displacement of Yrb2p RanBD by NES to occur effectively. The structure also reveals the major binding sites for FG repeats on Xpo1p.

  17. Sphingosine-1-phosphate promotes lymphangiogenesis by stimulating S1P1/Gi/PLC/Ca2+ signaling pathways.

    PubMed

    Yoon, Chang Min; Hong, Bok Sil; Moon, Hyung Geun; Lim, Seyoung; Suh, Pann-Ghill; Kim, Yoon-Keun; Chae, Chi-Bom; Gho, Yong Song

    2008-08-15

    The lymphatic system plays pivotal roles in mediating tissue fluid homeostasis and immunity, and excessive lymphatic vessel formation is implicated in many pathological conditions, which include inflammation and tumor metastasis. However, the molecular mechanisms that regulate lymphatic vessel formation remain poorly characterized. Sphingosine-1-phosphate (S1P) is a potent bioactive lipid that is implicated in a variety of biologic processes such as inflammatory responses and angiogenesis. Here, we first report that S1P acts as a lymphangiogenic mediator. S1P induced migration, capillary-like tube formation, and intracellular Ca(2+) mobilization, but not proliferation, in human lymphatic endothelial cells (HLECs) in vitro. Moreover, a Matrigel plug assay demonstrated that S1P promoted the outgrowth of new lymphatic vessels in vivo. HLECs expressed S1P1 and S1P3, and both RNA interference-mediated down-regulation of S1P1 and an S1P1 antagonist significantly blocked S1P-mediated lymphangiogenesis. Furthermore, pertussis toxin, U73122, and BAPTA-AM efficiently blocked S1P-induced in vitro lymphangiogenesis and intracellular Ca(2+) mobilization of HLECs, indicating that S1P promotes lymphangiogenesis by stimulating S1P1/G(i)/phospholipase C/Ca(2+) signaling pathways. Our results suggest that S1P is the first lymphangiogenic bioactive lipid to be identified, and that S1P and its receptors might serve as new therapeutic targets against inflammatory diseases and lymphatic metastasis in tumors.

  18. Identification of novel susceptibility loci for inflammatory bowel disease on chromosomes 1p, 3q, and 4q: Evidence for epistasis between 1p and IBD1

    PubMed Central

    Cho, Judy H.; Nicolae, Dan L.; Gold, Leslee H.; Fields, Carter T.; LaBuda, Michele C.; Rohal, Patrick M.; Pickles, Michael R.; Qin, Li; Fu, Yifan; Mann, Jasdeep S.; Kirschner, Barbara S.; Jabs, Ethylin Wang; Weber, James; Hanauer, Stephen B.; Bayless, Theodore M.; Brant, Steven R.

    1998-01-01

    The idiopathic inflammatory bowel diseases, Crohn’s disease (CD) and ulcerative colitis (UC), are chronic, frequently disabling diseases of the intestines. Segregation analyses, twin concordance, and ethnic differences in familial risks have established that CD and UC are complex, non-Mendelian, related genetic disorders. We performed a genome-wide screen using 377 autosomal markers, on 297 CD, UC, or mixed relative pairs from 174 families, 37% Ashkenazim. We observed evidence for linkage at 3q for all families (multipoint logarithm of the odds score (MLod) = 2.29, P = 5.7 × 10−4), with greatest significance for non-Ashkenazim Caucasians (MLod = 3.39, P = 3.92 × 10−5), and at chromosome 1p (MLod = 2.65, P = 2.4 × 10−4) for all families. In a limited subset of mixed families (containing one member with CD and another with UC), evidence for linkage was observed on chromosome 4q (MLod = 2.76, P = 1.9 × 10−4), especially among Ashkenazim. There was confirmatory evidence for a CD locus, overlapping IBD1, in the pericentromeric region of chromosome 16 (MLod = 1.69, P = 2.6 × 10−3), particularly among Ashkenazim (MLod = 1.51, P = 7.8 × 10−3); however, positive MLod scores were observed over a very broad region of chromosome 16. Furthermore, evidence for epistasis between IBD1 and chromosome 1p was observed. Thirteen additional loci demonstrated nominal (MLod > 1.0, P < 0.016) evidence for linkage. This screen provides strong evidence that there are several major susceptibility loci contributing to the genetic risk for CD and UC. PMID:9636179

  19. Identification of 1p36 deletion syndrome in patients with facial dysmorphism and developmental delay

    PubMed Central

    Seo, Go Hun; Kim, Ja Hye; Cho, Ja Hyang; Kim, Gu-Hwan; Seo, Eul-Ju; Lee, Beom Hee; Choi, Jin-Ho

    2016-01-01

    Purpose The 1p36 deletion syndrome is a microdeletion syndrome characterized by developmental delays/intellectual disability, craniofacial dysmorphism, and other congenital anomalies. To date, many cases of this syndrome have been reported worldwide. However, cases with this syndrome have not been reported in Korean populations anywhere. This study was performed to report the clinical and molecular characteristics of five Korean patients with the 1p36 deletion syndrome. Methods The clinical characteristics of the 5 patients were reviewed. Karyotyping and multiplex ligation-dependent probe amplification (MLPA) analyses were performed for genetic diagnoses. Results All 5 patients had typical dysmorphic features including frontal bossing, flat right parietal bone, low-set ears, straight eyebrows, down-slanting palpebral fissure, hypotelorism, flat nasal roots, midface hypoplasia, pointed chins, small lips, and variable degrees of developmental delay. Each patient had multiple and variable anomalies such as a congenital heart defect including ventricular septal defect, atrial septal defect, and patent duct arteriosus, ventriculomegaly, cryptorchism, or hearing loss. Karyotyping revealed the 1p36 deletion in only 1 patient, although it was confirmed in all 5 patients by MLPA analyses. Conclusion All the patients had the typical features of 1p36 deletion. These hallmarks can be used to identify other patients with this condition in their early years in order to provide more appropriate care. PMID:26893599

  20. SKI-1/S1P inhibitor PF-429242 impairs the onset of HCV infection.

    PubMed

    Blanchet, Matthieu; Sureau, Camille; Guévin, Carl; Seidah, Nabil G; Labonté, Patrick

    2015-03-01

    Worldwide, approximately 170 million individuals are afflicted with chronic hepatitis C virus (HCV) infection. To prevent the development of inherent diseases such as cirrhosis and hepatocellular carcinoma, tremendous efforts have been made, leading to the development of promising new treatments. However, their efficiency is still dependent on the viral genotype. Additionally, these treatments that target the virus directly can trigger the emergence of resistant variants. In a previous study, we have demonstrated that a long-term (72h) inhibition of SKI-1/S1P, a master lipogenic pathway regulator through activation of SREBP, resulted in impaired HCV genome replication and infectious virion secretion. In the present study, we sought to investigate the antiviral effect of the SKI-1/S1P small molecule inhibitor PF-429242 at the early steps of the HCV lifecycle. Our results indicate a very potent antiviral effect of the inhibitor early in the viral lifecycle and that the overall action of the compound relies on two different contributions. The first one is SREBP/SKI-1/S1P dependent and involves LDLR and NPC1L1 proteins, while the second one is SREBP independent. Overall, our study confirms that SKI-1/S1P is a relevant target to impair HCV infection and that PF-429242 could be a promising candidate in the field of HCV infection treatment.

  1. Delineating the phenotype of 1p36 deletion in adolescents and adults.

    PubMed

    Brazil, Ashley; Stanford, Kevin; Smolarek, Teresa; Hopkin, Robert

    2014-10-01

    1p36 deletion is the most common telomeric deletion syndrome, with an incidence of 1/5,000-1/10,000. A variety of clinical complications have been reported including seizures, hypotonia, heart malformations, cardiomyopathy, vision problems, and hearing loss. Approximately 90% are reported to have severe to profound intellectual disability and 75% to have absent expressive language. Little is known about long-term outcomes. The current literature suggests a poor prognosis for most patients. This study attempted to assess medical conditions and function of adolescent and adult patients with 1p36 deletion. A survey was distributed through three support groups to identify patients >12 years of age to assess functional status and medical problems in older patients with 1p36 deletion syndrome. 40 patients were identified between 12 and 46 years old. Among our survey sample, medical complications including seizures, hypotonia, structural heart defects, hearing loss, and vision problems, were similar to previous reports. However, functional skills were better than anticipated, with an overwhelming majority reported to independently sit, walk, and receive the majority of nutrition orally. Forty-four percent were reported to use complex speech abilities. While medical problems in patients with 1p36 deletion were similar to those that have been previously reported, we also demonstrated these same concerns persist into adolescence and adulthood. Additionally, patients were reported to have better functional skills than anticipated. Thus, quality of life and level of function appear to be better than anticipated from previous studies. © 2014 Wiley Periodicals, Inc.

  2. OEIS complex associated with chromosome 1p36 deletion: a case report and review.

    PubMed

    El-Hattab, Ayman W; Skorupski, Josh C; Hsieh, Michael H; Breman, Amy M; Patel, Ankita; Cheung, Sau Wai; Craigen, William J

    2010-02-01

    OEIS complex (Omphalocele, Exstrophy of the cloaca, Imperforate anus, and Spine abnormalities) is a rare defect with estimated incidence of 1 in 200,000 live births. Most cases are sporadic, with no obvious cause. However, it has been rarely reported in patients with family members having similar malformations or with chromosomal anomalies. In addition, OEIS complex has been observed in association with environmental exposures, twinning, and in vitro fertilization. Monosomy 1p36 is the most common terminal deletion syndrome, with a prevalence of 1 in 5,000 newborns. It is characterized by specific facial features, developmental delay, and heart, skeletal, genitourinary, and neurological defects. We describe an infant with OEIS complex and 1p36 deletion who had features of both disorders, including omphalocele, cloacal exstrophy, imperforate anus, sacral multiple segmentation, renal malposition and malrotation, genital anomalies, diastasis of the symphysis pubis, microbrachycephaly, large anterior fontanel, cardiac septal defects, rib fusion, a limb deformity, developmental delay, and typical facial features. Chromosomal microarray analysis detected a 2.4 Mb terminal deletion of chromosome 1p. This is the first reported case with OEIS complex in association with a chromosome 1p36 deletion.

  3. Prenatal diagnosis and prenatal imaging features of fetal monosomy 1p36.

    PubMed

    Lissauer, D; Larkins, S A; Sharif, S; MacPherson, L; Rhodes, C; Kilby, M D

    2007-09-01

    Deletion of the distal end of the short arm of chromosome 1 (1p36) is thought to be a common terminal chromosomal deletion. However, few cases prospectively diagnosed prenatally have been reported. In this case, prenatal ultrasound at 21 weeks of gestation noted the fetus to have mild ventriculomegaly (Vhanterior = 11 mm and Vhposterior = 12 mm) and increased nuchal edema (6 mm). Maternal serum alpha-fetoprotein was normal unlike in a majority of previously described cases. The prenatal ultrasound features were further clarified with fetal MRI. Chromosome analysis following amniocentesis demonstrated a 1p36 deletion, which was confirmed by fluorescence in situ hybridization (FISH). The syndrome associated with 1p36 deletion is well described in infants and is characterized by typical facial features (prominent forehead, straight eyebrows. deep-set eyes, flat nasal bridge and a pointed chin). Other associated features are neurodevelopmental delay, seizures, cardiomyopathy and neurosensory hearing impairment. This case supplements our knowledge of the prenatal features of 1p36. Identification of this deletion by direct chromosomal analysis can be technically difficult and vigilance is required to improve diagnosis. FISH analysis is an important diagnostic adjunct where the diagnosis is suspected following classical G-banding techniques. However, in this chromosomal anomaly there remain few characteristic prenatal signs that are readily diagnosed with prenatal imaging.

  4. Glutathione and Gts1p drive beneficial variability in the cadmium resistances of individual yeast cells

    PubMed Central

    Smith, Matthew C A; Sumner, Edward R; Avery, Simon V

    2007-01-01

    Phenotypic heterogeneity among individual cells within isogenic populations is widely documented, but its consequences are not well understood. Here, cell-to-cell variation in the stress resistance of Saccharomyces cerevisiae, particularly to cadmium, was revealed to depend on the antioxidant glutathione. Heterogeneity was decreased strikingly in gsh1 mutants. Furthermore, cells sorted according to differing reduced-glutathione (GSH) contents exhibited differing stress resistances. The vacuolar GSH-conjugate pathway of detoxification was implicated in heterogeneous Cd resistance. Metabolic oscillations (ultradian rhythms) in yeast are known to modulate single-cell redox and GSH status. Gts1p stabilizes these oscillations and was found to be required for heterogeneous Cd and hydrogen-peroxide resistance, through the same pathway as Gsh1p. Expression of GTS1 from a constitutive tet-regulated promoter suppressed oscillations and heterogeneity in GSH content, and resulted in decreased variation in stress resistance. This enabled manipulation of the degree of gene expression noise in cultures. It was shown that cells expressing Gts1p heterogeneously had a competitive advantage over more-homogeneous cell populations (with the same mean Gts1p expression), under continuous and fluctuating stress conditions. The results establish a novel molecular mechanism for single-cell heterogeneity, and demonstrate experimentally fitness advantages that depend on deterministic variation in gene expression within cell populations. PMID:17919285

  5. Creation of a S1P Lyase bacterial surrogate for structure-based drug design.

    PubMed

    Argiriadi, Maria A; Banach, David; Radziejewska, Elzbieta; Marchie, Susan; DiMauro, Jennifer; Dinges, Jurgen; Dominguez, Eric; Hutchins, Charles; Judge, Russell A; Queeney, Kara; Wallace, Grier; Harris, Christopher M

    2016-05-01

    S1P Lyase (SPL) has been described as a drug target in the treatment of autoimmune diseases. It plays an important role in maintaining intracellular levels of S1P thereby affecting T cell egress from lymphoid tissues. Several groups have already published approaches to inhibit S1P Lyase with small molecules, which in turn increase endogenous S1P concentrations resulting in immunosuppression. The use of structural biology has previously aided SPL inhibitor design. Novel construct design is at times necessary to provide a reagent for protein crystallography. Here we present a chimeric bacterial protein scaffold used for protein X-ray structures in the presence of early small molecule inhibitors. Mutations were introduced to the bacterial SPL from Symbiobacterium thermophilum which mimic the human enzyme. As a result, two mutant StSPL crystal structures resolved to 2.8Å and 2.2Å resolutions were solved and provide initial structural hypotheses for an isoxazole chemical series, whose optimization is discussed in the accompanying paper.

  6. A non-death role of the yeast metacaspase: Yca1p alters cell cycle dynamics.

    PubMed

    Lee, Robin E C; Puente, Lawrence G; Kaern, Mads; Megeney, Lynn A

    2008-08-13

    Caspase proteases are a conserved protein family predominantly known for engaging and executing apoptotic cell death. Nevertheless, in higher eukaryotes, caspases also influence a variety of cell behaviors including differentiation, proliferation and growth control. S. cerevisiae expresses a primordial caspase, yca1, and exhibits apoptosis-like death under certain stresses; however, the benefit of a dedicated death program to single cell organisms is controversial. In the absence of a clear rationale to justify the evolutionary retention of a death only pathway, we hypothesize that yca1 also influences non-apoptotic events. We report that genetic ablation and/or catalytic inactivation of Yca1p leads to a longer G1/S transition accompanied by slower growth in fermentation conditions. Downregulation of Yca1p proteolytic activity also results in failure to arrest during nocodazole treatment, indicating that Yca1p participates in the G2/M mitotic checkpoint. 20s proteasome activity and ROS staining of the Delta yca1 strain is indistinguishable from its isogenic control suggesting that putative regulation of the oxidative stress response by Yca1p does not instigate the cell cycle phenotype. Our results demonstrate multiple non-death roles for yca1 in the cell cycle.

  7. CDC64 Encodes Cytoplasmic Alanyl-tRNA Synthetase, Ala1p, of Saccharomyces cerevisiae

    PubMed Central

    Wrobel, Carolyn; Schmidt, Emmett V.; Polymenis, Michael

    1999-01-01

    The cdc64-1 mutation causes G1 arrest in Saccharomyces cerevisiae corresponding to a type II Start phenotype. We report that CDC64 encodes Ala1p, an alanyl-tRNA synthetase. Thus, cdc64-1 might affect charging of tRNAAla and thereby initiation of cell division. PMID:10601222

  8. Biochemical regulation of breast cancer cell expression of S1P2 (Edg-5) and S1P3 (Edg-3) G protein-coupled receptors for sphingosine 1-phosphate.

    PubMed

    Dolezalova, Hana; Shankar, Geetha; Huang, Mei-Chuan; Bikle, Daniel D; Goetzl, Edward J

    2003-03-01

    G protein-coupled receptors (GPCRs) for lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P) transduce signals to many functions of normal cells. Most human cancer cells upregulate S1P and LPA GPCRs, in patterns distinctive for each type of tumor. The findings that 1-alpha, 25-dihydroxy-vitamin D(3) (VD3) and all-trans retinoic acid (RA) differentially alter expression of the predominant S1P(3) (Edg-3) R and S1P(2) (Edg-5) R in human breast cancer cells (BCCs) permitted analyses of their individual activities, despite a lack of selective pharmacological probes. S1P-evoked increases in [Ca(2+)](i) in S1P(3) R-predominant BCCs were suppressed by concentrations of VD3 and RA which decreased expression of S1P(3) Rs, despite RA-induced increases in S1P(2) Rs. S1P-elicited chemokinetic migration of S1P(3) R-predominant BCCs across a type IV collagen-coated micropore filter also was inhibited by concentrations of VD3 and RA which decreased expression of S1P(3) Rs. The RA-induced increase in expression of S1P(2) Rs did not prevent suppression by RA of S1P-elicited chemokinesis, which appears to be mediated by S1P(3) Rs, but instead exposed S1P(2) R-mediated inhibition of epidermal growth factor-stimulated chemotaxis of BCCs. In contrast, expression of the predominant LPA(2) Rs, LPA-evoked increase in [Ca(2+)](i) and LPA-stimulated chemokinetic migration were suppressed concomitantly by RA but not VD3. Thus two structurally-homologous S1P Rs of BCCs differ in coupling to [Ca(2+)](i) signaling and have opposite effects on protein growth factor-stimulated chemotaxis.

  9. Swelling and pillaring of the layered precursor IPC-1P: tiny details determine everything.

    PubMed

    Shamzhy, Mariya; Mazur, Michal; Opanasenko, Maksym; Roth, Wieslaw J; Čejka, Jiří

    2014-07-21

    The influence of swelling (i.e. the size of tetraalkylammonium surfactant molecule, the presence of tetrapropylammonium hydroxide (TPAOH), pH) and pillaring (i.e. the ratio between the swollen precursor IPC-1P and tetraethyl orthosilicate) conditions on the structure and textural properties of the resulting materials was studied in detail for IPC-1P, which is the layered precursor of zeolite PCR. The swelling of IPC-1P proceeds efficiently under basic conditions both in mixed C(n)H(2n+1)N(CH3)3Cl/TPAOH systems and in C(n)H(2n+1)N(CH3)3OH (n = 8, 10, 12, 14, 16, 18) solutions at pH = 13-14. The intercalation of C(n)H(2n+1)N(+)(CH3)3 in IPC-1P resulted in the formation of expanded materials with interlayer distances growing with increasing length of the alkyl chain in C(n)H(2n+1)N(CH3)3(+): 1.59-1.86 (n = 8) < 1.89-2.11 (10) < 2.05-2.26 (12) = 2.08-2.26 (14) < 2.37-2.43 (16) < 2.57-2.71 (18) Å. IPC-2 zeolite was formed during calcination of IPC-1P samples swollen in C(n)H(2n+1)N(CH3)3OH solution, while PCR zeolite can be obtained by calcination of IPC-1P treated with either C(n)H(2n+1)N(CH3)3Cl/TPAOH or C(n)H(2n+1)N(CH3)3Cl. The pillaring of IPC-1P samples swollen with C(n)H(2n+1)N(CH3)3OH provided mesoporous materials with narrow pore size distribution in the range 2.5-3.5 nm. Pillared materials derived from the samples swollen in the presence of TPAOH were characterized by a broader pore size distribution. The optimal TEOS/IPC-1PSW ratio being sufficient for the formation of well-ordered pillared derivatives characterized by improved textural properties (S(BET) = 878 m(2) g(-1), V(total) = 0.599 cm(3) g(-1)) was found to be 1 : 1.5.

  10. Mp1p Is a Virulence Factor in Talaromyces (Penicillium) marneffei

    PubMed Central

    Zhang, Hongmin; Lo, Raymond K. C.; Cai, Jian-Pao; Au-Yeung, Rex K. H.; Ng, Wing-Fung; Tse, Herman; Wong, Samson S. Y.; Xu, Simin; Lam, Wai Hei; Tse, Man-Kit; Sze, Kong Hung; Kao, Richard Y.; Reiner, Neil E.; Hao, Quan; Yuen, Kwok-Yung

    2016-01-01

    Background Talaromyces marneffei is an opportunistic dimorphic fungus prevalent in Southeast Asia. We previously demonstrated that Mp1p is an immunogenic surface and secretory mannoprotein of T. marneffei. Since Mp1p is a surface protein that can generate protective immunity, we hypothesized that Mp1p and/or its homologs are virulence factors. Methodology/Principal Findings We examined the pathogenic roles of Mp1p and its homologs in a mouse model. All mice died 21 and 30 days after challenge with wild-type T. marneffei PM1 and MP1 complemented mutant respectively. None of the mice died 60 days after challenge with MP1 knockout mutant (P<0.0001). Seventy percent of mice died 60 days after challenge with MP1 knockdown mutant (P<0.0001). All mice died after challenge with MPLP1 to MPLP13 knockdown mutants, suggesting that only Mp1p plays a significant role in virulence. The mean fungal loads of PM1 and MP1 complemented mutant in the liver, lung, kidney and spleen were significantly higher than those of the MP1 knockout mutant. Similarly, the mean load of PM1 in the liver, lung and spleen were significantly higher than that of the MP1 knockdown mutant. Histopathological studies showed an abundance of yeast in the kidney, spleen, liver and lung with more marked hepatic and splenic necrosis in mice challenged with PM1 compared to MP1 knockout and MP1 knockdown mutants. Likewise, a higher abundance of yeast was observed in the liver and spleen of mice challenged with MP1 complemented mutant compared to MP1 knockout mutant. PM1 and MP1 complemented mutant survived significantly better than MP1 knockout mutant in macrophages at 48 hours (P<0.01) post-infection. The mean fungal counts of Pichia pastoris GS115-MP1 in the liver (P<0.001) and spleen (P<0.05) of mice were significantly higher than those of GS115 at 24 hours post-challenge. Conclusions/Significance Mp1p is a key virulence factor of T. marneffei. Mp1p mediates virulence by improving the survival of T. marneffei

  11. The role of Cdh1p in maintaining genomic stability in budding yeast.

    PubMed Central

    Ross, Karen E; Cohen-Fix, Orna

    2003-01-01

    Cdh1p, a substrate specificity factor for the cell cycle-regulated ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C), promotes exit from mitosis by directing the degradation of a number of proteins, including the mitotic cyclins. Here we present evidence that Cdh1p activity at the M/G(1) transition is important not only for mitotic exit but also for high-fidelity chromosome segregation in the subsequent cell cycle. CDH1 showed genetic interactions with MAD2 and PDS1, genes encoding components of the mitotic spindle assembly checkpoint that acts at metaphase to prevent premature chromosome segregation. Unlike cdh1delta and mad2delta single mutants, the mad2delta cdh1delta double mutant grew slowly and exhibited high rates of chromosome and plasmid loss. Simultaneous deletion of PDS1 and CDH1 caused extensive chromosome missegregation and cell death. Our data suggest that at least part of the chromosome loss can be attributed to kinetochore/spindle problems. Our data further suggest that Cdh1p and Sic1p, a Cdc28p/Clb inhibitor, have overlapping as well as nonoverlapping roles in ensuring proper chromosome segregation. The severe growth defects of both mad2delta cdh1delta and pds1delta cdh1dDelta strains were rescued by overexpressing Swe1p, a G(2)/M inhibitor of the cyclin-dependent kinase, Cdc28p/Clb. We propose that the failure to degrade cyclins at the end of mitosis leaves cdh1delta mutant strains with abnormal Cdc28p/Clb activity that interferes with proper chromosome segregation. PMID:14573464

  12. Mutational analysis of Mdm1p function in nuclear and mitochondrial inheritance.

    PubMed

    Fisk, H A; Yaffe, M P

    1997-08-11

    Nuclear and mitochondrial transmission to daughter buds of Saccharomyces cerevisiae depends on Mdm1p, an intermediate filament-like protein localized to numerous punctate structures distributed throughout the yeast cell cytoplasm. These structures disappear and organelle inheritance is disrupted when mdm1 mutant cells are incubated at the restrictive temperature. To characterize further the function of Mdm1p, new mutant mdm1 alleles that confer temperature-sensitive growth and defects in organelle inheritance but produce stable Mdm1p structures were isolated. Microscopic analysis of the new mdm1 mutants revealed three phenotypic classes: Class I mutants showed defects in both mitochondrial and nuclear transmission; Class II alleles displayed defective mitochondrial inheritance but had no effect on nuclear movement; and Class III mutants showed aberrant nuclear inheritance but normal mitochondrial distribution. Class I and II mutants also exhibited altered mitochondrial morphology, possessing primarily small, round mitochondria instead of the extended tubular structures found in wild-type cells. Mutant mdm1 alleles affecting nuclear transmission were of two types: Class Ia and IIIa mutants were deficient for nuclear movement into daughter buds, while Class Ib and IIIb mutants displayed a complete transfer of all nuclear DNA into buds. The mutations defining all three allelic classes mapped to two distinct domains within the Mdm1p protein. Genetic crosses of yeast strains containing different mdm1 alleles revealed complex genetic interactions including intragenic suppression, synthetic phenotypes, and intragenic complementation. These results support a model of Mdm1p function in which a network comprised of multimeric assemblies of the protein mediates two distinct cellular processes.

  13. PPARγ agonists upregulate sphingosine 1-phosphate (S1P) receptor 1 expression, which in turn reduces S1P-induced [Ca(2+)]i increases in renal mesangial cells.

    PubMed

    Koch, Alexander; Völzke, Anja; Puff, Bianca; Blankenbach, Kira; Meyer Zu Heringdorf, Dagmar; Huwiler, Andrea; Pfeilschifter, Josef

    2013-11-01

    We previously identified peroxisome proliferator-activated receptor gamma (PPARγ) agonists (thiazolidinediones, TZDs) as modulators of the sphingolipid metabolism in renal mesangial cells. TZDs upregulated sphingosine kinase 1 (SK-1) and increased the formation of intracellular sphingosine 1-phosphate (S1P), which in turn reduced the expression of pro-fibrotic connective tissue growth factor. Since S1P also acts as extracellular ligand at specific S1P receptors (S1PR, S1P1-5), we investigated here the effect of TZDs on S1PR expression in mesangial cells and evaluated the functional consequences by measuring S1P-induced increases in intracellular free Ca(2+) concentration ([Ca(2+)]i). Treatment with two different TZDs, troglitazone and rosiglitazone, enhanced S1P1 mRNA and protein expression in rat mesangial cells, whereas S1P2-5 expression levels were not altered. Upregulation of S1P1 mRNA upon TZD treatment was also detected in human mesangial cells and mouse glomeruli. PPARγ antagonism and promoter studies revealed that the TZD-dependent S1P1 mRNA induction involved a functional PPAR response element in the S1P1 promoter. Pharmacological approaches disclosed that S1P-induced [Ca(2+)]i increases in rat mesangial cells were predominantly mediated by S1P2 and S1P3. Interestingly, the transcriptional upregulation of S1P1 by TZDs resulted in a reduction of S1P-induced [Ca(2+)]i increases, which was reversed by the S1P1/3 antagonist VPC-23019, the protein kinase C (PKC) inhibitor PKC-412, and by S1P1 siRNA. These data suggest that PPARγ-dependent upregulation of S1P1 leads to an inhibition of S1P-induced Ca(2+) signaling in a PKC-dependent manner. Overall, these results reveal that TZDs not only modulate intracellular S1P levels but also regulate S1PR signaling by increasing S1P1 expression in mesangial cells.

  14. Expression of S1P metabolizing enzymes and receptors correlate with survival time and regulate cell migration in glioblastoma multiforme.

    PubMed

    Bien-Möller, Sandra; Lange, Sandra; Holm, Tobias; Böhm, Andreas; Paland, Heiko; Küpper, Johannes; Herzog, Susann; Weitmann, Kerstin; Havemann, Christoph; Vogelgesang, Silke; Marx, Sascha; Hoffmann, Wolfgang; Schroeder, Henry W S; Rauch, Bernhard H

    2016-03-15

    A signaling molecule which is involved in proliferation and migration of malignant cells is the lipid mediator sphingosine-1-phosphate (S1P). There are hints for a potential role of S1P signaling in malignant brain tumors such as glioblastoma multiforme (GBM) which is characterized by a poor prognosis. Therefore, a comprehensive expression analysis of S1P receptors (S1P1-S1P5) and S1P metabolizing enzymes in human GBM (n = 117) compared to healthy brain (n = 10) was performed to evaluate their role for patient´s survival. Furthermore, influence of S1P receptor inhibition on proliferation and migration were studied in LN18 GBM cells. Compared to control brain, mRNA levels of S1P1, S1P2, S1P3 and S1P generating sphingosine kinase-1 were elevated in GBM. Kaplan-Meier analyses demonstrated an association between S1P1 and S1P2 with patient´s survival times. In vitro, an inhibitory effect of the SphK inhibitor SKI-II on viability of LN18 cells was shown. S1P itself had no effect on viability but stimulated LN18 migration which was blocked by inhibition of S1P1 and S1P2. The participation of S1P1 and S1P2 in LN18 migration was further supported by siRNA-mediated silencing of these receptors. Immunoblots and inhibition experiments suggest an involvement of the PI3-kinase/AKT1 pathway in the chemotactic effect of S1P in LN18 cells.In summary, our data argue for a role of S1P signaling in proliferation and migration of GBM cells. Individual components of the S1P pathway represent prognostic factors for patients with GBM. Perspectively, a selective modulation of S1P receptor subtypes could represent a therapeutic approach for GBM patients and requires further evaluation.

  15. Investigating the molecular mechanisms through which FTY720-P causes persistent S1P1 receptor internalization

    PubMed Central

    Sykes, David A; Riddy, Darren M; Stamp, Craig; Bradley, Michelle E; McGuiness, Neil; Sattikar, Afrah; Guerini, Danilo; Rodrigues, Ines; Glaenzel, Albrecht; Dowling, Mark R; Mullershausen, Florian; Charlton, Steven J

    2014-01-01

    Background and Purpose The molecular mechanism underlying the clinical efficacy of FTY720-P is thought to involve persistent internalization and enhanced degradation of the S1P1 receptor subtype (S1P1R). We have investigated whether receptor binding kinetics and β-arrestin recruitment could play a role in the persistent internalization of the S1P1R by FTY720-P. Experimental Approach [3H]-FTY720-P and [33P]-S1P were used to label CHO-S1P1/3Rs for binding studies. Ligand efficacy was assessed through [35S]-GTPγS binding and β-arrestin recruitment. Metabolic stability was evaluated using a bioassay measuring intracellular Ca2+ release. CHO-S1P1/3R numbers were determined, following FTY720-P treatment using flow cytometry. Key Results The kinetic off-rate of [3H]-FTY720-P from the S1P1R was sixfold slower than from the S1P3R, and comparable to [33P]-S1P dissociation from S1P1/3Rs. S1P and FTY720-P stimulated [35S]-GTPγS incorporation to similar degrees, but FTY720-P was over 30-fold less potent at S1P3Rs. FTY720-P stimulated a higher level of β-arrestin recruitment at S1P1Rs, 132% of the total recruited by S1P. In contrast, FTY720-P was a weak partial agonist at S1P3R, stimulating just 29% of the total β-arrestin recruited by S1P. Internalization experiments confirmed that cell surface expression of the S1P1R but not the S1P3R was reduced following a pulse exposure to FTY720-P, which is metabolically stable unlike S1P. Conclusions and Implications FTY720-P and S1P activation of the S1P1R results in receptor internalization as a consequence of an efficient recruitment of β-arrestin. The combination of slow off-rate, efficacious β-arrestin recruitment and metabolic stability all contribute to FTY720-P's ability to promote prolonged S1P1R internalization and may be critical factors in its efficacy in the clinic. PMID:24641481

  16. Induction of apoptosis by quercetin is mediated through AMPKalpha1/ASK1/p38 pathway.

    PubMed

    Lee, Yun-Kyoung; Hwang, Jin-Taek; Kwon, Dae Young; Surh, Young-Joon; Park, Ock Jin

    2010-06-28

    Effective strategies for cancer prevention and treatment can be identified by understanding the mechanism of apoptotic pathways. In this study, we investigated the regulatory mechanism of quercetin-induced apoptosis through apoptosis signal-regulating kinase (ASK)-1 and mitogen-activated protein kinase pathways. Our results showed that quercetin increased apoptotic cell death through reactive oxygen species (ROS) generation and was responsible for ASK1 activation. Increasing ASK1 activity was accompanied by p38 activation. Interestingly, AMP-activated protein kinase (AMPK) seemed to be a critical controller of quercetin-regulated ASK1/p38 activation. Blocking AMPKalpha1 activity using Compound C, a synthetic inhibitor or siRNA showed that quercetin-activated ASK1 could not stimulate p38 activity. Thus, we suggested that quercetin-exerted apoptotic effects involve ROS/AMPKalpha1/ASK1/p38 signaling pathway, and AMPKalpha1 is a necessary element for apoptotic event induced by ASK1.

  17. Complex structural rearrangement features suggesting chromoanagenesis mechanism in a case of 1p36 deletion syndrome.

    PubMed

    Zanardo, Évelin Aline; Piazzon, Flavia Balbo; Dutra, Roberta Lelis; Dias, Alexandre Torchio; Montenegro, Marília Moreira; Novo-Filho, Gil Monteiro; Costa, Thaís Virgínia Moura Machado; Nascimento, Amom Mendes; Kim, Chong Ae; Kulikowski, Leslie Domenici

    2014-12-01

    Genome rearrangements are caused by the erroneous repair of DNA double-strand breaks, leading to several alterations that result in loss or gain of the structural genomic of a dosage-sensitive genes. However, the mechanisms that promote the complexity of rearrangements of congenital or developmental defects in human disease are unclear. The investigation of complex genomic abnormalities could help to elucidate the mechanisms and causes for the formation and facilitate the understanding of congenital or developmental defects in human disease. We here report one case of a patient with atypical clinical features of the 1p36 syndrome and the use of cytogenomic techniques to characterize the genomic alterations. Analysis by multiplex ligation-dependent probe amplification and array revealed a complex rearrangement in the 1p36.3 region with deletions and duplication interspaced by normal sequences. We also suggest that chromoanagenesis could be a possible mechanism involved in the repair and stabilization of this rearrangement.

  18. Ixr1p and the control of the Saccharomyces cerevisiae hypoxic response.

    PubMed

    Vizoso-Vázquez, Angel; Lamas-Maceiras, Mónica; Becerra, Manuel; González-Siso, M Isabel; Rodríguez-Belmonte, Esther; Cerdán, M Esperanza

    2012-04-01

    In Saccharomyces cerevisiae, adaptation to hypoxia/anaerobiosis requires the transcriptional induction or derepression of multiple genes organized in regulons controlled by specific transcriptional regulators. Ixr1p is a transcriptional regulatory factor that causes aerobic repression of several hypoxic genes (COX5B, TIR1, and HEM13) and also the activation of HEM13 during hypoxic growth. Analysis of the transcriptome of the wild-type strain BY4741 and its isogenic derivative Δixr1, grown in aerobic and hypoxic conditions, reveals differential regulation of genes related not only to the hypoxic and oxidative stress responses but also to the re-adaptation of catabolic and anabolic fluxes in response to oxygen limitation. The function of Ixr1p in the transcriptional regulation of genes from the sulfate assimilation pathway and other pathways producing α-keto acids is of biotechnological importance for industries based on yeast-derived fermentation products.

  19. Characterization of Vta1p, a class E Vps protein in Saccharomyces cerevisiae.

    PubMed

    Shiflett, Shelly L; Ward, Diane McVey; Huynh, Dinh; Vaughn, Michael B; Simmons, Jennifer C; Kaplan, Jerry

    2004-03-19

    We identified VTA1 in a screen for mutations that result in altered vacuole morphology. Deletion of VTA1 resulted in delayed trafficking of the lipophilic dye FM4-64 to the vacuole and altered vacuolar morphology when cells were exposed to the dye 5-(and 6)-carboxy-2',7'-dichlorofluorescein diacetate (CDCFDA). Deletion of class E vacuolar protein sorting (VPS) genes, which encode proteins that affect multivesicular body formation, also showed altered vacuolar morphology upon exposure to high concentrations of CDCFDA. These results suggest a VPS defect for Deltavta1 cells. Deletion of VTA1 did not affect growth on raffinose and only mildly affected carboxypeptidase S sorting. Turnover of the surface protein Ste3p, the a-factor receptor, was affected in Deltavta1 cells with the protein accumulating on the vacuolar membrane. Likewise the alpha-factor receptor Ste2p accumulated on the vacuolar membrane in Deltavta1 cells. We demonstrated that many class E VPS deletion strains are hyper-resistant to the cell wall disruption agent calcofluor white. Deletion of VTA1 or VPS60, another putative class E gene, resulted in calcofluor white hypersensitivity. A Vta1p-green fluorescent protein fusion protein transiently associated with a Pep12p-positive compartment. This localization was altered by deletion of many of the class E VPS genes, indicating that Vta1p binds to endosomes in a manner dependent on the assembly of the endosomal sorting complexes required for transport. Membrane-associated Vta1p co-purified with Vps60p, suggesting that Vta1p is a class E Vps protein that interacts with Vps60p on a prevacuolar compartment.

  20. Maladaptive exploratory behavior and neuropathology of the PS-1 P117L Alzheimer transgenic mice.

    PubMed

    Zufferey, Valérie; Vallet, Philippe G; Moeri, Michaël; Moulin-Sallanon, Marcelle; Piotton, Françoise; Marin, Pascale; Savioz, Armand

    2013-05-01

    Patients with the early-onset Alzheimer's disease P117L mutation in the presenilin-1 gene (PS-1) present pathological hallmarks in the hippocampus, the frontal cortex and the basal ganglia. In the present work we determined by immunohistochemistry which brain regions were injured in the transgenic PS-1 P117L mice, in comparison to their littermates, the B6D2 mice. Furthermore, as these regions are involved in novelty detection, we investigated the behavior of these mice in tests for object and place novelty recognition. Limited numbers of senile plaques and neurofibrillary tangles were detected in aged PS-1 P117L mice in the CA1 only, indicating that the disease is restrained to an initial neuropathological stage. Western blots showed a change in PSD-95 expression (p=0.03), not in NR2A subunit, NR2B subunit and synaptophysin expressions in the frontal cortex, suggesting specific synaptic alterations. The behavioral tests repeatedly revealed, despite a non-significant preference for object or place novelty, maladaptive exploratory behavior of the PS-1 P117L mice in novel environmental conditions, not due to locomotor problems. These mice, unlike the B6D2 mice, were less inhibited to visit the center of the cages (p=0.01) and they continued to move excessively in the presence of a displaced object (p=0.021). Overall, the PS-1 P117L mice appear to be in an initial Alzheimer's disease-like neuropathological stage, and they showed a lack of reaction toward novel environmental conditions.

  1. Strong association of chromosome 1p12 loci with thyroid cancer susceptibility.

    PubMed

    Baida, Aida; Akdi, Mounaim; González-Flores, Eddy; Galofré, Pere; Marcos, Ricard; Velázquez, Antonia

    2008-06-01

    Several genes directly related to thyroid cancer development have been described; nevertheless, the genetic pathways of this tumorigenesis process are unknown. Together with environmental factors, susceptibility genes could have an important role in thyroid cancer. Our previous studies suggest that the chromosome 1p12-13 is related to thyroid cancer incidence. Here, we extend the analysis with a case-control association study in a Spanish population. Thus, six single-nucleotide polymorphisms were genotyped, covering 2.4 Mb of the 1p12-13 region. A statistically significant association between thyroid cancer incidence and the rs2145418 and rs4658973 polymorphisms was found (P < 0.0001). No association was detected for the other four polymorphisms studied. The rs2145418 marker showed a significant odds ratio of 5.0 [95% confidence interval (95% CI), 2.85-8.83] and 9.2 (95% CI, 4.50-21.6) for heterozygous and homozygous G-variant alleles, respectively. For rs4658973, the odds ratios were 0.40 (95% CI, 0.26-0.62) and 0.07 (95% CI, 0.03-0.18) for heterozygous and homozygous G-variant alleles, respectively. These markers map into the 1p12 region, and no linkage disequilibrium was found between them, indicating an independent relation of these polymorphisms with thyroid cancer susceptibility. Our data provide evidence of a strong association of the chromosome 1p12 with thyroid cancer risk, and it is the first study describing susceptibility loci for thyroid cancer in this region.

  2. Comparing CN Features in Two Comets: 1P/Halley and 103P/Hartley 2

    NASA Astrophysics Data System (ADS)

    Samarasinha, Nalin H.; Lejoly, Cassandra; Barrera, Jose; Mueller, Beatrice; Schleicher, David

    2015-11-01

    Comets 1P/Halley and 103P/Hartley 2 show distinct CN features in their respecive comae. Both comets are non-principal-axis rotators. 1P/Halley is the proto-type for Halley-type comets with the Oort Cloud as its possible source region, whereas 103P/Hartley 2 is a Jupiter-Family comet that possibly originated from the Kuiper Belt. Both comets were spacecraft targets and studied widely from both space and from the ground.We will discuss the properties of CN features, and in particular the behavior of the derived outflow velocities based on the CN features present in the groundbased coma images of these two comets. The corresponding heliocentric distances for CN images of comet 1P/Halley range from approximately 0.8 AU to 2.0 AU (during its post-perihelion leg of the 1986 apparition). For CN images of comet 103P/Hartley 2, the corresponding heliocentric distances range from 1.31 AU through the perihelion (at 1.06 AU) to 1.25 AU (during its 2010 apparition).Ultimately, these results will be used to understand the rotational states and the activity behaviors of these two comets.

  3. Recurrent interstitial 1p36 deletions: Evidence for germline mosaicism and complex rearrangement breakpoints.

    PubMed

    Gajecka, Marzena; Saitta, Sulagna C; Gentles, Andrew J; Campbell, Lindsey; Ciprero, Karen; Geiger, Elizabeth; Catherwood, Anne; Rosenfeld, Jill A; Shaikh, Tamim; Shaffer, Lisa G

    2010-12-01

    Deletions of chromosome 1p36 are one of the most frequently encountered subtelomeric alterations. Clinical features of monosomy 1p36 include neurocognitive impairment, hearing loss, seizures, cardiac defects, and characteristic facial features. The majority of cases have occurred sporadically, implying that genomic instability plays a role in the prevalence of the syndrome. Here, we report two siblings with mild phenotypic features of the deletion syndrome, including developmental delay, hearing loss, and left ventricular non-compaction (LVNC). Microarray analysis using bacterial artificial chromosome and oligonucleotide microarrays indicated the deletions were identical, suggesting germline mosaicism. Parental phenotypes were normal, and analysis by fluorescence in situ hybridization (FISH) did not show mosaicism. These small interstitial deletions were not detectable by conventional subtelomeric FISH analysis. To investigate the mechanism of deletion further, the breakpoints were cloned and sequenced, demonstrating the presence of a complex rearrangement. Sequence analysis of genes in the deletion interval did not reveal any mutations on the intact homologue that may have contributed to the LVNC seen in both children. This is the first report of apparent germline mosaicism for this disorder. Thus, our findings have important implications for diagnostic approaches and for recurrence risk counseling in families with a child with monosomy 1p36. In addition, our results further refine the minimal critical region for LVNC and hearing loss.

  4. Is 1p36 deletion associated with anterior body wall defects?

    PubMed

    Çöllü, Medis; Yüksel, Şirin; Şirin, Başak Kumbasar; Abbasoğlu, Latif; Alanay, Yasemin

    2016-07-01

    Epispadias and exstrophy of the cloaca, also known as OEIS complex (omphalocele, exstrophy, imperforate anus, spinal defects), respectively constitute the most benign and severe ends of the bladder exstrophy-epispadias complex (BEEC) spectrum. In 2009, El-Hattab et al. reported the first patient with OEIS complex associated with a chromosome 1p36 deletion. Here we report a second patient with 1p36 deletion who also has classic bladder exstrophy, supporting the possible role of genes in this region in the development of BEEC. The absence of omphalocele and imperforate anus in our patient places him toward classic bladder exstrophy while presence of spina bifida and the absence of coccyx suggest an overlap with OEIS complex. An additional differential diagnosis is the pentalogy of Cantrell in our patient as he also has a diaphragmatic hernia and an incomplete sternum. This is the second observation of a ventral midline birth defect in association with 1p36 deletion syndrome, following El-Hattab et al.'s report [2009]. The three genes (NOCL2, DVL1, and MMP23B) discussed as possible candidates are also among the deleted ones in our patient, supporting the possible role of these genes in BEEC spectrum. © 2016 Wiley Periodicals, Inc.

  5. Flexibility of the exportins Cse1p and Xpot depicted by elastic network model.

    PubMed

    Hu, Mingwen; Kim, Byung

    2011-07-01

    Nucleocytoplasmic transport in eukaryotic cells involves many interactions between macromolecules, and has been an active area for many researchers. However, the precise mechanism still evades us and more efforts are needed to better understand it. In this study, the authors investigated exportins (Cse1p and Xpot) by elastic network interpolation (ENI) and elastic network based normal mode analysis (EN-NMA). Results of the study on Cse1p were in good agreement with the results obtained by molecular dynamics simulation in another study but with the benefit of time-efficiency. First, a formation of ring closure obtained by ENI was observed. Second, HEAT 1 to 3 and HEAT 14 to 17 had the largest values of root mean square deviation (RMSD) which indicated the flexibility of Cse1p during the transition. In the case of Xpot, a possible pathway from nuclear state to cytoplasmic state was shown, and the predicted pathway was also quantitatively analyzed in terms of RMSD. The results suggested two flexible regions of Xpot that might be important to the transporting mechanism. Moreover, the dominant mode of Xpot in the nuclear state obtained by EN-NMA not only showed the tendency to match the predicted pathway to the cytoplasmic state of Xpot, but also displayed the flexible regions of Xpot. A time-efficient computational approach was presented in this paper and the results indicated that the flexibility of tested exportins might be required to perform the biological function of transporting cargos.

  6. Sequestosome 1/p62, a scaffolding protein, is a newly identified partner of IRS-1 protein.

    PubMed

    Geetha, Thangiah; Zheng, Chen; Vishwaprakash, Nilmini; Broderick, Tom L; Babu, Jeganathan Ramesh

    2012-08-24

    Defects in the insulin-signaling pathway may lead to the development of skeletal muscle insulin resistance, which is one of the earliest abnormalities detected in individuals with the metabolic syndrome and predisposes them to develop type 2 diabetes. Previous studies have shown that deletion of the mouse sequestosome 1/p62 gene results in mature-onset obesity that progresses to insulin and leptin resistance and, ultimately, type 2 diabetes. Sequestosome 1/p62 is involved in receptor-mediated signal transduction and functions as an intracellular signal modulator or adaptor protein. Insulin receptor substrate-1 (IRS-1) plays a central role in transducing the insulin signal via phosphorylation, protein-protein interactions, and protein modifications. Mapping studies demonstrated that the SH(2) domain at the amino terminus of sequestosome 1/p62 interacts with IRS-1 upon insulin stimulation. Further, IRS-1 interacts with p62 through its YMXM motifs at Tyr-608, Tyr-628, and/or Tyr-658 in a manner similar to its interaction with p85 of phosphoinositol 3-kinase. Overexpression of p62 increased phosphorylation of Akt, GLUT4 translocation, and glucose uptake, providing evidence that p62 participates in the insulin-signaling pathway through its interactions with IRS-1.

  7. The yeast vacuolar ABC transporter Ybt1p regulates membrane fusion through Ca2+ transport modulation

    PubMed Central

    Sasser, Terry L.; Padolina, Mark; Fratti, Rutilio A.

    2013-01-01

    Ybt1p is a class C ABC transporter (ATP-binding cassette transporter) that is localized to the vacuole of Saccharomyces cerevisiae. Although Ybt1p was originally identified as a bile acid transporter, it has also been found to function in other capacities, including the translocation of phosphatidylcholine to the vacuole lumen, and the regulation of Ca2+ homoeostasis. In the present study we found that deletion of YBT1 enhanced in vitro homotypic vacuole fusion by up to 50 % relative to wild-type vacuoles. The increased vacuole fusion was not due to aberrant protein sorting of SNAREs (soluble N-ethylmaleimide-sensitive factor-attachment protein receptors) or recruitment of factors from the cytosol such as Ypt7p and the HOPS (homotypic fusion and vacuole protein sorting) tethering complex. In addition, ybt1Δ vacuoles displayed no observable differences in the formation of SNARE complexes, interactions between SNAREs and HOPS, or formation of vertex microdomains. However, the absence of Ybt1p caused significant changes in Ca2+ transport during fusion. One difference was the prolonged Ca2+ influx exhibited by ybt1Δ vacuoles at the start of the fusion reaction. We also observed a striking delay in SNARE-dependent Ca2+ efflux. As vacuole fusion can be inhibited by high Ca2+ concentrations, we suggest that the delayed efflux in ybt1Δ vacuoles leads to the enhanced SNARE function. PMID:22970809

  8. RACK1/Asc1p, a Ribosomal Node in Cellular Signaling

    PubMed Central

    Rachfall, Nicole; Schmitt, Kerstin; Bandau, Susanne; Smolinski, Nadine; Ehrenreich, Armin; Valerius, Oliver; Braus, Gerhard H.

    2013-01-01

    RACK1/Asc1p and its essential orthologues in higher eukaryotes, such as RACK1 in metazoa, are involved in several distinct cellular signaling processes. The implications of a total deletion have never been assessed in a comprehensive manner. This study reveals the major cellular processes affected in a Saccharomyces cerevisiae Δasc1 deletion background via de novo proteome and transcriptome analysis, as well as subsequent phenotypical characterizations. The deletion of ASC1 reduces iron uptake and causes nitrosative stress, both known indicators for hypoxia, which manifests in a shift of energy metabolism from respiration to fermentation in the Δasc1 strain. Asc1p further impacts cellular metabolism through its regulative role in the MAP kinase signal transduction pathways of invasive/filamentous growth and cell wall integrity. In the Δasc1 mutant strain, aberrations from the expected cellular response, mediated by these pathways, can be observed and are linked to changes in protein abundances of pathway-targeted transcription factors. Evidence of the translational regulation of such transcription factors suggests that ribosomal Asc1p is involved in signal transduction pathways and controls the biosynthesis of the respective final transcriptional regulators. PMID:23071099

  9. Binding properties of monoclonal antibodies recognizing external epitopes of the human MDR1 P-glycoprotein.

    PubMed

    Schinkel, A H; Arceci, R J; Smit, J J; Wagenaar, E; Baas, F; Dollé, M; Tsuruo, T; Mechetner, E B; Roninson, I B; Borst, P

    1993-09-30

    Monoclonal antibodies (MAbs) recognizing external epitopes of the human MDR1 P-glycoprotein have been used both for the detection of multidrug-resistant cells and as specific inhibitors of P-glycoprotein-mediated multidrug resistance. Using a panel of recently developed transfected or transgenic cell lines containing variants of the human MDR1 and MDR3 P-glycoproteins, we have compared the specificity and binding properties of the previously isolated MAbs MRK16, HYB-241, UIC2 and 4E3, and of the newly isolated MAb 7G4. The removal of 1, 2 or all 3 of the N-glycosylation sites present in the first extracellular loop of MDR1 P-glycoprotein did not significantly affect the binding of these MAbs. In contrast, 20 amino acid deletion in the first extracellular loop of MDR1 P-glycoprotein completely abolished binding of UIC2, whereas the binding of all other MAbs was hardly affected. None of the MAbs tested bound detectably to cell lines containing a high level of the human MDR3 P-glycoprotein. The differences in the binding specificity between UIC2 and the other tested antibodies parallel the reported functional differences in the ability of these antibodies to inhibit P-glycoprotein-mediated drug efflux.

  10. The membrane remodeling protein Pex11p activates the GTPase Dnm1p during peroxisomal fission

    PubMed Central

    Opalinski, Lukasz; Landgraf, Christiane; Costello, Joseph; Schrader, Michael; Krikken, Arjen M.; Knoops, Kèvin; Kram, Anita M.; Volkmer, Rudolf; van der Klei, Ida J.

    2015-01-01

    The initial phase of peroxisomal fission requires the peroxisomal membrane protein Peroxin 11 (Pex11p), which remodels the membrane, resulting in organelle elongation. Here, we identify an additional function for Pex11p, demonstrating that Pex11p also plays a crucial role in the final step of peroxisomal fission: dynamin-like protein (DLP)-mediated membrane scission. First, we demonstrate that yeast Pex11p is necessary for the function of the GTPase Dynamin-related 1 (Dnm1p) in vivo. In addition, our data indicate that Pex11p physically interacts with Dnm1p and that inhibiting this interaction compromises peroxisomal fission. Finally, we demonstrate that Pex11p functions as a GTPase activating protein (GAP) for Dnm1p in vitro. Similar observations were made for mammalian Pex11β and the corresponding DLP Drp1, indicating that DLP activation by Pex11p is conserved. Our work identifies a previously unknown requirement for a GAP in DLP function. PMID:25941407

  11. Sequestosome 1/p62, a Scaffolding Protein, Is a Newly Identified Partner of IRS-1 Protein*

    PubMed Central

    Geetha, Thangiah; Zheng, Chen; Vishwaprakash, Nilmini; Broderick, Tom L.; Babu, Jeganathan Ramesh

    2012-01-01

    Defects in the insulin-signaling pathway may lead to the development of skeletal muscle insulin resistance, which is one of the earliest abnormalities detected in individuals with the metabolic syndrome and predisposes them to develop type 2 diabetes. Previous studies have shown that deletion of the mouse sequestosome 1/p62 gene results in mature-onset obesity that progresses to insulin and leptin resistance and, ultimately, type 2 diabetes. Sequestosome 1/p62 is involved in receptor-mediated signal transduction and functions as an intracellular signal modulator or adaptor protein. Insulin receptor substrate-1 (IRS-1) plays a central role in transducing the insulin signal via phosphorylation, protein-protein interactions, and protein modifications. Mapping studies demonstrated that the SH2 domain at the amino terminus of sequestosome 1/p62 interacts with IRS-1 upon insulin stimulation. Further, IRS-1 interacts with p62 through its YMXM motifs at Tyr-608, Tyr-628, and/or Tyr-658 in a manner similar to its interaction with p85 of phosphoinositol 3-kinase. Overexpression of p62 increased phosphorylation of Akt, GLUT4 translocation, and glucose uptake, providing evidence that p62 participates in the insulin-signaling pathway through its interactions with IRS-1. PMID:22761437

  12. The Rtr1p CTD phosphatase autoregulates its mRNA through a degradation pathway involving the REX exonucleases

    PubMed Central

    Hodko, Domagoj; Ward, Taylor; Chanfreau, Guillaume

    2016-01-01

    Rtr1p is a phosphatase that impacts gene expression by modulating the phosphorylation status of the C-terminal domain of the large subunit of RNA polymerase II. Here, we show that Rtr1p is a component of a novel mRNA degradation pathway that promotes its autoregulation through turnover of its own mRNA. We show that the 3′UTR of the RTR1 mRNA contains a cis element that destabilizes this mRNA. RTR1 mRNA turnover is achieved through binding of Rtr1p to the RTR1 mRNP in a manner that is dependent on this cis element. Genetic evidence shows that Rtr1p-mediated decay of the RTR1 mRNA involves the 5′-3′ DExD/H-box RNA helicase Dhh1p and the 3′-5′ exonucleases Rex2p and Rex3p. Rtr1p and Rex3p are found associated with Dhh1p, suggesting a model for recruiting the REX exonucleases to the RTR1 mRNA for degradation. Rtr1p-mediated decay potentially impacts additional transcripts, including the unspliced BMH2 pre-mRNA. We propose that Rtr1p may imprint its RNA targets cotranscriptionally and determine their downstream degradation mechanism by directing these transcripts to a novel turnover pathway that involves Rtr1p, Dhh1p, and the REX family of exonucleases. PMID:26843527

  13. Ceramide synthase 2 facilitates S1P-dependent egress of thymocytes into the circulation in mice.

    PubMed

    Rieck, Michael; Kremser, Christiane; Jobin, Katarzyna; Mettke, Elisabeth; Kurts, Christian; Gräler, Markus; Willecke, Klaus; Kolanus, Waldemar

    2017-02-15

    Well-defined gradients of the lipid mediator sphingosine-1-phosphate (S1P) direct chemotactic egress of mature thymocytes from the thymus into the circulation. Although it is known that these gradients result from low S1P levels in the thymic parenchyma and high S1P concentrations at the exit sites and in the plasma, the biochemical mechanisms that regulate these differential S1P levels remain unclear. Several studies demonstrated that ceramide synthase 2 (Cers2) regulates the levels of the S1P precursor sphingosine. We, therefore, investigated whether Cers2 is involved in the regulation of S1P gradients and S1P-dependent egress into the circulation. By analyzing Cers2-deficient mice, we demonstrate that Cers2 limits the levels of S1P in thymus and blood to maintain functional S1P gradients that mediate thymocyte emigration into the circulation. This function is specific for Cers2, as we also show that Cers4 is not involved in the regulation of thymic egress. Our study identified Cers2 as an important regulator of S1P-dependent thymic egress, and thus contributes to the understanding of how S1P gradients are maintained in vivo.

  14. Edg8/S1P5: an oligodendroglial receptor with dual function on process retraction and cell survival.

    PubMed

    Jaillard, C; Harrison, S; Stankoff, B; Aigrot, M S; Calver, A R; Duddy, G; Walsh, F S; Pangalos, M N; Arimura, N; Kaibuchi, K; Zalc, B; Lubetzki, C

    2005-02-09

    Endothelial differentiation gene (Edg) proteins are G-protein-coupled receptors activated by lysophospholipid mediators: sphingosine-1-phosphate (S1P) or lysophosphatidic acid. We show that in the CNS, expression of Edg8/S1P5, a high-affinity S1P receptor, is restricted to oligodendrocytes and expressed throughout development from the immature stages to the mature myelin-forming cell. S1P activation of Edg8/S1P5 on O4-positive pre-oligodendrocytes induced process retraction via a Rho kinase/collapsin response-mediated protein signaling pathway, whereas no retraction was elicited by S1P on these cells derived from Edg8/S1P5-deficient mice. Edg8/S1P5-mediated process retraction was restricted to immature cells and was no longer observed at later developmental stages. In contrast, S1P activation promoted the survival of mature oligodendrocytes but not of pre-oligodendrocytes. The S1P-induced survival of mature oligodendrocytes was mediated through a pertussis toxin-sensitive, Akt-dependent pathway. Our data demonstrate that Edg8/S1P5 activation on oligodendroglial cells modulates two distinct functional pathways mediating either process retraction or cell survival and that these effects depend on the developmental stage of the cell.

  15. ER-associated SNAREs and Sey1p mediate nuclear fusion at two distinct steps during yeast mating.

    PubMed

    Rogers, Jason V; Arlow, Tim; Inkellis, Elizabeth R; Koo, Timothy S; Rose, Mark D

    2013-12-01

    During yeast mating, two haploid nuclei fuse membranes to form a single diploid nucleus. However, the known proteins required for nuclear fusion are unlikely to function as direct fusogens (i.e., they are unlikely to directly catalyze lipid bilayer fusion) based on their predicted structure and localization. Therefore we screened known fusogens from vesicle trafficking (soluble N-ethylmaleimide-sensitive factor attachment protein receptors [SNAREs]) and homotypic endoplasmic reticulum (ER) fusion (Sey1p) for additional roles in nuclear fusion. Here we demonstrate that the ER-localized SNAREs Sec20p, Ufe1p, Use1p, and Bos1p are required for efficient nuclear fusion. In contrast, Sey1p is required indirectly for nuclear fusion; sey1Δ zygotes accumulate ER at the zone of cell fusion, causing a block in nuclear congression. However, double mutants of Sey1p and Sec20p, Ufe1p, or Use1p, but not Bos1p, display extreme ER morphology defects, worse than either single mutant, suggesting that retrograde SNAREs fuse ER in the absence of Sey1p. Together these data demonstrate that SNAREs mediate nuclear fusion, ER fusion after cell fusion is necessary to complete nuclear congression, and there exists a SNARE-mediated, Sey1p-independent ER fusion pathway.

  16. Crz1p regulates pH homeostasis in Candida glabrata by altering membrane lipid composition.

    PubMed

    Yan, Dongni; Lin, Xiaobao; Qi, Yanli; Liu, Hui; Chen, Xiulai; Liu, Liming; Chen, Jian

    2016-09-23

    The asexual facultative aerobic haploid yeast Candida glabrata is widely used in the industrial production of various organic acids. To elucidate the physiological function of the transcription factor CgCrz1p and its role in tolerance to acid stress we deleted or overexpressed the corresponding gene CgCRZ1 Deletion of CgCRZ1 resulted in a 60% decrease in dry cell weight (DCW) and a 50% drop in cell viability compared to the wild type at pH 2.0. Expression of lipid metabolism-associated genes was also significantly down-regulated. Consequently, the proportion of C18:1 fatty acids, ratio of unsaturated to saturated fatty acids, and ergosterol content decreased by 30%, 46%, and 30%, respectively. Additionally, membrane integrity, fluidity, and H(+)-ATPase activity were reduced by 45%, 9%, and 50%, respectively. In contrast, overexpression of CgCrz1p increased C18:1 and ergosterol content by 16% and 40%, respectively. Overexpression also enhanced membrane integrity, fluidity, and H(+)-ATPase activity by 31%, 6%, and 20%, respectively. Moreover, in the absence of pH buffering, DCW and pyruvate titer increased by 48% and 60%, respectively, compared to the wild type. Together, these results suggest that CgCrz1p regulates tolerance to acidic conditions by altering membrane lipid composition in C. glabrata IMPORTANCE: The present study provides an insight into the metabolism of Candida glabrata under acidic conditions, such as those encountered during industrial production of organic acids. We found that overexpression of the transcription factor CgCrz1p improved viability, biomass, and pyruvate yields at low pH. Analysis of plasma membrane lipid composition indicated that CgCrz1p might play an important role in its integrity and fluidity, and enhanced the pumping of protons in acidic environments. We propose that altering the structure of the cell membrane may provide a successful strategy for increasing C glabrata productivity at low pH.

  17. The Integral Membrane Protein Snl1p Is Genetically Linked to Yeast Nuclear Pore Complex Function

    PubMed Central

    Ho, Albert K.; Raczniak, Gregory A.; Ives, Eric B.; Wente, Susan R.

    1998-01-01

    Integral membrane proteins are predicted to play key roles in the biogenesis and function of nuclear pore complexes (NPCs). Revealing how the transport apparatus is assembled will be critical for understanding the mechanism of nucleocytoplasmic transport. We observed that expression of the carboxyl-terminal 200 amino acids of the nucleoporin Nup116p had no effect on wild-type yeast cells, but it rendered the nup116 null strain inviable at all temperatures and coincidentally resulted in the formation of nuclear membrane herniations at 23°C. To identify factors related to NPC function, a genetic screen for high-copy suppressors of this lethal nup116-C phenotype was conducted. One gene (designated SNL1 for suppressor of nup116-C lethal) was identified whose expression was necessary and sufficient for rescuing growth. Snl1p has a predicted molecular mass of 18.3 kDa, a putative transmembrane domain, and limited sequence similarity to Pom152p, the only previously identified yeast NPC-associated integral membrane protein. By both indirect immunofluorescence microscopy and subcellular fractionation studies, Snl1p was localized to both the nuclear envelope and the endoplasmic reticulum. Membrane extraction and topology assays suggested that Snl1p was an integral membrane protein, with its carboxyl-terminal region exposed to the cytosol. With regard to genetic specificity, the nup116-C lethality was also suppressed by high-copy GLE2 and NIC96. Moreover, high-copy SNL1 suppressed the temperature sensitivity of gle2–1 and nic96-G3 mutant cells. The nic96-G3 allele was identified in a synthetic lethal genetic screen with a null allele of the closely related nucleoporin nup100. Gle2p physically associated with Nup116p in vitro, and the interaction required the N-terminal region of Nup116p. Therefore, genetic links between the role of Snl1p and at least three NPC-associated proteins were established. We suggest that Snl1p plays a stabilizing role in NPC structure and function

  18. Yos1p Is a Novel Subunit of the Yip1p–Yif1p Complex and Is Required for Transport between the Endoplasmic Reticulum and the Golgi Complex

    PubMed Central

    Heidtman, Matthew; Chen, Catherine Z.; Collins, Ruth N.; Barlowe, Charles

    2005-01-01

    Yeast Yip1p is a member of a conserved family of transmembrane proteins that interact with Rab GTPases. Previous studies also have indicated a role for Yip1p in the biogenesis of endoplasmic reticulum (ER)-derived COPII transport vesicles. In this report, we describe the identification and characterization of the uncharacterized open reading frame YER074W-A as a novel multicopy suppressor of the thermosensitive yip1-4 strain. We have termed this gene Yip One Suppressor 1 (YOS1). Yos1p is essential for growth and for function of the secretory pathway; depletion or inactivation of Yos1p blocks transport between the ER and the Golgi complex. YOS1 encodes an integral membrane protein of 87 amino acids that is conserved in eukaryotes. Yos1p localizes to ER and Golgi membranes and is efficiently packaged into ER-derived COPII transport vesicles. Yos1p associates with Yip1p and Yif1p, indicating Yos1p is a novel subunit of the Yip1p–Yif1p complex. PMID:15659647

  19. The Saccharomyces cerevisiae spindle pole body (SPB) component Nbp1p is required for SPB membrane insertion and interacts with the integral membrane proteins Ndc1p and Mps2p.

    PubMed

    Araki, Yasuhiro; Lau, Corine K; Maekawa, Hiromi; Jaspersen, Sue L; Giddings, Thomas H; Schiebel, Elmar; Winey, Mark

    2006-04-01

    The spindle pole body (SPB) in Saccharomyces cerevisiae functions to nucleate and organize spindle microtubules, and it is embedded in the nuclear envelope throughout the yeast life cycle. However, the mechanism of membrane insertion of the SPB has not been elucidated. Ndc1p is an integral membrane protein that localizes to SPBs, and it is required for insertion of the SPB into the nuclear envelope during SPB duplication. To better understand the function of Ndc1p, we performed a dosage suppressor screen using the ndc1-39 temperature-sensitive allele. We identified an essential SPB component, Nbp1p. NBP1 shows genetic interactions with several SPB genes in addition to NDC1, and two-hybrid analysis revealed that Nbp1p binds to Ndc1p. Furthermore, Nbp1p is in the Mps2p-Bbp1p complex in the SPB. Immunoelectron microscopy confirmed that Nbp1p localizes to the SPB, suggesting a function at this location. Consistent with this hypothesis, nbp1-td (a degron allele) cells fail in SPB duplication upon depletion of Nbp1p. Importantly, these cells exhibit a "dead" SPB phenotype, similar to cells mutant in MPS2, NDC1, or BBP1. These results demonstrate that Nbp1p is a SPB component that acts in SPB duplication at the point of SPB insertion into the nuclear envelope.

  20. Differential S1P Receptor Profiles on M1- and M2-Polarized Macrophages Affect Macrophage Cytokine Production and Migration

    PubMed Central

    Müller, Jan; von Bernstorff, Wolfram; Heidecke, Claus-Dieter

    2017-01-01

    Introduction. Macrophages are key players in complex biological processes. In response to environmental signals, macrophages undergo polarization towards a proinflammatory (M1) or anti-inflammatory (M2) phenotype. Sphingosine 1-phosphate (S1P) is a bioactive lysophospholipid that acts via 5 G-protein coupled receptors (S1P1–5) in order to influence a broad spectrum of biological processes. This study assesses S1P receptor expression on macrophages before and after M1 and M2 polarization and performs a comparative analysis of S1P signalling in the two activational states of macrophages. Methods. Bone marrow derived macrophages (BMDM) from C57 BL/6 mice were cultured under either M1- or M2-polarizing conditions. S1P-receptor expression was determined by quantitative RT-PCR. Influence of S1P on macrophage activation, migration, phagocytosis, and cytokine secretion was assessed in vitro. Results. All 5 S1P receptor subclasses were expressed in macrophages. Culture under both M1- and M2-polarizing conditions led to significant downregulation of S1P1. In contrast, M1-polarized macrophages significantly downregulated S1P4. The expression of the remaining three S1P receptors did not change. S1P increased expression of iNOS under M2-polarizing conditions. Furthermore, S1P induced chemotaxis in M1 macrophages and changed cytokine production in M2 macrophages. Phagocytosis was not affected by S1P-signalling. Discussion. The expression of different specific S1P receptor profiles may provide a possibility to selectively influence M1- or M2-polarized macrophages. PMID:28367448

  1. Assay to measure the secretion of sphingosine-1-phosphate from cells induced by S1P lyase inhibitors.

    PubMed

    Loetscher, Erika; Schneider, Karolina; Beerli, Christian; Billich, Andreas

    2013-04-12

    Inhibitors of the sphingosine-1-phosphate (S1P) degrading enzyme S1P lyase (SPL) may be useful in the therapy of inflammatory diseases by preventing lymphocyte recruitment to diseased tissues. Here we describe a cellular assay for such inhibitors, which takes advantage of the observation that a fraction of the intracellular S1P accumulated in the presence of SPL inhibitors is secreted into the medium of cultured cells. The secreted S1P is then quantified using an S1P-sensitive reporter cell line. In the routine assay protocol, human HEK293T cells are treated with SPL inhibitors in the presence of phosphatase inhibitors and sphingosine; while the phosphatase inhibitors are included to prevent the degradation of S1P secreted from the cells, sphingosine is added as source for intracellular S1P that is prone to SPL degradation. The secreted S1P in the supernatant of the cell cultures is then quantified by measuring calcium flux induced in CHO-K1 cells expressing the human S1P3 receptor. Using this method SPL inhibitors were shown to induce a concentration-dependent increase of extracellular S1P under the conditions used; thus, the assay allows for the ranking of SPL inhibitors according to their potency on living cells.

  2. MMP2 and MMP9 participate in S1P-induced invasion of follicular ML-1 thyroid cancer cells.

    PubMed

    Kalhori, Veronica; Törnquist, Kid

    2015-03-15

    The bioactive lipid sphingosine-1-phosphate (S1P) has emerged as a potent inducer of cancer cell migration and invasion. Previously, we have shown that S1P induces invasion of ML-1 follicular thyroid cancer cells via S1P receptors 1 and 3 (S1P1,3). Matrix metalloproteinases (MMPs) are zinc-dependent proteolytic enzymes used by cells for degradation of the extracellular matrix during invasion and migration. In the present study, we examined the role of MMP2 and MMP9 for S1P-induced invasion of ML-1 cells, and found that S1P regulates the secretion and activity of MMP2 and MMP9 via S1P1,3. Both pharmacological inhibitors and siRNA knockdown of MMP2 and MMP9 could attenuate S1P-induced invasion. Additionally, we show that calpains and Rac1 mediate S1P-induced secretion of MMP2 and MMP9. In conclusion, MMP2 and MMP9 participate in S1P-evoked follicular ML-1 thyroid cancer cell invasion.

  3. Bitopic Sphingosine 1-Phosphate Receptor 3 (S1P3) Antagonist Rescue from Complete Heart Block: Pharmacological and Genetic Evidence for Direct S1P3 Regulation of Mouse Cardiac Conduction

    PubMed Central

    Sanna, M. Germana; Vincent, Kevin P.; Repetto, Emanuela; Nguyen, Nhan; Brown, Steven J.; Abgaryan, Lusine; Riley, Sean W.; Leaf, Nora B.; Cahalan, Stuart M.; Kiosses, William B.; Kohno, Yasushi; Brown, Joan Heller; McCulloch, Andrew D.

    2016-01-01

    The molecular pharmacology of the G protein–coupled receptors for sphingosine 1-phosphate (S1P) provides important insight into established and new therapeutic targets. A new, potent bitopic S1P3 antagonist, SPM-354, with in vivo activity, has been used, together with S1P3-knockin and S1P3-knockout mice to define the spatial and functional properties of S1P3 in regulating cardiac conduction. We show that S1P3 is a key direct regulator of cardiac rhythm both in vivo and in isolated perfused hearts. 2-Amino-2-[2-(4-octylphenyl)ethyl]propane-1,3-diol in vivo and S1P in isolated hearts induced a spectrum of cardiac effects, ranging from sinus bradycardia to complete heart block, as measured by a surface electrocardiogram in anesthetized mice and in volume-conducted Langendorff preparations. The agonist effects on complete heart block are absent in S1P3-knockout mice and are reversed in wild-type mice with SPM-354, as characterized and described here. Homologous knockin of S1P3-mCherry is fully functional pharmacologically and is strongly expressed by immunohistochemistry confocal microscopy in Hyperpolarization Activated Cyclic Nucleotide Gated Potassium Channel 4 (HCN4)-positive atrioventricular node and His-Purkinje fibers, with relative less expression in the HCN4-positive sinoatrial node. In Langendorff studies, at constant pressure, SPM-354 restored sinus rhythm in S1P-induced complete heart block and fully reversed S1P-mediated bradycardia. S1P3 distribution and function in the mouse ventricular cardiac conduction system suggest a direct mechanism for heart block risk that should be further studied in humans. A richer understanding of receptor and ligand usage in the pacemaker cells of the cardiac system is likely to be useful in understanding ventricular conduction in health, disease, and pharmacology. PMID:26494861

  4. Fine-Mapping of the 1p11.2 Breast Cancer Susceptibility Locus.

    PubMed

    Horne, Hisani N; Chung, Charles C; Zhang, Han; Yu, Kai; Prokunina-Olsson, Ludmila; Michailidou, Kyriaki; Bolla, Manjeet K; Wang, Qin; Dennis, Joe; Hopper, John L; Southey, Melissa C; Schmidt, Marjanka K; Broeks, Annegien; Muir, Kenneth; Lophatananon, Artitaya; Fasching, Peter A; Beckmann, Matthias W; Fletcher, Olivia; Johnson, Nichola; Sawyer, Elinor J; Tomlinson, Ian; Burwinkel, Barbara; Marme, Frederik; Guénel, Pascal; Truong, Thérèse; Bojesen, Stig E; Flyger, Henrik; Benitez, Javier; González-Neira, Anna; Anton-Culver, Hoda; Neuhausen, Susan L; Brenner, Hermann; Arndt, Volker; Meindl, Alfons; Schmutzler, Rita K; Brauch, Hiltrud; Hamann, Ute; Nevanlinna, Heli; Khan, Sofia; Matsuo, Keitaro; Iwata, Hiroji; Dörk, Thilo; Bogdanova, Natalia V; Lindblom, Annika; Margolin, Sara; Mannermaa, Arto; Kosma, Veli-Matti; Chenevix-Trench, Georgia; Wu, Anna H; Ven den Berg, David; Smeets, Ann; Zhao, Hui; Chang-Claude, Jenny; Rudolph, Anja; Radice, Paolo; Barile, Monica; Couch, Fergus J; Vachon, Celine; Giles, Graham G; Milne, Roger L; Haiman, Christopher A; Marchand, Loic Le; Goldberg, Mark S; Teo, Soo H; Taib, Nur A M; Kristensen, Vessela; Borresen-Dale, Anne-Lise; Zheng, Wei; Shrubsole, Martha; Winqvist, Robert; Jukkola-Vuorinen, Arja; Andrulis, Irene L; Knight, Julia A; Devilee, Peter; Seynaeve, Caroline; García-Closas, Montserrat; Czene, Kamila; Darabi, Hatef; Hollestelle, Antoinette; Martens, John W M; Li, Jingmei; Lu, Wei; Shu, Xiao-Ou; Cox, Angela; Cross, Simon S; Blot, William; Cai, Qiuyin; Shah, Mitul; Luccarini, Craig; Baynes, Caroline; Harrington, Patricia; Kang, Daehee; Choi, Ji-Yeob; Hartman, Mikael; Chia, Kee Seng; Kabisch, Maria; Torres, Diana; Jakubowska, Anna; Lubinski, Jan; Sangrajrang, Suleeporn; Brennan, Paul; Slager, Susan; Yannoukakos, Drakoulis; Shen, Chen-Yang; Hou, Ming-Feng; Swerdlow, Anthony; Orr, Nick; Simard, Jacques; Hall, Per; Pharoah, Paul D P; Easton, Douglas F; Chanock, Stephen J; Dunning, Alison M; Figueroa, Jonine D

    2016-01-01

    The Cancer Genetic Markers of Susceptibility genome-wide association study (GWAS) originally identified a single nucleotide polymorphism (SNP) rs11249433 at 1p11.2 associated with breast cancer risk. To fine-map this locus, we genotyped 92 SNPs in a 900kb region (120,505,799-121,481,132) flanking rs11249433 in 45,276 breast cancer cases and 48,998 controls of European, Asian and African ancestry from 50 studies in the Breast Cancer Association Consortium. Genotyping was done using iCOGS, a custom-built array. Due to the complicated nature of the region on chr1p11.2: 120,300,000-120,505,798, that lies near the centromere and contains seven duplicated genomic segments, we restricted analyses to 429 SNPs excluding the duplicated regions (42 genotyped and 387 imputed). Per-allelic associations with breast cancer risk were estimated using logistic regression models adjusting for study and ancestry-specific principal components. The strongest association observed was with the original identified index SNP rs11249433 (minor allele frequency (MAF) 0.402; per-allele odds ratio (OR) = 1.10, 95% confidence interval (CI) 1.08-1.13, P = 1.49 x 10-21). The association for rs11249433 was limited to ER-positive breast cancers (test for heterogeneity P≤8.41 x 10-5). Additional analyses by other tumor characteristics showed stronger associations with moderately/well differentiated tumors and tumors of lobular histology. Although no significant eQTL associations were observed, in silico analyses showed that rs11249433 was located in a region that is likely a weak enhancer/promoter. Fine-mapping analysis of the 1p11.2 breast cancer susceptibility locus confirms this region to be limited to risk to cancers that are ER-positive.

  5. Fine-Mapping of the 1p11.2 Breast Cancer Susceptibility Locus

    PubMed Central

    Horne, Hisani N.; Chung, Charles C.; Zhang, Han; Yu, Kai; Prokunina-Olsson, Ludmila; Michailidou, Kyriaki; Bolla, Manjeet K.; Wang, Qin; Dennis, Joe; Hopper, John L.; Southey, Melissa C.; Schmidt, Marjanka K.; Broeks, Annegien; Muir, Kenneth; Lophatananon, Artitaya; Fasching, Peter A.; Beckmann, Matthias W.; Fletcher, Olivia; Johnson, Nichola; Sawyer, Elinor J.; Tomlinson, Ian; Burwinkel, Barbara; Marme, Frederik; Guénel, Pascal; Truong, Thérèse; Bojesen, Stig E.; Flyger, Henrik; Benitez, Javier; González-Neira, Anna; Anton-Culver, Hoda; Neuhausen, Susan L.; Brenner, Hermann; Arndt, Volker; Meindl, Alfons; Schmutzler, Rita K.; Brauch, Hiltrud; Hamann, Ute; Nevanlinna, Heli; Khan, Sofia; Matsuo, Keitaro; Iwata, Hiroji; Dörk, Thilo; Bogdanova, Natalia V.; Lindblom, Annika; Margolin, Sara; Mannermaa, Arto; Kosma, Veli-Matti; Chenevix-Trench, Georgia; Wu, Anna H.; ven den Berg, David; Smeets, Ann; Zhao, Hui; Chang-Claude, Jenny; Rudolph, Anja; Radice, Paolo; Barile, Monica; Couch, Fergus J.; Vachon, Celine; Giles, Graham G.; Milne, Roger L.; Haiman, Christopher A.; Marchand, Loic Le; Goldberg, Mark S.; Teo, Soo H.; Taib, Nur A. M.; Kristensen, Vessela; Borresen-Dale, Anne-Lise; Zheng, Wei; Shrubsole, Martha; Winqvist, Robert; Jukkola-Vuorinen, Arja; Andrulis, Irene L.; Knight, Julia A.; Devilee, Peter; Seynaeve, Caroline; García-Closas, Montserrat; Czene, Kamila; Darabi, Hatef; Hollestelle, Antoinette; Martens, John W. M.; Li, Jingmei; Lu, Wei; Shu, Xiao-Ou; Cox, Angela; Cross, Simon S.; Blot, William; Cai, Qiuyin; Shah, Mitul; Luccarini, Craig; Baynes, Caroline; Harrington, Patricia; Kang, Daehee; Choi, Ji-Yeob; Hartman, Mikael; Chia, Kee Seng; Kabisch, Maria; Torres, Diana; Jakubowska, Anna; Lubinski, Jan; Sangrajrang, Suleeporn; Brennan, Paul; Slager, Susan; Yannoukakos, Drakoulis; Shen, Chen-Yang; Hou, Ming-Feng; Swerdlow, Anthony; Orr, Nick; Simard, Jacques; Hall, Per; Pharoah, Paul D. P.

    2016-01-01

    The Cancer Genetic Markers of Susceptibility genome-wide association study (GWAS) originally identified a single nucleotide polymorphism (SNP) rs11249433 at 1p11.2 associated with breast cancer risk. To fine-map this locus, we genotyped 92 SNPs in a 900kb region (120,505,799–121,481,132) flanking rs11249433 in 45,276 breast cancer cases and 48,998 controls of European, Asian and African ancestry from 50 studies in the Breast Cancer Association Consortium. Genotyping was done using iCOGS, a custom-built array. Due to the complicated nature of the region on chr1p11.2: 120,300,000–120,505,798, that lies near the centromere and contains seven duplicated genomic segments, we restricted analyses to 429 SNPs excluding the duplicated regions (42 genotyped and 387 imputed). Per-allelic associations with breast cancer risk were estimated using logistic regression models adjusting for study and ancestry-specific principal components. The strongest association observed was with the original identified index SNP rs11249433 (minor allele frequency (MAF) 0.402; per-allele odds ratio (OR) = 1.10, 95% confidence interval (CI) 1.08–1.13, P = 1.49 x 10-21). The association for rs11249433 was limited to ER-positive breast cancers (test for heterogeneity P≤8.41 x 10-5). Additional analyses by other tumor characteristics showed stronger associations with moderately/well differentiated tumors and tumors of lobular histology. Although no significant eQTL associations were observed, in silico analyses showed that rs11249433 was located in a region that is likely a weak enhancer/promoter. Fine-mapping analysis of the 1p11.2 breast cancer susceptibility locus confirms this region to be limited to risk to cancers that are ER-positive. PMID:27556229

  6. Candida albicans virulence and drug-resistance requires the O-acyltransferase Gup1p

    PubMed Central

    2010-01-01

    Background GUP1 gene was primarily identified in Saccharomyces cerevisiae being connected with glycerol uptake defects in association with osmotic stress response. Soon after, Gup1p was implicated in a complex and extensive series of phenotypes involving major cellular processes. These include membrane and wall maintenance, lipid composition, bud-site selection, cytoskeleton orientation, vacuole morphology, secretory/endocytic pathway, GPI anchors remodelling, and lipid-ordered domains assembly, which is compatible with their inclusion in the Membrane Bound O-acyl transferases (MBOAT) family. In mammals, it has been described as a negative regulator of the Sonic hedgehog pathway involved in morphogenesis, differentiation, proliferation, among other processes. Results We show that Candida albicans Gup1p strongly interferes with the capacity of cells to develop hyphae, to adhere, to invade, and to form a biofilm, all of which are significant virulence factors. Furthermore, the mutant colonies exhibited an aberrant morphology/differentiation pattern. Identically to S. cerevisiae, Cagup1Δ null mutant was more resistant to antifungals like fluconazole, ketoconazole, and clotrimazole, and displayed an abnormal even sterol distribution at the plasma membrane. Conclusions This work is the first study in the opportunistic yeast Candida albicans, showing a role for the GUP1 gene in virulence as well as in the mechanisms underlying antifungal resistance. Moreover, its impact is even more significant since these results, taken together with all the knowledge about GUP1 gene (from S. cerevisiae and mammals) give consistence to the possibility that Gup1p may be part of a yeast morphogenic pathway parallel to the mammalian Hedgehog. PMID:20843317

  7. Ezrin and BCAR1/p130Cas mediate breast cancer growth as 3-D spheroids.

    PubMed

    Konstantinovsky, Sophya; Davidson, Ben; Reich, Reuven

    2012-08-01

    CAS proteins and Ezrin, Radixin, Moesin (ERM) family members act as intracellular scaffolds and are involved in interactions with the cytoskeleton, respectively. Both protein families have previously been associated with metastasis and poor prognosis in cancer. Our group recently reported on the overexpression of EZR/VIL2 and BCAR1 and their protein products in breast carcinoma effusions compared to primary breast carcinoma. In the present study, the role of these two proteins was studied in semi-normal MCF10A cells and metastatic MDA-MB-231 breast carcinoma cells cultured in tri-dimensional (3-D) conditions that were hypothesized to reproduce the in vivo conditions of breast cancer metastasis. MCF10A cells formed spheroid-shaped colonies without any Matrigel invasion, while MDA-MB-231 cells displayed an invasive phenotype and showed satellite projections that bridged multiple cell colonies in 3-D culture. E-cadherin was expressed in MCF10A, but not in MDA-MB-231 cells. The temporal expression of ezrin and BCAR1/p130Cas at the mRNA and protein level differed in the two cell lines upon 3-D culturing on Matrigel. Upregulation of BCAR1/p130cas was observed in the transition of MDA-MB-231 from attached to detached culture. Silencing of Ezrin and p130Cas in MDA-MB-231 cells by short hairpin RNA resulted in decreased invasive potential, and p130Cas silencing further resulted in smaller spheroid/colony formation. Our data show that MCF10A and MDA-MB-231 cells differ in their ability to form spheroids, in expression of E-cadherin and in the expression of Ezrin and BCAR1/p130Cas in 3-D cultures on Matrigel, suggesting a role in tumor progression in breast carcinoma.

  8. Genomewide Linkage Analysis of Obsessive Compulsive Disorder Implicates Chromosome 1p36

    PubMed Central

    Mathews, Carol A.; Badner, Judith A.; Andresen, J. Michael; Sheppard, Brooke; Himle, Joseph A.; Grant, Jon E.; Williams, Kyle A; Chavira, Denise A.; Azzam, Amin; Schwartz, Maxine; Reus, Victor I.; Kim, Suck Won; Cook, Edwin H.; Hanna, Gregory L.

    2012-01-01

    Background Obsessive compulsive disorder (OCD) has a complex etiology involving both genetic and environmental factors. However, the genetic causes of OCD are largely unknown, despite the identification of several promising candidate genes and linkage regions. Methods Our objective was to conduct genetic linkage studies of the type of OCD thought to have the strongest genetic etiology (i.e., childhood-onset OCD), in 33 Caucasian families with ≥2 childhood-onset OCD-affected individuals from the United States (US) (N=245 individuals with genotype data). Parametric and non-parametric genome-wide linkage analyses were conducted with Morgan and Merlin in these families using a selected panel of single nucleotide repeat polymorphisms (SNPs) from the Illumina 610-Quad Bead Chip. The initial analyses were followed by fine-mapping analyses in genomic regions with initial heterogeneity LOD (HLOD) scores of ≥2.0. Results We identified five areas of interest (HLOD score ≥2) on chromosomes 1p36, 2p14, 5q13, 6p25, and 10p13. The strongest result was on chromosome 1p36.33-p36.32 (HLOD=3.77, suggestive evidence for linkage after fine-mapping). At this location, several of the families showed haplotypes co-segregating with OCD. Conclusions The results of this study represent the strongest linkage finding for OCD in a primary analysis to date, and suggest that chromosome 1p36, and possibly several other genomic regions, may harbor susceptibility loci for OCD. Multiple brain-expressed genes lie under the primary linkage peak (approximately 4 mb in size). Follow-up studies, including replication in additional samples and targeted sequencing of the areas of interest, are needed to confirm these findings and to identify specific OCD risk variants. PMID:22633946

  9. HIV-1 p6-Another viral interaction partner to the host cellular protein cyclophilin A.

    PubMed

    Solbak, Sara M Ø; Reksten, Tove R; Röder, Rene; Wray, Victor; Horvli, Ole; Raae, Arnt J; Henklein, Petra; Henklein, Peter; Fossen, Torgils

    2012-04-01

    The 52-amino acid human immunodeficiency virus type 1 (HIV-1) p6 protein has previously been recognized as a docking site for several cellular and viral binding factors and is important for the formation of infectious viruses. A particular structural feature of p6 is the notably high relative content of proline residues, located at positions 5, 7, 10, 11, 24, 30, 37 and 49 in the sequence. Proline cis/trans isomerism was detected for all these proline residues to such an extent that more than 40% of all p6 molecules contain at least one proline in a cis conformation. 2D (1)H nuclear magnetic resonance analysis of full-length HIV-1 p6 and p6 peptides established that cyclophilin A (CypA) interacts as a peptidyl-prolyl cis/trans isomerase with all proline residues of p6. Only catalytic amounts of CypA were necessary for the interaction with p6 to occur, strongly suggesting that the observed interaction is highly relevant in vivo. In addition, surface plasmon resonance studies revealed binding of full-length p6 to CypA, and that this binding was significantly stronger than any of its N- or C-terminal peptides. This study demonstrates the first identification of an interaction between HIV-1 p6 and the host cellular protein CypA. The mode of interaction involves both transient enzyme-substrate interactions and a more stable binding. The binding motifs of p6 to Tsg-101, ALIX and Vpr coincide with binding regions and catalytic sites of p6 to CypA, suggesting a potential role of CypA in modulating functional interactions of HIV-1.

  10. Late-onset Stargardt-like macular dystrophy maps to chromosome 1p13

    SciTech Connect

    Kaplan, J.; Gerber, S.; Rozet, J.M.

    1994-09-01

    Stargardt`s disease (MIM 248200), originally described in 1909, is an autosomal recessive condition of childhood, characterized by a sudden and bilateral loss of central vision. Typically, it has an early onset (7 to 12 years), a rapidly progressive course and a poor final outcome. The central area of the retina (macula) displays pigmentary changes in a ring form with depigmentation and atrophy of the retinal pigmentary epithelium (RPE). Perimacular yellowish spots, termed fundus flavimaculatus, are observed in a high percentage of patients. We have recently reported the genetic mapping of Stargardt`s disease to chromosome 1p13. On the other hand, considering that fundus flavimaculatus (MIM 230100) is another form of fleck fundus disease, with a Stargardt-like retinal aspect but with a late-onset and a more progressive course, we decided to test the hypothesis of allelism between typical Stargardt`s disease and late-onset autosomal recessive fundus flavimaculatus. Significant pairwise lod scores were obtained in each of four multiplex families (11 affected individuals, 12 relatives) with four markers of the 1p13 region (Z = 4.79, 4.64, 3.07, 3.16 at loci D1S435, D1S424, D1S236, and D1S415, respectively at {theta} = 0). Multipoint analysis showed that the best estimate for location of the disease gene is between D1S424 and D1S236 (maximum lod score of 5.20) as also observed in Stargardt`s disease. Our results are consistent with the location of the gene responsible of the late-onset Stargardt-like macular dystrophy in the 1p13 region and raise the hypothesis of either allelic mutational events or contiguous genes in this chromosomal region. The question of possible relationship with some age-related macular dystrophies in now open to debate.

  11. Dying at 23 with 1p36 deletion syndrome: Laura's family story.

    PubMed

    Tandy, P A

    2012-09-01

    Laura was unusual. She had always been different and at times difficult. She was born with a genetic disorder, diagnosed as 1p36 deletion syndrome when she was 21 years old. At 23 she suffered her first cardiac arrest at home and entered the hospital system for the first time apart from infancy. After initially appearing to do well, she suffered a second cardiac arrest 10 weeks after admission. This was followed by an irreversible deterioration and she died 14 weeks after admission. We her family had been with her throughout her traumatic experience. This is our story.

  12. {alpha}-cluster and {alpha}-binucleon states of 1p-shell nuclei

    SciTech Connect

    Wolsky, R.; Gnilozub, I. A.; Kurgalin, S. D.; Tchuvil'sky, Yu. M.

    2010-08-15

    States that exhibit the properties of an {alpha}-cluster or an {alpha}-binucleon condensate are studied in 1p-shell nuclei. The generalized Hamiltonian of the Elliott SU(3) model is used to classify these states and to calculate their spectra. The results of the calculations are found to be in good agreement with experimental data. States not observed so far in the cluster spectra of the {sup 12}C, {sup 16}O, {sup 10}Be, and {sup 12}Be nuclei are predicted.

  13. The appearance of the feet in Pfeiffer syndrome caused by FGFR1 P252R mutation.

    PubMed

    Rossi, Massimiliano; Jones, Rachel L; Norbury, Gail; Bloch-Zupan, Agnès; Winter, Robin M

    2003-10-01

    Patients affected by Pfeiffer syndrome generally present with syndromic craniosynostosis and typical limb defects including broad thumbs, wide halluces with varus deformity, toe syndactyly and sometimes elbow ankylosis. This autosomal dominant condition can be caused by mutations in either fibroblast growth factor receptor gene type 1 or 2 (FGFR1 or FGFR2). We report four new affected families showing an FGFR1 P252R mutation and emphasize the characteristic malformations of the feet in this form of Pfeiffer syndrome. In one family this was the only abnormality.

  14. A convenient synthesis of benzo[c]naphtho[2,1-p]chrysene

    SciTech Connect

    Hagen, S.; Scott, L.T.

    1996-10-04

    The polycyclic aromatic hydrocarbon (PAH) benzo[c]-naphtho[2,1-p]chrysene (1) has recently attracted renewed attention as a potential precursor for the synthesis of bowl-shaped fullerene substructures. The published synthetic approaches to 1, however, are lengthy and entail one or more photocyclizations of stilbene-type compounds that suffer from competing [2 + 2]cycloaddition reactions at normal concentrations and are thereby rendered quite inefficient. The authors report here a convenient four-step synthesis of 1 that can be performed on a multigram scale starting from the commercially available {alpha}-tetralone (2) and 2-bromonaphthalene. 12 refs.

  15. RAP1GA1: A candidate tumor suppressor locus in 1p36.1

    SciTech Connect

    Ranade, K.; Hussussian, C.J.; Higgins, P.

    1994-09-01

    The rap1/Krev-1 gene (RAP1A) encodes a p21-related protein that suppresses transformation by activated p21{sup ras}. The GTPase activating protein (GAP) gene for p21{sup rap1A} (RAP1GA1) has recently been assigned to chromosome 1p36.1-p35, a region of the genome that is frequently involved in deletions and rearrangements in several different tumors including breast, colon and hepatocellular carcinomas, melanoma, and neuroblastoma. GAP genes negatively regulate the activity of p21 proteins by catalyzing the conversion of the active GTP-bound forms to the inactive GDP-bound forms. The physiological function of p21{sup rap1A}-GAP makes it a strong candidate as a tumor suppressor gene that may have a role in the development of one or more of these malignancies. We have refined the localization of RAP1GA1 by linkage analysis with a highly informative (CA){sub n} repeat contained within the gene, and demonstrated that it is within the minimal deleted region for breast and colon carcinomas, and that it is excluded from the minimally deleted region in melanoma and neuroblastoma. Genetic mapping in the mouse demonstrated that Rap1ga1 is located {approximately}10 cM proximal to Pnd and therefore maps within the interval containing the modifier of Min gene (Mom-1) and the plasmocytoma susceptibility locus (Pcts). The human RAP1GA1 gene contains at least 27 exons. The coding region contains 22 exons, and there are at least five 5{prime}-UT exons that are assembled in a complex pattern of alternative splicing in different tissues. The localization of RAP1GA1 makes it a very strong candidate for a role as a modifier gene involved in the common secondary abnormalities involving 1p36 in several different carcinomas. The potential role of RAP1GA1 in these malignancies is currently being investigated by sequence analysis of breast and colon carcinomas with loss of heterozygosity in 1p36.

  16. HMGA1P7-pseudogene regulates H19 and Igf2 expression by a competitive endogenous RNA mechanism

    PubMed Central

    De Martino, Marco; Forzati, Floriana; Marfella, Marianna; Pellecchia, Simona; Arra, Claudio; Terracciano, Luigi; Fusco, Alfredo; Esposito, Francesco

    2016-01-01

    Recent studies have revealed that pseudogene transcripts can function as competing endogenous RNAs, and thereby can also contribute to cancer when dysregulated. We have recently identified two pseudogenes, HMGA1P6 and HMGA1P7 for the HMGA1 gene whose overexpression has a critical role in cancer progression. These pseudogenes work as competitive endogenous RNA decoys for HMGA1 and other cancer related genes suggesting their role in carcinogenesis. Looking for new HMGA1 pseudogene ceRNAs, we performed RNA sequencing technology on mouse embryonic fibroblasts deriving from transgenic mice overexpressing HMGA1P7. Here, we report that HMGA1P7 mRNA sustains the H19 and Igf2 overexpression by acting as miRNA decoy. Lastly, the expression of HMGA1P7 was significantly correlated with H19 and IGF2 levels in human breast cancer thereby suggesting a role for HMGA1P7 deregulation in this neoplasia. PMID:27874091

  17. Precision measurement of the mass of the hc(1P1) state of charmonium.

    PubMed

    Dobbs, S; Metreveli, Z; Seth, K K; Tomaradze, A; Libby, J; Powell, A; Wilkinson, G; Ecklund, K M; Love, W; Savinov, V; Lopez, A; Mendez, H; Ramirez, J; Ge, J Y; Miller, D H; Shipsey, I P J; Xin, B; Adams, G S; Anderson, M; Cummings, J P; Danko, I; Hu, D; Moziak, B; Napolitano, J; He, Q; Insler, J; Muramatsu, H; Park, C S; Thorndike, E H; Yang, F; Artuso, M; Blusk, S; Khalil, S; Li, J; Mountain, R; Nisar, S; Randrianarivony, K; Sultana, N; Skwarnicki, T; Stone, S; Wang, J C; Zhang, L M; Bonvicini, G; Cinabro, D; Dubrovin, M; Lincoln, A; Naik, P; Rademacker, J; Asner, D M; Edwards, K W; Reed, J; Briere, R A; Ferguson, T; Tatishvili, G; Vogel, H; Watkins, M E; Rosner, J L; Alexander, J P; Cassel, D G; Duboscq, J E; Ehrlich, R; Fields, L; Galik, R S; Gibbons, L; Gray, R; Gray, S W; Hartill, D L; Heltsley, B K; Hertz, D; Hunt, J M; Kandaswamy, J; Kreinick, D L; Kuznetsov, V E; Ledoux, J; Mahlke-Krüger, H; Mohapatra, D; Onyisi, P U E; Patterson, J R; Peterson, D; Riley, D; Ryd, A; Sadoff, A J; Shi, X; Stroiney, S; Sun, W M; Wilksen, T; Athar, S B; Patel, R; Yelton, J; Rubin, P; Eisenstein, B I; Karliner, I; Mehrabyan, S; Lowrey, N; Selen, M; White, E J; Wiss, J; Mitchell, R E; Shepherd, M R; Besson, D; Pedlar, T K; Cronin-Hennessy, D; Gao, K Y; Hietala, J; Kubota, Y; Klein, T; Lang, B W; Poling, R; Scott, A W; Zweber, P

    2008-10-31

    A precision measurement of the mass of the h_{c}(1P1) state of charmonium has been made using a sample of 24.5x10;{6} psi(2S) events produced in e;{+}e;{-} annihilation at the Cornell Electron Storage Ring (CESR). The reaction used was psi(2S)-->pi;{0}h_{c}, pi;{0}-->gammagamma, h_{c}-->gammaeta_{c}, and the reaction products were detected in the CLEO-c detector. Data have been analyzed both for the inclusive reaction and for the exclusive reactions in which eta_{c} decays are reconstructed in 15 hadronic decay channels. Consistent results are obtained in the two analyses. The averaged results of the present measurements are M(h_{c})=3525.28+/-0.19(stat.)+/-0.12(syst.) MeV, and B(psi(2S)-->pi;{0}h_{c})xB(h_{c}-->gammaeta_{c})=(4.19+/-0.32+/-0.45)x10;{-4}. Using the ;{3}P_{J} centroid mass, DeltaM_{hf}(1P) identical withM(chi_{cJ})-M(h_{c})=+0.02+/-0.19+/-0.13 MeV.

  18. Targeted Proteomics-Driven Computational Modeling of Macrophage S1P Chemosensing.

    PubMed

    Manes, Nathan P; Angermann, Bastian R; Koppenol-Raab, Marijke; An, Eunkyung; Sjoelund, Virginie H; Sun, Jing; Ishii, Masaru; Germain, Ronald N; Meier-Schellersheim, Martin; Nita-Lazar, Aleksandra

    2015-10-01

    Osteoclasts are monocyte-derived multinuclear cells that directly attach to and resorb bone. Sphingosine-1-phosphate (S1P)(1) regulates bone resorption by functioning as both a chemoattractant and chemorepellent of osteoclast precursors through two G-protein coupled receptors that antagonize each other in an S1P-concentration-dependent manner. To quantitatively explore the behavior of this chemosensing pathway, we applied targeted proteomics, transcriptomics, and rule-based pathway modeling using the Simmune toolset. RAW264.7 cells (a mouse monocyte/macrophage cell line) were used as model osteoclast precursors, RNA-seq was used to identify expressed target proteins, and selected reaction monitoring (SRM) mass spectrometry using internal peptide standards was used to perform absolute abundance measurements of pathway proteins. The resulting transcript and protein abundance values were strongly correlated. Measured protein abundance values, used as simulation input parameters, led to in silico pathway behavior matching in vitro measurements. Moreover, once model parameters were established, even simulated responses toward stimuli that were not used for parameterization were consistent with experimental findings. These findings demonstrate the feasibility and value of combining targeted mass spectrometry with pathway modeling for advancing biological insight.

  19. Inhibition of the Formation of the Spf1p Phosphoenzyme by Ca2+*

    PubMed Central

    Corradi, Gerardo R.; Czysezon, Nicolas A.; Mazzitelli, Luciana R.; Sarbia, Nicolas; Adamo, Hugo P.

    2016-01-01

    P5-ATPases are important for processes associated with the endosomal-lysosomal system of eukaryotic cells. In humans, the loss of function of P5-ATPases causes neurodegeneration. In the yeast Saccharomyces cerevisiae, deletion of P5-ATPase Spf1p gives rise to endoplasmic reticulum stress. The reaction cycle of P5-ATPases is poorly characterized. Here, we showed that the formation of the Spf1p catalytic phosphoenzyme was fast in a reaction medium containing ATP, Mg2+, and EGTA. Low concentrations of Ca2+ in the phosphorylation medium decreased the rate of phosphorylation and the maximal level of phosphoenzyme. Neither Mn2+ nor Mg2+ had an inhibitory effect on the formation of the phosphoenzyme similar to that of Ca2+. The Km for ATP in the phosphorylation reaction was ∼1 μm and did not significantly change in the presence of Ca2+. Half-maximal phosphorylation was attained at 8 μm Mg2+, but higher concentrations partially protected from Ca2+ inhibition. In conditions similar to those used for phosphorylation, Ca2+ had a small effect accelerating dephosphorylation and minimally affected ATPase activity, suggesting that the formation of the phosphoenzyme was not the limiting step of the ATP hydrolytic cycle. PMID:26858246

  20. Control of Mitotic Spindle Position by the Saccharomyces cerevisiae Formin Bni1p

    PubMed Central

    Lee, Laifong; Klee, Saskia K.; Evangelista, Marie; Boone, Charles; Pellman, David

    1999-01-01

    Alignment of the mitotic spindle with the axis of cell division is an essential process in Saccharomyces cerevisiae that is mediated by interactions between cytoplasmic microtubules and the cell cortex. We found that a cortical protein, the yeast formin Bni1p, was required for spindle orientation. Two striking abnormalities were observed in bni1Δ cells. First, the initial movement of the spindle pole body (SPB) toward the emerging bud was defective. This phenotype is similar to that previously observed in cells lacking the kinesin Kip3p and, in fact, BNI1 and KIP3 were found to be in the same genetic pathway. Second, abnormal pulling interactions between microtubules and the cortex appeared to cause preanaphase spindles in bni1Δ cells to transit back and forth between the mother and the bud. We therefore propose that Bni1p may localize or alter the function of cortical microtubule-binding sites in the bud. Additionally, we present evidence that other bipolar bud site determinants together with cortical actin are also required for spindle orientation. PMID:10085293

  1. Inhibition of the Formation of the Spf1p Phosphoenzyme by Ca2.

    PubMed

    Corradi, Gerardo R; Czysezon, Nicolas A; Mazzitelli, Luciana R; Sarbia, Nicolas; Adamo, Hugo P

    2016-04-01

    P5-ATPases are important for processes associated with the endosomal-lysosomal system of eukaryotic cells. In humans, the loss of function of P5-ATPases causes neurodegeneration. In the yeastSaccharomyces cerevisiae, deletion of P5-ATPase Spf1p gives rise to endoplasmic reticulum stress. The reaction cycle of P5-ATPases is poorly characterized. Here, we showed that the formation of the Spf1p catalytic phosphoenzyme was fast in a reaction medium containing ATP, Mg(2+), and EGTA. Low concentrations of Ca(2+)in the phosphorylation medium decreased the rate of phosphorylation and the maximal level of phosphoenzyme. Neither Mn(2+)nor Mg(2+)had an inhibitory effect on the formation of the phosphoenzyme similar to that of Ca(2+) TheKmfor ATP in the phosphorylation reaction was ∼1 μmand did not significantly change in the presence of Ca(2+) Half-maximal phosphorylation was attained at 8 μmMg(2+), but higher concentrations partially protected from Ca(2+)inhibition. In conditions similar to those used for phosphorylation, Ca(2+)had a small effect accelerating dephosphorylation and minimally affected ATPase activity, suggesting that the formation of the phosphoenzyme was not the limiting step of the ATP hydrolytic cycle.

  2. Targeted Proteomics-Driven Computational Modeling of Macrophage S1P Chemosensing*

    PubMed Central

    Manes, Nathan P.; Angermann, Bastian R.; Koppenol-Raab, Marijke; An, Eunkyung; Sjoelund, Virginie H.; Sun, Jing; Ishii, Masaru; Germain, Ronald N.; Meier-Schellersheim, Martin; Nita-Lazar, Aleksandra

    2015-01-01

    Osteoclasts are monocyte-derived multinuclear cells that directly attach to and resorb bone. Sphingosine-1-phosphate (S1P)1 regulates bone resorption by functioning as both a chemoattractant and chemorepellent of osteoclast precursors through two G-protein coupled receptors that antagonize each other in an S1P-concentration-dependent manner. To quantitatively explore the behavior of this chemosensing pathway, we applied targeted proteomics, transcriptomics, and rule-based pathway modeling using the Simmune toolset. RAW264.7 cells (a mouse monocyte/macrophage cell line) were used as model osteoclast precursors, RNA-seq was used to identify expressed target proteins, and selected reaction monitoring (SRM) mass spectrometry using internal peptide standards was used to perform absolute abundance measurements of pathway proteins. The resulting transcript and protein abundance values were strongly correlated. Measured protein abundance values, used as simulation input parameters, led to in silico pathway behavior matching in vitro measurements. Moreover, once model parameters were established, even simulated responses toward stimuli that were not used for parameterization were consistent with experimental findings. These findings demonstrate the feasibility and value of combining targeted mass spectrometry with pathway modeling for advancing biological insight. PMID:26199343

  3. Tof1p regulates DNA damage responses during S phase in Saccharomyces cerevisiae.

    PubMed Central

    Foss, E J

    2001-01-01

    A tof1 mutant was recovered in a screen aimed at identifying genes involved specifically in the S phase branch of the MEC1-dependent DNA damage response pathway. The screen was based on the observation that mutants missing this branch are particularly dependent on the cell cycle-wide branch and, therefore, on RAD9, for surviving DNA damage. tof1 and rad9 conferred synergistic sensitivity to MMS, UV, and HU, and the double mutant was incapable of slowing S phase in response to MMS, inducing RNR3 transcription in response to UV, and phosphorylating Rad53p in response to HU. TOF1's contribution to DNA damage response appeared to be restricted to S phase, since TOF1 did not contribute to UV-induced transcription during G1 or to the cdc13-1-induced block to anaphase in G2/M. I suggest a model in which Tof1p functions to link Mec1p with Rad53p. PMID:11156979

  4. Phospholipid transfer protein deficiency decreases the content of S1P in HDL via the loss of its transfer capability.

    PubMed

    Yu, Yang; Guo, Shoudong; Feng, Yumei; Feng, Lei; Cui, Yingjie; Song, Guohua; Luo, Tian; Zhang, Ke; Wang, Yiwei; Jiang, Xian-Cheng; Qin, Shucun

    2014-02-01

    Sphingosine-1-phosphate (S1P) is an amphiphilic signaling molecule, which is enriched in functional high density lipoprotein (HDL) and shows arterial protection. The distribution of S1P is changed with increased plasma phospholipid transfer protein (PLTP) activity and impaired HDL function in patients with coronary heart diseases. Therefore, we hypothesized that PLTP might transfer S1P among cells or lipoproteins. We found that plasma S1P contents were decreased by 60.1 % in PLTP knockout mice (PLTP-/-, N = 5) compared with their wild type littermates (WT, N = 5) (151.70 ± 38.59 vs. 379.32 ± 59.90 nmol/l, P<0.01). S1P content in HDL fraction (HDL-S1P) from PLTP-/- was decreased by 64.7 % compared with WT (49.36 ± 1.49 vs. 139.76 ± 2.94 nmol/l, P<0.01). The results of the S1P transfer assay indicated that PLTP could facilitate S1P transport from erythrocytes to HDL at 37 °C in D-Hanks buffer. Plasma content of apolipoprotein M, a specific adaptor of S1P, was not changed in PLTP-/- compared with WT. Therefore, we concluded that PLTP was a key factor to maintain plasma HDL-S1P, and PLTP deficiency could decrease the S1P content in plasma lipoproteins, which involves its capability of transferring S1P from erythrocyte to HDL.

  5. The Extracellular Domain of the Saccharomyces cerevisiae Sln1p Membrane Osmolarity Sensor Is Necessary for Kinase Activity

    PubMed Central

    Ostrander, Darin B.; Gorman, Jessica A.

    1999-01-01

    The function of the extracellular domain (ECD) of Sln1p, a plasma membrane two-transmembrane domain (TMD) sensor of the high-osmolarity glycerol (HOG) response pathway, has been studied in the yeast Saccharomyces cerevisiae. Truncations of SLN1 that retain an intact kinase domain are capable of complementing the lethality of an sln1Δ strain. By observing levels of Hog1p phosphorylation as well as the phosphorylation state of Sln1p, the kinase activities of various SLN1 constructions were determined. In derivatives that do not contain the first TMD, Sln1p activity was no longer dependent on medium osmolarity but appeared to be constitutively active even under conditions of high osmolarity. Removal of the first TMD (ΔTMD1 construct) gave a protein that was strongly phosphorylated whereas Hog1p was largely dephosphorylated, as expected if the active form of Sln1p is phosphorylated. When both TMDs as well as the ECD were deleted, so that the kinase domain is cytosolic, Sln1p was not phosphorylated whereas Hog1p became constitutively hyperphosphorylated. Surprisingly, this hyperactivity of the HOG mitogen-activated protein kinase signaling pathway was not sufficient to result in cell lethality. When the ECD of the ΔTMD1 construct was replaced with a leucine zipper motif, Sln1p was hyperactive, so that Hog1p became mostly unphosphorylated. In contrast, when the Sln1p/leucine zipper construct was crippled by a mutation of one of the internal leucines, the Sln1 kinase was inactive. These experiments are consistent with the hypothesis that the ECD of Sln1p functions as a dimerization and activation domain but that osmotic regulation of activity requires the presence of the first TMD. PMID:10198019

  6. The Wsc1p Cell Wall Signaling Protein Controls Biofilm (Mat) Formation Independently of Flo11p in Saccharomyces cerevisiae

    PubMed Central

    Sarode, Neha; Davis, Sarah E.; Tams, Robert N.; Reynolds, Todd B.

    2013-01-01

    Saccharomyces cerevisiae strains of the ∑1278b background generate biofilms, referred to as mats, on low-density agar (0.3%) plates made with rich media (YPD). Mat formation involves adhesion of yeast cells to the surface of the agar substrate and each other as the biofilm matures, resulting in elaborate water channels that create filigreed patterns of cells. The cell wall adhesion protein Flo11p is required for mat formation; however, genetic data indicate that other unknown effectors are also required. For example, mutations in vacuolar protein sorting genes that affect the multivesicular body pathway, such as vps27∆, cause mat formation defects independently of Flo11p, presumably by affecting an unidentified signaling pathway. A cell wall signaling protein, Wsc1p, found at the plasma membrane is affected for localization and function by vps27∆. We found that a wsc1∆ mutation disrupted mat formation in a Flo11p-independent manner. Wsc1p appears to impact mat formation through the Rom2p-Rho1p signaling module, by which Wsc1p also regulates the cell wall. The Bck1p, Mkk1/Mkk2, Mpk1p MAP kinase signaling cascade is known to regulate the cell wall downstream of Wsc1p-Rom2p-Rho1p but, surprisingly, these kinases do not affect mat formation. In contrast, Wsc1p may impact mat formation by affecting Skn7p instead. Skn7p can also receive signaling inputs from the Sln1p histidine kinase; however, mutational analysis of specific histidine kinase receiver residues in Skn7p indicate that Sln1p does not play an important role in mat formation, suggesting that Skn7p primarily acts downstream of Wsc1p to regulate mat formation. PMID:24318926

  7. Fus1p interacts with components of the Hog1p mitogen-activated protein kinase and Cdc42p morphogenesis signaling pathways to control cell fusion during yeast mating.

    PubMed Central

    Nelson, Bryce; Parsons, Ainslie B; Evangelista, Marie; Schaefer, Karen; Kennedy, Kathy; Ritchie, Steven; Petryshen, Tracey L; Boone, Charles

    2004-01-01

    Cell fusion in the budding yeast Saccharomyces cerevisiae is a temporally and spatially regulated process that involves degradation of the septum, which is composed of cell wall material, and occurs between conjugating cells within a prezygote, followed by plasma membrane fusion. The plasma membrane protein Fus1p is known to be required for septum degradation during cell fusion, yet its role at the molecular level is not understood. We identified Sho1p, an osmosensor for the HOG MAPK pathway, as a binding partner for Fus1 in a two-hybrid screen. The Sho1p-Fus1p interaction occurs directly and is mediated through the Sho1p-SH3 domain and a proline-rich peptide ligand on the Fus1p COOH-terminal cytoplasmic region. The cell fusion defect associated with fus1Delta mutants is suppressed by a sho1Delta deletion allele, suggesting that Fus1p negatively regulates Sho1p signaling to ensure efficient cell fusion. A two-hybrid matrix containing fusion proteins and pheromone response pathway signaling molecules reveals that Fus1p may participate in a complex network of interactions. In particular, the Fus1p cytoplasmic domain interacts with Chs5p, a protein required for secretion of specialized Chs3p-containing vesicles during bud development, and chs5Delta mutants were defective in cell surface localization of Fus1p. The Fus1p cytoplasmic domain also interacts with the activated GTP-bound form of Cdc42p and the Fus1p-SH3 domain interacts with Bni1p, a yeast formin that participates in cell fusion and controls the assembly of actin cables to polarize secretion in response to Cdc42p signaling. Taken together, our results suggest that Fus1p acts as a scaffold for the assembly of a cell surface complex involved in polarized secretion of septum-degrading enzymes and inhibition of HOG pathway signaling to promote cell fusion. PMID:15020407

  8. Sphingosine-1-phosphate (S1P): A Potential Biomarker and Therapeutic Target for Endothelial Dysfunction and Sepsis?

    PubMed

    Winkler, Martin S; Nierhaus, Axel; Poppe, Annika; Greiwe, Gillis; Gräler, Markus; Daum, Guenter

    2016-12-02

    Sepsis is an acute life-threatening multiple organ failure caused by a dysregulated host response to infection. Endothelial dysfunction, particularly barrier disruption leading to increased vascular permeability, edema and insufficient tissue oxygenation is critical to sepsis pathogenesis. Sphingosine-1-phosphate (S1P) is a signaling lipid that regulates important pathophysiological processes including vascular endothelial cell permeability, inflammation and coagulation. It is present at high concentrations in blood and lymph and at very low concentrations in tissues due to the activity of the S1P-degrading enzyme S1P-lyase in tissue cells. Recently, four preclinical observational studies determined S1P levels in serum or plasma of sepsis patients, and all found reduced S1P levels associated with the disease. Based on these findings, this review summarizes S1P-regulated processes pertaining to endothelial functions, discusses the possible use of S1P as a marker and possibilities how to manipulate S1P levels and S1P receptor activation to restore endothelial integrity, dampen the inflammatory host response, and improve organ function in sepsis.

  9. Cu/Zn superoxide dismutase and the proton ATPase Pma1p of Saccharomyces cerevisiae

    SciTech Connect

    Baron, J. Allen; Chen, Janice S.; Culotta, Valeria C.

    2015-07-03

    In eukaryotes, the Cu/Zn containing superoxide dismutase (SOD1) plays a critical role in oxidative stress protection as well as in signaling. We recently demonstrated a function for Saccharomyces cerevisiae Sod1p in signaling through CK1γ casein kinases and identified the essential proton ATPase Pma1p as one likely target. The connection between Sod1p and Pma1p was explored further by testing the impact of sod1Δ mutations on cells expressing mutant alleles of Pma1p that alter activity and/or post-translational regulation of this ATPase. We report here that sod1Δ mutations are lethal when combined with the T912D allele of Pma1p in the C-terminal regulatory domain. This “synthetic lethality” was reversed by intragenic suppressor mutations in Pma1p, including an A906G substitution that lies within the C-terminal regulatory domain and hyper-activates Pma1p. Surprisingly the effect of sod1Δ mutations on Pma1-T912D is not mediated through the Sod1p signaling pathway involving the CK1γ casein kinases. Rather, Sod1p sustains life of cells expressing Pma1-T912D through oxidative stress protection. The synthetic lethality of sod1Δ Pma1-T912D cells is suppressed by growing cells under low oxygen conditions or by treatments with manganese-based antioxidants. We now propose a model in which Sod1p maximizes Pma1p activity in two ways: one involving signaling through CK1γ casein kinases and an independent role for Sod1p in oxidative stress protection. - Highlights: • In yeast, the anti-oxidant enzyme SOD1 promotes activity of the proton ATPase Pma1p. • Cells expressing a T912D variant of Pma1p are not viable without SOD1. • SOD1 is needed to protect Pma1-T912D expressing cells from severe oxidative damage. • SOD1 activates Pma1p through casein kinase signaling and oxidative stress protection.

  10. FISH analysis of hematological neoplasias with 1p36 rearrangements allows the definition of a cluster of 2.5 Mb included in the minimal region deleted in 1p36 deletion syndrome.

    PubMed

    Lahortiga, Idoya; Vázquez, Iria; Belloni, Elena; Román, José P; Gasparini, Patrizia; Novo, Francisco J; Zudaire, Isabel; Pelicci, Pier G; Hernández, Jesús M; Calasanz, María J; Odero, María D

    2005-05-01

    Rearrangements in the distal region of the short arm of chromosome 1 are recurrent aberrations in a broad spectrum of human neoplasias. However, neither the location of the breakpoints (BP) on 1p36 nor the candidate genes have been fully determined. We have characterized, by fluorescence in situ hybridization (FISH), the BP in 26 patients with hematological neoplasias and 1p36 rearrangements in the G-banding karyotype. FISH allowed a better characterization of all samples analyzed. Nine cases (35%) showed reciprocal translocations, 15 (58%) unbalanced rearrangements, and two (7%) deletions. We describe two new recurrent aberrations. In 18 of the 26 cases analyzed the BP were located in band 1p36, which is 25.5 Mb long. In 14 of these 18 cases (78%) and without distinction between myeloid and lymphoid neoplasias, the BP clustered in a 2.5 Mb region located between 1p36.32 and the telomere. Interestingly, this region is contained in the 10.5 Mb cluster on 1p36.22-1pter defined in cases with 1p36 deletion syndrome. The 2.5 Mb region, located on 1p36.32-1pter, has a higher frequency of occurrence of tandem repeats and segmental duplications larger than 1 kb, when compared with the 25.5 Mb of the complete 1p36 band. This could explain its proneness for involvement in chromosomal rearrangements in hematological neoplasias.

  11. Observation of the {sup 1}P{sub 1} state of charmonium

    SciTech Connect

    Rubin, P.; Cawlfield, C.; Eisenstein, B.I.; Gollin, G.D.; Karliner, I.; Kim, D.; Lowrey, N.; Naik, P.; Sedlack, C.; Selen, M.; White, E. J.; Williams, J.; Wiss, J.; Edwards, K.W.; Besson, D.; Pedlar, T.K.; Cronin-Hennessy, D.; Gao, K.Y.; Gong, D.T.; Hietala, J.

    2005-11-01

    The spin-singlet P-wave state of charmonium, h{sub c}({sup 1}P{sub 1}), has been observed in the decay {psi}(2S){yields}{pi}{sup 0}h{sub c} followed by h{sub c}{yields}{gamma}{eta}{sub c}. Inclusive and exclusive analyses of the M(h{sub c}) spectrum have been performed. Two complementary inclusive analyses select either a range of energies for the photon emitted in h{sub c}{yields}{gamma}{eta}{sub c} or a range of values of M({eta}{sub c}). These analyses, consistent with one another within statistics, yield M(h{sub c})=[3524.9{+-}0.7 (stat){+-}0.4 (sys)] MeV/c{sup 2} and a product of the branching ratios B{sub {psi}}({psi}(2S){yields}{pi}{sup 0}h{sub c})xB{sub h}(h{sub c}{yields}{gamma}{eta}{sub c})=[3.5{+-}1.0 (stat){+-}0.7 (sys)]x10{sup -4}. When the {eta}{sub c} is reconstructed in seven exclusive decay modes, 17.5{+-}4.5 h{sub c} events are seen with an average mass M(h{sub c})=[3523.6{+-}0.9 (stat){+-}0.5 (sys)] MeV/c{sup 2}, and B{sub {psi}}B{sub h}=[5.3{+-}1.5 (stat){+-}1.0 (sys)]x10{sup -4}. If combined, the inclusive and exclusive data samples yield an overall mass M(h{sub c})=[3524.4{+-}0.6 (stat){+-}0.4 (sys)] MeV/c{sup 2} and product of branching ratios B{sub {psi}}B{sub h}=[4.0{+-}0.8 (stat){+-}0.7 (sys)]x10{sup -4}. The h{sub c} mass implies a P-wave hyperfine splitting {delta}M{sub HF}(1P){identical_to}-M(1{sup 1}P{sub 1})=[1.0{+-}0.6 (stat){+-}0.4 (sys)] MeV/c{sup 2}.

  12. Mitochondrial localization of the OAS1 p46 isoform associated with a common single nucleotide polymorphism

    PubMed Central

    2014-01-01

    Background The expression of 2′-5′-Oligoadenylate synthetases (OASs) is induced by type 1 Interferons (IFNs) in response to viral infection. The OAS proteins have a unique ability to produce 2′-5′ Oligoadenylates, which bind and activate the ribonuclease RNase L. The RNase L degrades cellular RNAs which in turn inhibits protein translation and induces apoptosis. Several single nucleotide polymorphisms (SNPs) in the OAS1 gene have been associated with disease. We have investigated the functional effect of two common SNPs in the OAS1 gene. The SNP rs10774671 affects splicing to one of the exons in the OAS1 gene giving rise to differential expression of the OAS1 isoforms, and the SNP rs1131454 (former rs3741981) resides in exon 3 giving rise to OAS1 isoforms with either a Glycine or a Serine at position 162 in the core OAS unit. Results We have used three human cell lines with different genotypes in the OAS1 SNP rs10774671, HeLa cells with the AA genotype, HT1080 cells with AG, and Daudi cells with GG. The main OAS1 isoform expressed in Daudi and HT1080 cells was p46, and the main OAS1 isoform expressed in HeLa cells was p42. In addition, low levels of the OAS1 p52 mRNA was detected in HeLa cells and p48 mRNA in Daudi cells, and trace amounts of p44a mRNA were detected in the three cell lines treated with type 1 interferon. We show that the OAS1 p46 isoform was localized in the mitochondria in Daudi cells, whereas the OAS1 isoforms in HeLa cells were primarily localized in cytoplasmic vacuoles/lysosomes. By using recombinantly expressed OAS1 mutant proteins, we found that the OAS1 SNP rs1131454 (former rs3741981) did not affect the enzymatic OAS1 activity. Conclusions The SNP rs10774671 determines differential expression of the OAS1 isoforms. In Daudi and HT1080 cells the p46 isoform is the most abundantly expressed isoform associated with the G allele, whereas in HeLa cells the most abundantly expressed isoform is p42 associated with the A allele. The SNP rs

  13. Identification of a locus (LCA9) for Leber's congenital amaurosis on chromosome 1p36.

    PubMed

    Keen, T Jeffrey; Mohamed, Moin D; McKibbin, Martin; Rashid, Yasmin; Jafri, Hussain; Maumenee, Irene H; Inglehearn, Chris F

    2003-05-01

    Leber's congenital amaurosis (LCA) is the most common cause of inherited childhood blindness and is characterised by severe retinal degeneration at or shortly after birth. We have identified a new locus, LCA9, on chromosome 1p36, at which the disease segregates in a single consanguineous Pakistani family. Following a whole genome linkage search, an autozygous region of 10 cM was identified between the markers D1S1612 and D1S228. Multipoint linkage analysis generated a lod score of 4.4, strongly supporting linkage to this region. The critical disease interval contains at least 5.7 Mb of DNA and around 50 distinct genes. One of these, retinoid binding protein 7 (RBP7), was screened for mutations in the family, but none was found.

  14. Therapeutic Strategies and Pharmacological Tools Influencing S1P Signaling and Metabolism.

    PubMed

    Vogt, Dominik; Stark, Holger

    2017-01-01

    During the last two decades the study of the sphingolipid anabolic, catabolic, and signaling pathways has attracted enormous interest. Especially the introduction of fingolimod into market as first p.o. therapeutic for the treatment of multiple sclerosis has boosted this effect. Although the complex regulation of sphingosine-1-phosphate (S1P) and other catabolic and anabolic sphingosine-related compounds is not fully understood, the influence on different (patho)physiological states from inflammation to cytotoxicity as well as the availability of versatile pharmacological tools that represent new approaches to study these states are described. Here, we have summarized various aspects concerning the many faces of sphingolipid function modulation by different pharmacological tools up to clinical candidates. Due to the immense heterogeneity of physiological or pharmacological actions and complex cross regulations, it is difficult to predict their role in upcoming therapeutic approaches. Currently, inflammatory, immunological, and/or antitumor aspects are discussed.

  15. Interactions within the yeast t-SNARE Sso1p that control SNARE complex assembly.

    PubMed

    Munson, M; Chen, X; Cocina, A E; Schultz, S M; Hughson, F M

    2000-10-01

    In the eukaryotic secretory and endocytic pathways, transport vesicles shuttle cargo among intracellular organelles and to and from the plasma membrane. Cargo delivery entails fusion of the transport vesicle with its target, a process thought to be mediated by membrane bridging SNARE protein complexes. Temporal and spatial control of intracellular trafficking depends in part on regulating the assembly of these complexes. In vitro, SNARE assembly is inhibited by the closed conformation adopted by the syntaxin family of SNAREs. To visualize this closed conformation directly, the X-ray crystal structure of a yeast syntaxin, Sso1p, has been determined and refined to 2.1 A resolution. Mutants designed to destabilize the closed conformation exhibit accelerated rates of SNARE assembly. Our results provide insight into the mechanism of SNARE assembly and its intramolecular and intermolecular regulation.

  16. Screening for hydrolytic enzymes reveals Ayr1p as a novel triacylglycerol lipase in Saccharomyces cerevisiae.

    PubMed

    Ploier, Birgit; Scharwey, Melanie; Koch, Barbara; Schmidt, Claudia; Schatte, Jessica; Rechberger, Gerald; Kollroser, Manfred; Hermetter, Albin; Daum, Günther

    2013-12-13

    Saccharomyces cerevisiae, as well as other eukaryotes, preserves fatty acids and sterols in a biologically inert form, as triacylglycerols and steryl esters. The major triacylglycerol lipases of the yeast S. cerevisiae identified so far are Tgl3p, Tgl4p, and Tgl5p (Athenstaedt, K., and Daum, G. (2003) YMR313c/TGL3 encodes a novel triacylglycerol lipase located in lipid particles of Saccharomyces cerevisiae. J. Biol. Chem. 278, 23317-23323; Athenstaedt, K., and Daum, G. (2005) Tgl4p and Tgl5p, two triacylglycerol lipases of the yeast Saccharomyces cerevisiae, are localized to lipid particles. J. Biol. Chem. 280, 37301-37309). We observed that upon cultivation on oleic acid, triacylglycerol mobilization did not come to a halt in a yeast strain deficient in all currently known triacylglycerol lipases, indicating the presence of additional not yet characterized lipases/esterases. Functional proteome analysis using lipase and esterase inhibitors revealed a subset of candidate genes for yet unknown hydrolytic enzymes on peroxisomes and lipid droplets. Based on the conserved GXSXG lipase motif, putative functions, and subcellular localizations, a selected number of candidates were characterized by enzyme assays in vitro, gene expression analysis, non-polar lipid analysis, and in vivo triacylglycerol mobilization assays. These investigations led to the identification of Ayr1p as a novel triacylglycerol lipase of yeast lipid droplets and confirmed the hydrolytic potential of the peroxisomal Lpx1p in vivo. Based on these results, we discuss a possible link between lipid storage, lipid mobilization, and peroxisomal utilization of fatty acids as a carbon source.

  17. Velocity-dependent optical potential for neutron elastic scattering from 1 p -shell nuclei

    NASA Astrophysics Data System (ADS)

    Ghabar, I. N.; Jaghoub, M. I.

    2015-06-01

    Background: The conventional optical model is quite successful in describing the nucleon elastic scattering data from medium and heavy nuclei. However, its success in describing the light 1 p -shell nuclei is somewhat limited. The velocity-dependent optical potential resulted in a significant improvement in describing the elastic angular distributions for light nuclei in the low energy region. Purpose: To extend the formalism of the velocity-dependent potential to higher energies, and to assess its importance in describing neutron elastic scattering data from light 1 p -shell nuclei at high energies. Method: We fit the angular distribution data for neutron elastic scattering from 12C and 16O using (i) the velocity-dependent optical potential and (ii) the conventional optical potential. The results of the two models are then compared. At low energies, we compare our angular distribution fits with the fits of other works that exist in the literature. Furthermore, the total integrated cross sections in addition to the analyzing power are calculated using the velocity-dependent optical potential and compared to the experimental data. Results: The velocity-dependent potential resulted in significant improvements in describing the angular distributions particularly in the large-angle scattering region and for certain energy ranges. This model is important where the experimental data show structural effects from nuclear surface deformations, which are important in light nuclei. Furthermore, the calculated total elastic cross sections and analyzing power are in good agreement with the experimental data. Conclusions: The velocity-dependent potential gives rise to surface-peaked real terms in the optical model. Such terms account, at least partly, for the structural effects seen in the angular distribution data. The energy range over which the surface terms are needed is found to depend on the target nucleus. Other works that have introduced real surface terms in the optical

  18. Deletion of FPS1, encoding aquaglyceroporin Fps1p, improves xylose fermentation by engineered Saccharomyces cerevisiae.

    PubMed

    Wei, Na; Xu, Haiqing; Kim, Soo Rin; Jin, Yong-Su

    2013-05-01

    Accumulation of xylitol in xylose fermentation with engineered Saccharomyces cerevisiae presents a major problem that hampers economically feasible production of biofuels from cellulosic plant biomass. In particular, substantial production of xylitol due to unbalanced redox cofactor usage by xylose reductase (XR) and xylitol dehydrogenase (XDH) leads to low yields of ethanol. While previous research focused on manipulating intracellular enzymatic reactions to improve xylose metabolism, this study demonstrated a new strategy to reduce xylitol formation and increase carbon flux toward target products by controlling the process of xylitol secretion. Using xylitol-producing S. cerevisiae strains expressing XR only, we determined the role of aquaglyceroporin Fps1p in xylitol export by characterizing extracellular and intracellular xylitol. In addition, when FPS1 was deleted in a poorly xylose-fermenting strain with unbalanced XR and XDH activities, the xylitol yield was decreased by 71% and the ethanol yield was substantially increased by nearly four times. Experiments with our optimized xylose-fermenting strain also showed that FPS1 deletion reduced xylitol production by 21% to 30% and increased ethanol yields by 3% to 10% under various fermentation conditions. Deletion of FPS1 decreased the xylose consumption rate under anaerobic conditions, but the effect was not significant in fermentation at high cell density. Deletion of FPS1 resulted in higher intracellular xylitol concentrations but did not significantly change the intracellular NAD(+)/NADH ratio in xylose-fermenting strains. The results demonstrate that Fps1p is involved in xylitol export in S. cerevisiae and present a new gene deletion target, FPS1, and a mechanism different from those previously reported to engineer yeast for improved xylose fermentation.

  19. Stress-activated protein kinase-mediated down-regulation of the cell integrity pathway mitogen-activated protein kinase Pmk1p by protein phosphatases.

    PubMed

    Madrid, Marisa; Núñez, Andrés; Soto, Teresa; Vicente-Soler, Jero; Gacto, Mariano; Cansado, José

    2007-11-01

    Fission yeast mitogen-activated protein kinase (MAPK) Pmk1p is involved in morphogenesis, cytokinesis, and ion homeostasis as part of the cell integrity pathway, and it becomes activated under multiple stresses, including hyper- or hypotonic conditions, glucose deprivation, cell wall-damaging compounds, and oxidative stress. The only protein phosphatase known to dephosphorylate and inactivate Pmk1p is Pmp1p. We show here that the stress-activated protein kinase (SAPK) pathway and its main effector, Sty1p MAPK, are essential for proper deactivation of Pmk1p under hypertonic stress in a process regulated by Atf1p transcription factor. We demonstrate that tyrosine phosphatases Pyp1p and Pyp2p, and serine/threonine phosphatase Ptc1p, that negatively regulate Sty1p activity and whose expression is dependent on Sty1p-Atf1p function, are involved in Pmk1p dephosphorylation under osmostress. Pyp1p and Ptc1p, in addition to Pmp1p, also control the basal level of MAPK Pmk1p activity in growing cells and associate with, and dephosphorylate Pmk1p both in vitro and in vivo. Our results with Ptc1p provide the first biochemical evidence for a PP2C-type phosphatase acting on more than one MAPK in yeast cells. Importantly, the SAPK-dependent down-regulation of Pmk1p through Pyp1p, Pyp2p, and Ptc1p was not complete, and Pyp1p and Ptc1p phosphatases are able to negatively regulate MAPK Pmk1p activity by an alternative regulatory mechanism. Our data also indicate that Pmk1p phosphorylation oscillates as a function of the cell cycle, peaking at cell separation during cytokinesis, and that Pmp1p phosphatase plays a main role in regulating this process.

  20. Stress-activated Protein Kinase-mediated Down-Regulation of the Cell Integrity Pathway Mitogen-activated Protein Kinase Pmk1p by Protein Phosphatases

    PubMed Central

    Madrid, Marisa; Núñez, Andrés; Soto, Teresa; Vicente-Soler, Jero; Cansado, José

    2007-01-01

    Fission yeast mitogen-activated protein kinase (MAPK) Pmk1p is involved in morphogenesis, cytokinesis, and ion homeostasis as part of the cell integrity pathway, and it becomes activated under multiple stresses, including hyper- or hypotonic conditions, glucose deprivation, cell wall-damaging compounds, and oxidative stress. The only protein phosphatase known to dephosphorylate and inactivate Pmk1p is Pmp1p. We show here that the stress-activated protein kinase (SAPK) pathway and its main effector, Sty1p MAPK, are essential for proper deactivation of Pmk1p under hypertonic stress in a process regulated by Atf1p transcription factor. We demonstrate that tyrosine phosphatases Pyp1p and Pyp2p, and serine/threonine phosphatase Ptc1p, that negatively regulate Sty1p activity and whose expression is dependent on Sty1p-Atf1p function, are involved in Pmk1p dephosphorylation under osmostress. Pyp1p and Ptc1p, in addition to Pmp1p, also control the basal level of MAPK Pmk1p activity in growing cells and associate with, and dephosphorylate Pmk1p both in vitro and in vivo. Our results with Ptc1p provide the first biochemical evidence for a PP2C-type phosphatase acting on more than one MAPK in yeast cells. Importantly, the SAPK-dependent down-regulation of Pmk1p through Pyp1p, Pyp2p, and Ptc1p was not complete, and Pyp1p and Ptc1p phosphatases are able to negatively regulate MAPK Pmk1p activity by an alternative regulatory mechanism. Our data also indicate that Pmk1p phosphorylation oscillates as a function of the cell cycle, peaking at cell separation during cytokinesis, and that Pmp1p phosphatase plays a main role in regulating this process. PMID:17761528

  1. The Effect of Gender on the N1-P2 Auditory Complex while Listening and Speaking with Altered Auditory Feedback

    ERIC Educational Resources Information Center

    Swink, Shannon; Stuart, Andrew

    2012-01-01

    The effect of gender on the N1-P2 auditory complex was examined while listening and speaking with altered auditory feedback. Fifteen normal hearing adult males and 15 females participated. N1-P2 components were evoked while listening to self-produced nonaltered and frequency shifted /a/ tokens and during production of /a/ tokens during nonaltered…

  2. T-bet–dependent S1P5 expression in NK cells promotes egress from lymph nodes and bone marrow

    PubMed Central

    Jenne, Craig N.; Enders, Anselm; Rivera, Richard; Watson, Susan R.; Bankovich, Alexander J.; Pereira, Joao P.; Xu, Ying; Roots, Carla M.; Beilke, Joshua N.; Banerjee, Arnob; Reiner, Steven L.; Miller, Sara A.; Weinmann, Amy S.; Goodnow, Chris C.

    2009-01-01

    During a screen for ethylnitrosourea-induced mutations in mice affecting blood natural killer (NK) cells, we identified a strain, designated Duane, in which NK cells were reduced in blood and spleen but increased in lymph nodes (LNs) and bone marrow (BM). The accumulation of NK cells in LNs reflected a decreased ability to exit into lymph. This strain carries a point mutation within Tbx21 (T-bet), which generates a defective protein. Duane NK cells have a 30-fold deficiency in sphingosine-1-phosphate receptor 5 (S1P5) transcript levels, and S1P5-deficient mice exhibit an egress defect similar to Duane. Chromatin immunoprecipitation confirms binding of T-bet to the S1pr5 locus. S1P-deficient mice exhibit a more severe NK cell egress block, and the FTY720-sensitive S1P1 also plays a role in NK cell egress from LNs. S1P5 is not inhibited by CD69, a property that may facilitate trafficking of activated NK cells to effector sites. Finally, the accumulation of NK cells within BM of S1P-deficient mice was associated with reduced numbers in BM sinusoids, suggesting a role for S1P in BM egress. In summary, these findings identify S1P5 as a T-bet–induced gene that is required for NK cell egress from LNs and BM. PMID:19808259

  3. Monosomy 1p36 uncovers a role for OX40 in survival of activated CD4+ T cells.

    PubMed

    Suhoski, M M; Perez, E E; Heltzer, M L; Laney, A; Shaffer, L G; Saitta, S; Nachman, S; Spinner, N B; June, C H; Orange, J S

    2008-08-01

    Monosomy 1p36 is a subtelomeric deletion syndrome associated with congenital anomalies presumably due to haploinsufficiency of multiple genes. Although immunodeficiency has not been reported, genes encoding costimulatory molecules of the TNF receptor superfamily (TNFRSF) are within 1p36 and may be affected. In one patient with monosomy 1p36, comparative genome hybridization and fluorescence in- situ hybridization confirmed that TNFRSF member OX40 was included within the subtelomeric deletion. T cells from this patient had decreased OX40 expression after stimulation. Specific, ex vivo T cell activation through OX40 revealed enhanced proliferation, and reduced viability of patient CD4+ T cells, providing evidence for the association of monosomy 1p36 with reduced OX40 expression, and decreased OX40-induced T cell survival. These results support a role for OX40 in human immunity, and calls attention to the potential for haploinsufficiency deletions of TNFRSF costimulatory molecules in monosomy 1p36.

  4. Increased mRNA Levels of Sphingosine Kinases and S1P Lyase and Reduced Levels of S1P Were Observed in Hepatocellular Carcinoma in Association with Poorer Differentiation and Earlier Recurrence.

    PubMed

    Uranbileg, Baasanjav; Ikeda, Hitoshi; Kurano, Makoto; Enooku, Kenichiro; Sato, Masaya; Saigusa, Daisuke; Aoki, Junken; Ishizawa, Takeaki; Hasegawa, Kiyoshi; Kokudo, Norihiro; Yatomi, Yutaka

    2016-01-01

    Although sphingosine 1-phosphate (S1P) has been reported to play an important role in cancer pathophysiology, little is known about S1P and hepatocellular carcinoma (HCC). To clarify the relationship between S1P and HCC, 77 patients with HCC who underwent surgical treatment were consecutively enrolled in this study. In addition, S1P and its metabolites were quantitated by LC-MS/MS. The mRNA levels of sphingosine kinases (SKs), which phosphorylate sphingosine to generate S1P, were increased in HCC tissues compared with adjacent non-HCC tissues. Higher mRNA levels of SKs in HCC were associated with poorer differentiation and microvascular invasion, whereas a higher level of SK2 mRNA was a risk factor for intra- and extra-hepatic recurrence. S1P levels, however, were unexpectedly reduced in HCC compared with non-HCC tissues, and increased mRNA levels of S1P lyase (SPL), which degrades S1P, were observed in HCC compared with non-HCC tissues. Higher SPL mRNA levels in HCC were associated with poorer differentiation. Finally, in HCC cell lines, inhibition of the expression of SKs or SPL by siRNA led to reduced proliferation, invasion and migration, whereas overexpression of SKs or SPL enhanced proliferation. In conclusion, increased SK and SPL mRNA expression along with reduced S1P levels were more commonly observed in HCC tissues compared with adjacent non-HCC tissues and were associated with poor differentiation and early recurrence. SPL as well as SKs may be therapeutic targets for HCC treatment.

  5. Pseudogene BMI1P1 expression as a novel predictor for acute myeloid leukemia development and prognosis

    PubMed Central

    Zhou, Ling-Yu; Zhai, Ling-Ling; Yin, Jia-Yu; Vanessa, Minse Evola-Deniz; Zhou, Jiao; Zhang, Jing; Tang, Xi; Lin, Jiang; Qian, Jun; Deng, Zhao-Qun

    2016-01-01

    The BMI1P1 levels of 144 de novo AML patients and 36 healthy donors were detected by real-time quantitative PCR (RQ-PCR). BMI1P1 was significantly down-regulated in AML compared with control (P < 0.001). A receiver operating characteristic (ROC) curve revealed that BMI1P1 expression could differentiate patients with AML from control subjects (AUC = 0.895, 95% CI: 0.835–0.954, P < 0.001). The percentage of blasts in bone marrow (BM) was significantly lower in BMI1P1 high-expressed group versus low-expressed group (P = 0.008). BMI1P1 high-expressed cases had significantly higher complete remission (CR) than BMI1P1 low-expressed cases (P = 0.023). Furthermore, Kaplan–Meier demonstrated that both whole AML cohort and non-M3-AML patients with low BMI1P1 expression showed shorter leukemia free survival (LFS, P = 0.002 and P = 0.01, respectively) and overall survival (OS, P < 0.001 and P = 0.011, respectively) than those with high BMI1P1 expression. Multivariate analysis also showed that BMI1P1 over-expression was an independent favorable prognostic factor for OS in both whole and non-M3 cohort of AML patients (HR = 0.462, 95% CI = 0.243–0.879, P = 0.019 and HR = 0.483, 95% CI = 0.254–0.919, P = 0.027). To further investigate the significance of BMI1P1 expression in the follow-up of AML patients, we monitored the BMI1P1 level in 26 de novo AML patients and found that the BMI1P1 level increased significantly from the initial diagnosis to post-CR (P < 0.001). These results indicated that BMI1P1 might contribute to the diagnosis of AML and the assessment of therapeutic effect. PMID:27329719

  6. Partial monosomy of chromosome 1p36.3: Characterization of the critical region and delineation of a syndrome

    SciTech Connect

    Reish, O.; Berry, S.A.; Hirsch, B.

    1995-12-04

    We describe 5 patients ranging in age from 3 to 47 years, with karyotypic abnormalities resulting in monosomy for portion of 1p36.3, microcephaly, mental retardation, prominent forehead, deep-set eyes, depressed nasal bridge, flat midface, relative prognathism, and abnormal ears. Four patients have small hands and feet. All exhibited selfabusive behavior. Additional findings in some of the patients include brain anomalies, optic atrophy, hearing loss and skeletal deformities. The breakpoints within chromosome 1 were designated at 1p36.31 (3 cases), 1p36.32 (1 case) and 1p36.33 (1 case). Thus, the smallest region of deletion overlap is 1p36.33{r_arrow}pter. Detection of the abnormal 1 relied on high resolution G-band analysis. Fluorescence in situ hybridization (FISH) utilizing a DNA probe (Oncor D1Z2) containing the repetitive sequences in distal 1p36, confirmed a deletion of one 1 homologue in all 5 cases. The abnormal 1 resulted from a de novo deletion in only one patient. The remaining patients were either confirmed (3 cases) or suspected (1 case) to have unbalanced translocations. Despite the additional genetic imbalance present in these four cases, monosomy of 1p36.33 appears to be responsible for a specific clinical phenotype. Characterization of this phenotype should assist in the clinical diagnosis of this chromosome abnormality. 26 refs., 4 figs., 2 tabs.

  7. Accurate, fast and cost-effective diagnostic test for monosomy 1p36 using real-time quantitative PCR.

    PubMed

    Cunha, Pricila da Silva; Pena, Heloisa B; D'Angelo, Carla Sustek; Koiffmann, Celia P; Rosenfeld, Jill A; Shaffer, Lisa G; Stofanko, Martin; Gonçalves-Dornelas, Higgor; Pena, Sérgio Danilo Junho

    2014-01-01

    Monosomy 1p36 is considered the most common subtelomeric deletion syndrome in humans and it accounts for 0.5-0.7% of all the cases of idiopathic intellectual disability. The molecular diagnosis is often made by microarray-based comparative genomic hybridization (aCGH), which has the drawback of being a high-cost technique. However, patients with classic monosomy 1p36 share some typical clinical characteristics that, together with its common prevalence, justify the development of a less expensive, targeted diagnostic method. In this study, we developed a simple, rapid, and inexpensive real-time quantitative PCR (qPCR) assay for targeted diagnosis of monosomy 1p36, easily accessible for low-budget laboratories in developing countries. For this, we have chosen two target genes which are deleted in the majority of patients with monosomy 1p36: PRKCZ and SKI. In total, 39 patients previously diagnosed with monosomy 1p36 by aCGH, fluorescent in situ hybridization (FISH), and/or multiplex ligation-dependent probe amplification (MLPA) all tested positive on our qPCR assay. By simultaneously using these two genes we have been able to detect 1p36 deletions with 100% sensitivity and 100% specificity. We conclude that qPCR of PRKCZ and SKI is a fast and accurate diagnostic test for monosomy 1p36, costing less than 10 US dollars in reagent costs.

  8. Yeast Asc1p and Mammalian RACK1 Are Functionally Orthologous Core 40S Ribosomal Proteins That Repress Gene Expression

    PubMed Central

    Gerbasi, Vincent R.; Weaver, Connie M.; Hill, Salisha; Friedman, David B.; Link, Andrew J.

    2004-01-01

    Translation of mRNA into protein is a fundamental step in eukaryotic gene expression requiring the large (60S) and small (40S) ribosome subunits and associated proteins. By modern proteomic approaches, we previously identified a novel 40S-associated protein named Asc1p in budding yeast and RACK1 in mammals. The goals of this study were to establish Asc1p or RACK1 as a core conserved eukaryotic ribosomal protein and to determine the role of Asc1p or RACK1 in translational control. We provide biochemical, evolutionary, genetic, and functional evidence showing that Asc1p or RACK1 is indeed a conserved core component of the eukaryotic ribosome. We also show that purified Asc1p-deficient ribosomes have increased translational activity compared to that of wild-type yeast ribosomes. Further, we demonstrate that asc1Δ null strains have increased levels of specific proteins in vivo and that this molecular phenotype is complemented by either Asc1p or RACK1. Our data suggest that one of Asc1p's or RACK1's functions is to repress gene expression. PMID:15340087

  9. Yeast Asc1p and mammalian RACK1 are functionally orthologous core 40S ribosomal proteins that repress gene expression.

    PubMed

    Gerbasi, Vincent R; Weaver, Connie M; Hill, Salisha; Friedman, David B; Link, Andrew J

    2004-09-01

    Translation of mRNA into protein is a fundamental step in eukaryotic gene expression requiring the large (60S) and small (40S) ribosome subunits and associated proteins. By modern proteomic approaches, we previously identified a novel 40S-associated protein named Asc1p in budding yeast and RACK1 in mammals. The goals of this study were to establish Asc1p or RACK1 as a core conserved eukaryotic ribosomal protein and to determine the role of Asc1p or RACK1 in translational control. We provide biochemical, evolutionary, genetic, and functional evidence showing that Asc1p or RACK1 is indeed a conserved core component of the eukaryotic ribosome. We also show that purified Asc1p-deficient ribosomes have increased translational activity compared to that of wild-type yeast ribosomes. Further, we demonstrate that asc1Delta null strains have increased levels of specific proteins in vivo and that this molecular phenotype is complemented by either Asc1p or RACK1. Our data suggest that one of Asc1p's or RACK1's functions is to repress gene expression.

  10. Structures of the yeast dynamin-like GTPase Sey1p provide insight into homotypic ER fusion

    PubMed Central

    Yan, Liming; Sun, Sha; Wang, Wei; Shi, Juanming; Hu, Xiaoyu; Wang, Shiyan; Su, Dan; Lou, Zhiyong

    2015-01-01

    Homotypic membrane fusion of the endoplasmic reticulum is mediated by dynamin-like guanosine triphosphatases (GTPases), which include atlastin (ATL) in metazoans and Sey1p in yeast. In this paper, we determined the crystal structures of the cytosolic domain of Sey1p derived from Candida albicans. The structures reveal a stalk-like, helical bundle domain following the GTPase, which represents a previously unidentified configuration of the dynamin superfamily. This domain is significantly longer than that of ATL and critical for fusion. Sey1p forms a side-by-side dimer in complex with GMP-PNP or GDP/AlF4− but is monomeric with GDP. Surprisingly, Sey1p could mediate fusion without GTP hydrolysis, even though fusion was much more efficient with GTP. Sey1p was able to replace ATL in mammalian cells, and the punctate localization of Sey1p was dependent on its GTPase activity. Despite the common function of fusogenic GTPases, our results reveal unique features of Sey1p. PMID:26370501

  11. The effect of S1P receptor signaling pathway on the survival and drug resistance in multiple myeloma cells.

    PubMed

    Fu, Di; Li, Yingchun; Li, Jia; Shi, Xiaoyan; Yang, Ronghui; Zhong, Yuan; Wang, Huihan; Liao, Aijun

    2017-01-01

    Multiple myeloma (MM) remains incurable by conventional chemotherapy. Sphingosine-1-phosphate (S1P) receptor-mediated signaling has been recently demonstrated to have critical roles in cell survival and drug resistance in a number of hematological malignancies. To dissect the roles of S1P receptor pathway in MM, we systematically examined cell viability and protein expression associated with cell survival and drug resistance in MM cell lines upon treatment with either pathway activator (S1P) or inhibitor (FTY720). Our results reveal that FTY720 inhibits cell proliferation by downregulating expression of target genes, while S1P has an opposite effect. Knocking down of S1P receptor S1P5R results in a reduction of cell survival-related gene expression; however, it does not have impacts on expression of drug resistance genes. These results suggest that S1P signaling plays a role in cell proliferation and drug resistance in MM, and targeting this pathway will provide a new therapeutic direction for MM management.

  12. Sphingosine kinase 2 (Sphk2) regulates platelet biogenesis by providing intracellular sphingosine 1-phosphate (S1P).

    PubMed

    Zhang, Lin; Urtz, Nicole; Gaertner, Florian; Legate, Kyle R; Petzold, Tobias; Lorenz, Michael; Mazharian, Alexandra; Watson, Steve P; Massberg, Steffen

    2013-08-01

    Human megakaryocytes (MKs) release trillions of platelets each day into the circulation to maintain normal homeostatic platelet levels. We have previously shown that extracellular sphingosine 1-phosphate (S1P) plays a key role in thrombopoiesis via its receptor S1pr1. In addition to its role as an extracellular mediator, S1P can also function as a second messenger in the intracellular compartment. Although signaling via intracellular S1P is involved in various cellular processes, a role in thrombopoiesis has not been examined. Sphingosine kinases are the key enzymes that produce intracellular S1P. Here we report that sphingosine kinase 2 (Sphk2) is the major messenger RNA species present in MKs. Sphk2 predominantly localizes to the nucleus and is the major source of intracellular S1P in MKs. Loss of Sphk2 significantly reduced intracellular S1P in MKs and downregulated the expression and activity of Src family kinases (SFKs). Loss of Sphk2 and inhibition of SFK activity resulted in defective intravascular proplatelet shedding, the final stage of thrombopoiesis. Correspondingly, mice lacking Sphk2 in the hematopoietic system display thrombocytopenia. Together, our data suggest that Sphk2 provides the source of intracellular S1P that controls thrombopoiesis, which is associated with SFK expression and activity in MKs.

  13. Identification of a Nfs1p-bound persulfide intermediate in Fe-S cluster synthesis by intact mitochondria

    PubMed Central

    Pandey, Alok; Yoon, Heeyong; Lyver, Elise R.; Dancis, Andrew; Pain, Debkumar

    2012-01-01

    Cysteine desulfurases generate a covalent persulfide intermediate from cysteine, and this activated form of sulfur is essential for the synthesis of iron-sulfur (Fe-S) clusters. In yeast mitochondria, there is a complete machinery for Fe-S cluster synthesis, including a cysteine desulfurase, Nfs1p. Here we show that following supplementation of isolated mitochondria with [35S]cysteine, a radiolabeled persulfide could be detected on Nfs1p. The persulfide persisted under conditions that did not permit Fe-S cluster formation, such as nucleotide and/or iron depletion of mitochondria. By contrast, under permissive conditions, the radiolabeled Nfs1p persulfide was greatly reduced and radiolabeled aconitase was formed, indicating transfer of persulfide to downstream Fe-S cluster recipients. Nfs1p in mitochondria was found to be relatively more resistant to inactivation by N-ethylmaleimide (NEM) as compared with a prokaryotic cysteine desulfurase. Mitochondria treated with NEM (1 mM) formed the persulfide on Nfs1p but failed to generate Fe-S clusters on aconitase, likely due to inactivation of downstream recipient(s) of the Nfs1p persulfide. Thus the Nfs1p-bound persulfide as described here represents a precursor en route to Fe-S cluster synthesis in mitochondria. PMID:22813754

  14. Identification of a Nfs1p-bound persulfide intermediate in Fe-S cluster synthesis by intact mitochondria.

    PubMed

    Pandey, Alok; Yoon, Heeyong; Lyver, Elise R; Dancis, Andrew; Pain, Debkumar

    2012-09-01

    Cysteine desulfurases generate a covalent persulfide intermediate from cysteine, and this activated form of sulfur is essential for the synthesis of iron-sulfur (Fe-S) clusters. In yeast mitochondria, there is a complete machinery for Fe-S cluster synthesis, including a cysteine desulfurase, Nfs1p. Here we show that following supplementation of isolated mitochondria with [(35)S]cysteine, a radiolabeled persulfide could be detected on Nfs1p. The persulfide persisted under conditions that did not permit Fe-S cluster formation, such as nucleotide and/or iron depletion of mitochondria. By contrast, under permissive conditions, the radiolabeled Nfs1p persulfide was greatly reduced and radiolabeled aconitase was formed, indicating transfer of persulfide to downstream Fe-S cluster recipients. Nfs1p in mitochondria was found to be relatively more resistant to inactivation by N-ethylmaleimide (NEM) as compared with a prokaryotic cysteine desulfurase. Mitochondria treated with NEM (1 mM) formed the persulfide on Nfs1p but failed to generate Fe-S clusters on aconitase, likely due to inactivation of downstream recipient(s) of the Nfs1p persulfide. Thus the Nfs1p-bound persulfide as described here represents a precursor en route to Fe-S cluster synthesis in mitochondria.

  15. Accurate, Fast and Cost-Effective Diagnostic Test for Monosomy 1p36 Using Real-Time Quantitative PCR

    PubMed Central

    Cunha, Pricila da Silva; Pena, Heloisa B.; D'Angelo, Carla Sustek; Koiffmann, Celia P.; Rosenfeld, Jill A.; Shaffer, Lisa G.; Stofanko, Martin; Gonçalves-Dornelas, Higgor; Pena, Sérgio Danilo Junho

    2014-01-01

    Monosomy 1p36 is considered the most common subtelomeric deletion syndrome in humans and it accounts for 0.5–0.7% of all the cases of idiopathic intellectual disability. The molecular diagnosis is often made by microarray-based comparative genomic hybridization (aCGH), which has the drawback of being a high-cost technique. However, patients with classic monosomy 1p36 share some typical clinical characteristics that, together with its common prevalence, justify the development of a less expensive, targeted diagnostic method. In this study, we developed a simple, rapid, and inexpensive real-time quantitative PCR (qPCR) assay for targeted diagnosis of monosomy 1p36, easily accessible for low-budget laboratories in developing countries. For this, we have chosen two target genes which are deleted in the majority of patients with monosomy 1p36: PRKCZ and SKI. In total, 39 patients previously diagnosed with monosomy 1p36 by aCGH, fluorescent in situ hybridization (FISH), and/or multiplex ligation-dependent probe amplification (MLPA) all tested positive on our qPCR assay. By simultaneously using these two genes we have been able to detect 1p36 deletions with 100% sensitivity and 100% specificity. We conclude that qPCR of PRKCZ and SKI is a fast and accurate diagnostic test for monosomy 1p36, costing less than 10 US dollars in reagent costs. PMID:24839341

  16. Mmr1p is a mitochondrial factor for Myo2p-dependent inheritance of mitochondria in the budding yeast.

    PubMed

    Itoh, Takashi; Toh-E, Akio; Matsui, Yasushi

    2004-07-07

    Class V myosins play a pivotal role in organelle distribution. In the budding yeast, Myo2p, a class V myosin, is essential for mitochondrial distribution. We identified MMR1 as a high-dose suppressor of the myo2 mitochondrial defect and that Mmr1p resides restrictively on the bud-localizing mitochondria and forms a complex with Myo2p tail. Mmr1p loss delayed mitochondrial transfer to buds and completely abolished mitochondrial distribution in the absence of Ypt11p, which promotes mitochondrial distribution by complex formation with Myo2p tail. The myo2-573 mutation, which causes a mitochondrial distribution defect and inactivates the Mmr1p function, reduced association between Myo2p and Mmr1p and depolarized Mmr1p localization on mitochondria. These strongly suggest that Mmr1p is a key mitochondrial component of the link between Myo2p and mitochondria for Myo2p-dependent mitochondrial distribution. Genetical analysis revealed that the Mmr1p-Myo2p pathway is independent of the Ypt11p-Myo2p pathway, suggesting that an essential system for mitochondrial distribution is composed of two independent Myo2p pathways.

  17. Glutathione depletion activates the yeast vacuolar transient receptor potential channel, Yvc1p, by reversible glutathionylation of specific cysteines

    PubMed Central

    Chandel, Avinash; Das, Krishna K.; Bachhawat, Anand K.

    2016-01-01

    Glutathione depletion and calcium influx into the cytoplasm are two hallmarks of apoptosis. We have been investigating how glutathione depletion leads to apoptosis in yeast. We show here that glutathione depletion in yeast leads to the activation of two cytoplasmically inward-facing channels: the plasma membrane, Cch1p, and the vacuolar calcium channel, Yvc1p. Deletion of these channels partially rescues cells from glutathione depletion–induced cell death. Subsequent investigations on the Yvc1p channel, a homologue of the mammalian TRP channels, revealed that the channel is activated by glutathionylation. Yvc1p has nine cysteine residues, of which eight are located in the cytoplasmic regions and one on the transmembrane domain. We show that three of these cysteines, Cys-17, Cys-79, and Cys-191, are specifically glutathionylated. Mutation of these cysteines to alanine leads to a loss in glutathionylation and a concomitant loss in calcium channel activity. We further investigated the mechanism of glutathionylation and demonstrate a role for the yeast glutathione S-transferase Gtt1p in glutathionylation. Yvc1p is also deglutathionylated, and this was found to be mediated by the yeast thioredoxin, Trx2p. A model for redox activation and deactivation of the yeast Yvc1p channel is presented. PMID:27708136

  18. Modulation of cellular S1P levels with a novel, potent and specific inhibitor of sphingosine kinase-1.

    PubMed

    Schnute, Mark E; McReynolds, Matthew D; Kasten, Tom; Yates, Matthew; Jerome, Gina; Rains, John W; Hall, Troii; Chrencik, Jill; Kraus, Michelle; Cronin, Ciaran N; Saabye, Matthew; Highkin, Maureen K; Broadus, Richard; Ogawa, Shinji; Cukyne, Kristin; Zawadzke, Laura E; Peterkin, Vincent; Iyanar, Kaliapan; Scholten, Jeffrey A; Wendling, Jay; Fujiwara, Hideji; Nemirovskiy, Olga; Wittwer, Arthur J; Nagiec, Marek M

    2012-05-15

    SphK (sphingosine kinase) is the major source of the bioactive lipid and GPCR (G-protein-coupled receptor) agonist S1P (sphingosine 1-phosphate). S1P promotes cell growth, survival and migration, and is a key regulator of lymphocyte trafficking. Inhibition of S1P signalling has been proposed as a strategy for treatment of inflammatory diseases and cancer. In the present paper we describe the discovery and characterization of PF-543, a novel cell-permeant inhibitor of SphK1. PF-543 inhibits SphK1 with a K(i) of 3.6 nM, is sphingosine-competitive and is more than 100-fold selective for SphK1 over the SphK2 isoform. In 1483 head and neck carcinoma cells, which are characterized by high levels of SphK1 expression and an unusually high rate of S1P production, PF-543 decreased the level of endogenous S1P 10-fold with a proportional increase in the level of sphingosine. In contrast with past reports that show that the growth of many cancer cell lines is SphK1-dependent, specific inhibition of SphK1 had no effect on the proliferation and survival of 1483 cells, despite a dramatic change in the cellular S1P/sphingosine ratio. PF-543 was effective as a potent inhibitor of S1P formation in whole blood, indicating that the SphK1 isoform of sphingosine kinase is the major source of S1P in human blood. PF-543 is the most potent inhibitor of SphK1 described to date and it will be useful for dissecting specific roles of SphK1-driven S1P signalling.

  19. High-precision spectroscopy of the HD+ molecule at the 1-p.p.b. level

    NASA Astrophysics Data System (ADS)

    Biesheuvel, J.; Karr, J.-Ph.; Hilico, L.; Eikema, K. S. E.; Ubachs, W.; Koelemeij, J. C. J.

    2017-01-01

    Recently we reported a high-precision optical frequency measurement of the ( v, L): (0, 2)→(8, 3) vibrational overtone transition in trapped deuterated molecular hydrogen (HD+) ions at 10 mK temperature. Achieving a resolution of 0.85 parts-per-billion (p.p.b.), we found the experimental value [ ν 0 = 383, 407, 177.38 (41) MHz] to be in agreement with the value from molecular theory [ ν th 383, 407, 177.150 (15) MHz] within 0.6 (1.1) p.p.b. (Biesheuvel et al. in Nat Commun 7:10385, 2016). This enabled an improved test of molecular theory (including QED), new constraints on the size of possible effects due to `new physics,' and the first determination of the proton-electron mass ratio from a molecule. Here, we provide the details of the experimental procedure, spectral analysis, and the assessment of systematic frequency shifts. Our analysis focuses in particular on deviations of the HD+ velocity distribution from thermal (Gaussian) distributions under the influence of collisions with fast ions produced during (laser-induced) chemical reactions, as such deviations turn out to significantly shift the hyperfine-less vibrational frequency as inferred from the saturated and Doppler-broadened spectrum, which contains partly unresolved hyperfine structure.

  20. The origin of chaos in the orbit of comet 1P/Halley

    NASA Astrophysics Data System (ADS)

    Boekholt, T. C. N.; Pelupessy, F. I.; Heggie, D. C.; Portegies Zwart, S. F.

    2016-10-01

    According to Muñoz-Gutiérrez et al. the orbit of comet 1P/Halley is chaotic with a surprisingly small Lyapunov time-scale of order its orbital period. In this work we analyse the origin of chaos in Halley's orbit and the growth of perturbations, in order to get a better understanding of this unusually short time-scale. We perform N-body simulations to model Halley's orbit in the Solar system and measure the separation between neighbouring trajectories. To be able to interpret the numerical results, we use a semi-analytical map to demonstrate different growth modes, i.e. linear, oscillatory or exponential, and transitions between these modes. We find the Lyapunov time-scale of Halley's orbit to be of order 300 yr, which is significantly longer than previous estimates in the literature. This discrepancy could be due to the different methods used to measure the Lyapunov time-scale. A surprising result is that next to Jupiter, also encounters with Venus contribute to the exponential growth in the next 3000 yr. Finally, we note an interesting application of the sub-linear, oscillatory growth mode to an ensemble of bodies moving through the Solar system. Whereas in the absence of encounters with a third body the ensemble spreads out linearly in time, the accumulation of weak encounters can increase the lifetime of such systems due to the oscillatory behaviour.

  1. Signaling, polyubiquitination, trafficking, and inclusions: sequestosome 1/p62's role in neurodegenerative disease.

    PubMed

    Wooten, Marie W; Hu, Xiao; Babu, J Ramesh; Seibenhener, M Lamar; Geetha, Thangiah; Paine, Michael G; Wooten, Michael C

    2006-01-01

    Aggregated misfolded proteins are hallmarks of most neurodegenerative diseases. In a chronic disease state, including pathologic situations of oxidative stress, these proteins are sequestered into inclusions. Accumulation of aggregated proteins can be prevented by chaperones, or by targeting their degradation to the UPS. If the accumulation of these proteins exceeds their degradation, they may impair the function of the proteasome. Alternatively, the function of the proteasome may be preserved by directing aggregated proteins to the autophagy-lysosome pathway for degradation. Sequestosome 1/p62 has recently been shown to interact with polyubiquitinated proteins through its UBA domain and may direct proteins to either the UPS or autophagosome. P62 is present in neuronal inclusions of individuals with Alzheimer's disease and other neurodegenerative diseases. Herein, we review p62's role in signaling, aggregation, and inclusion formation, and specifically as a possible contributor to Alzheimer's disease. The use of p62 as a potential target for the development of therapeutics and as a disease biomarker is also discussed.

  2. Comet 1P/Halley Multifluid MHD Model for the Giotto Fly-by

    NASA Astrophysics Data System (ADS)

    Rubin, M.; Combi, M. R.; Daldorff, L. K. S.; Gombosi, T. I.; Hansen, K. C.; Shou, Y.; Tenishev, V. M.; Tóth, G.; van der Holst, B.; Altwegg, K.

    2014-02-01

    The interaction of comets with the solar wind has been the focus of many studies including numerical modeling. We compare the results of our multifluid MHD simulation of comet 1P/Halley to data obtained during the flyby of the European Space Agency's Giotto spacecraft in 1986. The model solves the full set of MHD equations for the individual fluids representing the solar wind protons, the cometary light and heavy ions, and the electrons. The mass loading, charge-exchange, dissociative ion-electron recombination, and collisional interactions between the fluids are taken into account. The computational domain spans over several million kilometers, and the close vicinity of the comet is resolved to the details of the magnetic cavity. The model is validated by comparison to the corresponding Giotto observations obtained by the Ion Mass Spectrometer, the Neutral Mass Spectrometer, the Giotto magnetometer experiment, and the Johnstone Plasma Analyzer instrument. The model shows the formation of the bow shock, the ion pile-up, and the diamagnetic cavity and is able to reproduce the observed temperature differences between the pick-up ion populations and the solar wind protons. We give an overview of the global interaction of the comet with the solar wind and then show the effects of the Lorentz force interaction between the different plasma populations.

  3. [Effect of carbamazepine on epilepsy with 1p36 deletion syndrome].

    PubMed

    Nabatame, Shin; Okinaga, Takeshi; Imai, Katsumi; Kamio, Noriko; Kagitani-Shimono, Kuriko; Nagai, Toshisaburo; Kurosawa, Kenji; Ozono, Keiichi

    2007-07-01

    The 1p36 deletion syndrome is caused by submicroscopic deletion in the subtelomeric region of chromosome 1. Epilepsy is one of the most important features of the syndrome, in addition to the characteristic facial appearance, cardiac anomaly, dysphagia, deafness, mental retardation and growth delay. We identified three patients with this syndrome and assessed the features of complicated epilepsy. In all cases, epilepsy developed during infancy. The seizure types were mainly focal seizure and multiple seizure types including tonic seizure and tonic-clonic seizure. Interictal electroencephalogram showed focal abnormalities. Noticeably, two developed epileptic spasms and hypsarrhythmia in electroencephalogram, just after the administration of carbamazepine (CBZ). Including cases showing epileptic spasms, their epilepsy was easily tractable with anti-epileptic drugs, which could be withdrawn as they aged. All had deleted potassium channel beta subunit (KCNAB2) and gamma-aminobutyric acid A receptor delta (GABRD). CBZ may aggravate various epileptic syndromes, especially, those caused by GABA-A receptor gene mutation. Our cases may suggest the novel correspondence of GABA-A receptor-related epilepsy syndrome and exacerbation of epilepsy triggered by CBZ.

  4. HSF1 stress response pathway regulates autophagy receptor SQSTM1/p62-associated proteostasis

    PubMed Central

    Watanabe, Yoshihisa; Tsujimura, Atsushi; Taguchi, Katsutoshi; Tanaka, Masaki

    2017-01-01

    ABSTRACT Proteostasis is important for protecting cells from harmful proteins and is mainly controlled by the HSF1 (heat shock transcription factor 1) stress response pathway. This pathway facilitates protein refolding by molecular chaperones; however, it is unclear whether it functions in autophagy or inclusion formation. The autophagy receptor SQSTM1/p62 is involved in selective autophagic clearance and inclusion formation by harmful proteins, and its phosphorylation at S349, S403, and S407 is required for binding to substrates. Here, we demonstrate that casein kinase 1 phosphorylates the SQSTM1 S349 residue when harmful proteins accumulate. Investigation of upstream factors showed that both SQSTM1 S349 and SQSTM1 S403 residues were phosphorylated in an HSF1 dependent manner. Inhibition of SQSTM1 phosphorylation suppressed inclusion formation by ubiquitinated proteins and prevented colocalization of SQSTM1 with aggregation-prone proteins. Moreover, HSF1 inhibition impaired aggregate-induced autophagosome formation and elimination of protein aggregates. Our findings indicate that HSF1 triggers SQSTM1-mediated proteostasis. PMID:27846364

  5. Legionella pneumophila S1P-lyase targets host sphingolipid metabolism and restrains autophagy

    PubMed Central

    Rolando, Monica; Escoll, Pedro; Nora, Tamara; Botti, Joëlle; Boitez, Valérie; Daniels, Craig; Abraham, Gilu; Stogios, Peter J.; Skarina, Tatiana; Christophe, Charlotte; Dervins-Ravault, Delphine; Cazalet, Christel; Hilbi, Hubert; Rupasinghe, Thusitha W. T.; Tull, Dedreia; McConville, Malcolm J.; Ong, Sze Ying; Hartland, Elizabeth L.; Codogno, Patrice; Levade, Thierry; Naderer, Thomas; Savchenko, Alexei; Buchrieser, Carmen

    2016-01-01

    Autophagy is an essential component of innate immunity, enabling the detection and elimination of intracellular pathogens. Legionella pneumophila, an intracellular pathogen that can cause a severe pneumonia in humans, is able to modulate autophagy through the action of effector proteins that are translocated into the host cell by the pathogen’s Dot/Icm type IV secretion system. Many of these effectors share structural and sequence similarity with eukaryotic proteins. Indeed, phylogenetic analyses have indicated their acquisition by horizontal gene transfer from a eukaryotic host. Here we report that L. pneumophila translocates the effector protein sphingosine-1 phosphate lyase (LpSpl) to target the host sphingosine biosynthesis and to curtail autophagy. Our structural characterization of LpSpl and its comparison with human SPL reveals high structural conservation, thus supporting prior phylogenetic analysis. We show that LpSpl possesses S1P lyase activity that was abrogated by mutation of the catalytic site residues. L. pneumophila triggers the reduction of several sphingolipids critical for macrophage function in an LpSpl-dependent and -independent manner. LpSpl activity alone was sufficient to prevent an increase in sphingosine levels in infected host cells and to inhibit autophagy during macrophage infection. LpSpl was required for efficient infection of A/J mice, highlighting an important virulence role for this effector. Thus, we have uncovered a previously unidentified mechanism used by intracellular pathogens to inhibit autophagy, namely the disruption of host sphingolipid biosynthesis. PMID:26831115

  6. Adenosine induces apoptosis through TNFR1/RIPK1/P38 axis in colon cancer cells.

    PubMed

    Yu, Shunji; Hou, Daisen; Chen, Ping; Zhang, Qi; Lv, Bin; Ma, Yunfang; Liu, Fuchen; Liu, Hui; Song, Evelyn J; Yang, Dongqin; Liu, Jie

    2015-05-08

    Adenosine, a metabolite of ATP, ubiquitously exists in a wide range of organs and tissues. We previously reported that adenosine was implicated in apoptosis in many cancer cells by extrinsic and/or intrinsic pathways. Here, we found that adenosine suppresses the cell growth by induction of apoptosis of human colonic cancer cells through a novel mechanism. Adenosine suppresses the cell growth of human SW620 and SW480 colon cells in an adenosine transporter and adenosine kinase dependent manner. Moreover, the cell growth suppression is induced by apoptosis through activation of caspase-3 and PARP, and accumulation of ROS in cells. Importantly, we found that adenosine increases the expression of TNFR1 and RIPK1 and the phosphorylation of p38. Knockdown of TNFR1 or RIPK1 impairs the activation of p38, blocks the cleavage of PARP, and provides partially, yet significantly protection from cell death, including reducing the ROS generation in the colon cancer cells. These results indicate that a TNFR1/RIPK1/P38 axis is present in adenosine-induced apoptosis of colonic cancer cells. This axis triggers apoptosis and plays crucial roles in relay of the death signaling. Our study also provides additional experimental evidence for adenosine as a potent therapeutic drug in cancer therapy.

  7. cep-1/p53-dependent dysplastic pathology of the aging C. elegans gonad.

    PubMed

    McGee, Mathew D; Day, Nicholas; Graham, Jill; Melov, Simon

    2012-04-01

    The C. elegans germline and somatic gonad are actively developing until the animal reaches adulthood, and then continue to undergo striking changes as the animal ages. Reported changes include a depletion of available sperm, a decrease in oocyte quality up till mid-life, a reduction in germline nuclei, a decrease in fertility, and an accumulation of DNA in the midbody of aging C. elegans. Here, we have focused on the aging gonad in old animals, and show in detail that the aging gonad undergoes a massive uterine growth composed of endoreduplicating oocytes, yolk, and expanses of chromatin. We use a novel series of imaging techniques in combination with histological methodology for reconstructing aged worms in 3-dimensions, and show in old animals growing masses swelling inside the uterus to occupy most of the diameter of the worm. We link this accelerated growth to the cep-1/p53 tumor suppressor. Because cep-1 is required for DNA damage induced apoptosis, and daf-2 limits longevity, these results suggest a role for age-related DNA damage in dysplastic uterine growths, which in some respects resemble premalignant changes that can occur in aging mammals.

  8. Legionella pneumophila S1P-lyase targets host sphingolipid metabolism and restrains autophagy.

    PubMed

    Rolando, Monica; Escoll, Pedro; Nora, Tamara; Botti, Joëlle; Boitez, Valérie; Bedia, Carmen; Daniels, Craig; Abraham, Gilu; Stogios, Peter J; Skarina, Tatiana; Christophe, Charlotte; Dervins-Ravault, Delphine; Cazalet, Christel; Hilbi, Hubert; Rupasinghe, Thusitha W T; Tull, Dedreia; McConville, Malcolm J; Ong, Sze Ying; Hartland, Elizabeth L; Codogno, Patrice; Levade, Thierry; Naderer, Thomas; Savchenko, Alexei; Buchrieser, Carmen

    2016-02-16

    Autophagy is an essential component of innate immunity, enabling the detection and elimination of intracellular pathogens. Legionella pneumophila, an intracellular pathogen that can cause a severe pneumonia in humans, is able to modulate autophagy through the action of effector proteins that are translocated into the host cell by the pathogen's Dot/Icm type IV secretion system. Many of these effectors share structural and sequence similarity with eukaryotic proteins. Indeed, phylogenetic analyses have indicated their acquisition by horizontal gene transfer from a eukaryotic host. Here we report that L. pneumophila translocates the effector protein sphingosine-1 phosphate lyase (LpSpl) to target the host sphingosine biosynthesis and to curtail autophagy. Our structural characterization of LpSpl and its comparison with human SPL reveals high structural conservation, thus supporting prior phylogenetic analysis. We show that LpSpl possesses S1P lyase activity that was abrogated by mutation of the catalytic site residues. L. pneumophila triggers the reduction of several sphingolipids critical for macrophage function in an LpSpl-dependent and -independent manner. LpSpl activity alone was sufficient to prevent an increase in sphingosine levels in infected host cells and to inhibit autophagy during macrophage infection. LpSpl was required for efficient infection of A/J mice, highlighting an important virulence role for this effector. Thus, we have uncovered a previously unidentified mechanism used by intracellular pathogens to inhibit autophagy, namely the disruption of host sphingolipid biosynthesis.

  9. TRIM21 ubiquitylates SQSTM1/p62 and suppresses protein sequestration to regulate redox homeostasis

    PubMed Central

    Pan, Ji-An; Sun, Yu; Jiang, Ya-Ping; Bott, Alex J.; Jaber, Nadia; Dou, Zhixun; Yang, Bin; Chen, Juei-Suei; Catanzaro, Joseph M.; Du, Chunying; Ding, Wen-Xing; Diaz-Meco, Maria T.; Moscat, Jorge; Ozato, Keiko; Lin, Richard Z.; Zong, Wei-Xing

    2016-01-01

    Summary TRIM21 is a RING finger domain-containing ubiquitin E3 ligase whose expression is elevated in autoimmune disease. While TRIM21 plays an important role in immune activation during pathogen infection, little is known about its inherent cellular function. Here we show that TRIM21 plays an essential role in redox regulation by directly interacting with SQSTM1/p62 and ubiquitylating p62 at lysine(K)7 via K63-linkage. As p62 oligomerizes and sequesters client proteins in inclusions, the TRIM21-mediated p62 ubiquitylation abrogates p62 oligomerization and sequestration of proteins including Keap1, a negative regulator of antioxidant response. TRIM21-deficient cells display an enhanced antioxidant response and reduced cell death in response to oxidative stress. Genetic ablation of TRIM21 in mice confers protection from oxidative damages caused by arsenic-induced liver insult and pressure overload heart injury. Therefore, TRIM21 plays an essential role in p62-regulated redox homeostasis and may be a viable target for treating pathological conditions resulting from oxidative damage. PMID:26942676

  10. A conserved amphipathic helix is required for membrane tubule formation by Yop1p

    PubMed Central

    Brady, Jacob P.; Claridge, Jolyon K.; Smith, Peter G.; Schnell, Jason R.

    2015-01-01

    The integral membrane proteins of the DP1 (deleted in polyposis) and reticulon families are responsible for maintaining the high membrane curvature required for both smooth endoplasmic reticulum (ER) tubules and the edges of ER sheets, and mutations in these proteins lead to motor neuron diseases, such as hereditary spastic paraplegia. Reticulon/DP1 proteins contain reticulon homology domains (RHDs) that have unusually long hydrophobic segments and are proposed to adopt intramembrane helical hairpins that stabilize membrane curvature. We have characterized the secondary structure and dynamics of the DP1 family protein produced from the YOP1 gene (Yop1p) and identified a C-terminal conserved amphipathic helix (APH) that, on its own, interacts strongly with negatively charged membranes and is necessary for membrane tubule formation. Analyses of DP1 and reticulon family members indicate that most, if not all, contain C-terminal sequences capable of forming APHs. Together, these results indicate that APHs play a previously unrecognized role in RHD membrane curvature stabilization. PMID:25646439

  11. Triage of oxidation-prone proteins by Sqstm1/p62 within the mitochondria

    SciTech Connect

    Lee, Minjung; Shin, Jaekyoon

    2011-09-16

    Highlights: {yields} The mitochondrion contains its own protein quality control system. {yields} p62 localizes within the mitochondria and forms mega-dalton sized complexes. {yields} p62 interacts with oxidation-prone proteins and the proteins of quality control. {yields} In vitro delivery of p62 improves mitochondrial functions. {yields} p62 is implicated as a participant in mitochondrial protein quality control. -- Abstract: As the mitochondrion is vulnerable to oxidative stress, cells have evolved several strategies to maintain mitochondrial integrity, including mitochondrial protein quality control mechanisms and autophagic removal of damaged mitochondria. Involvement of an autophagy adaptor, Sqstm1/p62, in the latter process has been recently described. In the present study, we provide evidence that a portion of p62 directly localizes within the mitochondria and supports stable electron transport by forming heterogeneous protein complexes. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF) of mitochondrial proteins co-purified with p62 revealed that p62 interacts with several oxidation-prone proteins, including a few components of the electron transport chain complexes, as well as multiple chaperone molecules and redox regulatory enzymes. Accordingly, p62-deficient mitochondria exhibited compromised electron transport, and the compromised function was partially restored by in vitro delivery of p62. These results suggest that p62 plays an additional role in maintaining mitochondrial integrity at the vicinity of target machineries through its function in relation to protein quality control.

  12. A YAC contig encompassing the recessive Stargardt disease gene (STGD) on chromosome 1p.

    PubMed Central

    Anderson, K L; Baird, L; Lewis, R A; Chinault, A C; Otterud, B; Leppert, M; Lupski, J R

    1995-01-01

    Stargardt disease (STGD) and fundus flavimaculatus are infrequent autosomal recessive conditions characterized by a juvenile macular dystrophy and variable degrees of peripheral retinal changes. Linkage analysis performed in 47 STGD/fundus flavimaculatus families demonstrated significant linkage to 13 polymorphic DNA markers on chromosome 1p. The maximum combined two-point lod score was 32.7 (maximum recombination fraction [phi max] = .006) with the polymorphic marker D1S188. Our data demonstrate that STGD and fundus flavimaculatus are the same disorder clinically and genetically and provide further evidence for genetic homogeneity of this phenotype. Analysis of recombination events on disease chromosomes placed the STGD gene within a 4-cM interval between markers D1S435 and D1S236. A physical map was constructed of a YAC contig flanking STGD, from markers D1S500 to D1S495, and includes the critical interval delineated by historical recombinants. This contig spans approximately 31 cM, with one gap (3-5 cM) that is outside the 4-cM critical region. Localization of STGD to a single YAC contig will facilitate its positional cloning. Images Figure 3 PMID:8533764

  13. Negative Ion Chemistry in the Coma of Comet 1P/Halley

    NASA Technical Reports Server (NTRS)

    Cordiner, M. A.; Charnley, S. B.

    2012-01-01

    Negative ions (anions) were identified in the coma of comet 1P/Halley from in-situ measurements performed by the Giotto spacecraft in 1986. These anions were detected with masses in the range 7-110 amu, but with insufficient mass resolution to permit unambiguous identification. We present details of a new chemical-hydrodynamic model for the coma of comet Halley that includes - for the first time - atomic and molecular anions, in addition to a comprehensive hydrocarbon chemistry. Anion number densities arc calculated as a function of radius in the coma, and compared with the Giotto results. Important anion production mechanisms arc found to include radiative electron attachment, polar photodissociation, dissociative electron attachment, and proton transfer. The polyyne anions C4H(-) and C6H(-) arc found to be likely candidates to explain the Giotto anion mass spectrum in the range 49-73 amu. Thc CN(-) anion probably makes a significant contribution to the mass spectrum at 26 amu. Larger carbon-chain anions such as C8H(1) can explain the peak near 100 amu provided there is a source of large carbon-chain-bearing molecules from the cometary nucleus.

  14. Mapping of guanylin to murine chromosome 4 and human chromosome 1p34-p35

    SciTech Connect

    Sciaky, D.; Cohen, M.B.; Jenkins, N.A.

    1995-03-20

    Guanylin is a 15-amino-acid peptide similar in structure and in function to ST{sub a}, the heat stable enterotoxin of enterotoxigenic Escherichia coli (4). Both guanylin and ST{sub a} bind guanylyl cyclase-C (GC-C), resulting in increased levels of intracellular cGMP and induction of Cl- secretion (4) via the cystic fibrosis transmembrane regulator (CFM) (2). Guanylin is a highly regulated intestinal gene that is differentially expressed along the duodenal-to-colonic and villus-to-crypt axes. Guanylin mRNA abundance is maximal in the distal small intestine and proximal colon, where the mRNA is detected mainly in differentiated villus epithelial cells and superficial colonic epithelial cells, respectively. The murine guanylin gene (Guca2) has been isolated and sequenced; the gene is 1.7 kb and consists of 3 exons. We report here the mapping of Guca2 to mouse chromosome 4 by linkage analysis and to human chromosome region 1p34-p35 using fluorescence in situ hybridization (FISH). 20 refs., 2 figs.

  15. Comet 1P/Halley multifluid MHD model for the Giotto fly-by

    SciTech Connect

    Rubin, M.; Altwegg, K.; Combi, M. R.; Daldorff, L. K. S.; Gombosi, T. I.; Hansen, K. C.; Shou, Y.; Tenishev, V. M.; Tóth, G.; Van der Holst, B.

    2014-02-01

    The interaction of comets with the solar wind has been the focus of many studies including numerical modeling. We compare the results of our multifluid MHD simulation of comet 1P/Halley to data obtained during the flyby of the European Space Agency's Giotto spacecraft in 1986. The model solves the full set of MHD equations for the individual fluids representing the solar wind protons, the cometary light and heavy ions, and the electrons. The mass loading, charge-exchange, dissociative ion-electron recombination, and collisional interactions between the fluids are taken into account. The computational domain spans over several million kilometers, and the close vicinity of the comet is resolved to the details of the magnetic cavity. The model is validated by comparison to the corresponding Giotto observations obtained by the Ion Mass Spectrometer, the Neutral Mass Spectrometer, the Giotto magnetometer experiment, and the Johnstone Plasma Analyzer instrument. The model shows the formation of the bow shock, the ion pile-up, and the diamagnetic cavity and is able to reproduce the observed temperature differences between the pick-up ion populations and the solar wind protons. We give an overview of the global interaction of the comet with the solar wind and then show the effects of the Lorentz force interaction between the different plasma populations.

  16. Alcohol alters hepatic FoxO1, p53, and mitochondrial SIRT5 deacetylation function

    SciTech Connect

    Lieber, Charles S. Leo, Maria Anna; Wang, Xiaolei; DeCarli, Leonore M.

    2008-08-22

    Chronic alcohol consumption affects the gene expression of a NAD-dependent deacetylase Sirtuis 1 (SIRT1) and the peroxisome proliferator-activated receptor-{gamma} coactivator1{alpha} (PGC-1{alpha}). Our aim was to verify that it also alters the forkhead (FoxO1) and p53 transcription factor proteins, critical in the hepatic response to oxidative stress and regulated by SIRT1 through its deacetylating capacity. Accordingly, rats were pair-fed the Lieber-DeCarli alcohol-containing liquid diets for 28 days. Alcohol increased hepatic mRNA expression of FoxO1 (p = 0.003) and p53 (p = 0.001) while corresponding protein levels remained unchanged. However phospho-FoxO1 and phospho-Akt (protein kinase) were both decreased by alcohol consumption (p = 0.04 and p = 0.02, respectively) while hepatic p53 was found hyperacetylated (p = 0.017). Furthermore, mitochondrial SIRT5 was reduced (p = 0.0025), and PGC-1{alpha} hyperacetylated (p = 0.027), establishing their role in protein modification. Thus, alcohol consumption disrupts nuclear-mitochondrial interactions by post-translation protein modifications, which contribute to alteration of mitochondrial biogenesis through the newly discovered reduction of SIRT5.

  17. Oxidative damage to the promoter region of SQSTM1/p62 is common to neurodegenerative disease

    PubMed Central

    Du, Yifeng; Wooten, Michael C; Wooten, Marie W.

    2009-01-01

    Recently we reported that declined SQSTM1/p62 expression in Alzheimer disease brain was age-correlated with oxidative damage to the p62 promoter. The objective of this study was to examine whether oxidative damage to the p62 promoter is common to DNA recovered from brain of individuals with neurodegenerative disease. Increased 8-OHdG staining was observed in brain sections from Alzheimer’s disease (AD), Parkinson disease (PD), Huntington disease (HD), Frontotemporal dementia (FTD), and Pick’s disease compared to control subjects. In parallel, the p62 promoter exhibited elevated oxidative damage in samples from various diseases compared to normal brain, and damage was negatively correlated with p62 expression in FTD samples. Oxidative damage to the p62 promoter induced by H2O2 treatment decreased its transcriptional activity. In keeping with this observation, the transcriptional activity of a Sp-1 element deletion mutant displayed reduced stimulus-induced activity. These findings reveal that oxidative damage to the p62 promoter decreased its transcriptional activity and might therefore account for decreased expression of p62. Altogether these results suggest that pharmacological means to increase p62 expression may be beneficial in delaying the onset of neurodegeneration. PMID:19481605

  18. A YAC contig encompassing the recessive Stargardt disease gene (STGD) on chromosome 1p

    SciTech Connect

    Anderson, K.L.; Lewis, R.A.; Chinault, A.C.

    1995-12-01

    Stargardt disease (STGD) and fundus flavimaculatus are infrequent autosomal recessive conditions characterized by a juvenile macular dystrophy and variable degrees of peripheral retinal changes. Linkage analysis performed in 47 STGD/fundus flavimaculatus families demonstrated significant linkage to 13 polymorphic DNA markers on chromosome 1p. The maximum combined two-point lod score was 32.7 (maximum recombination fraction [{theta}{sub max}] = .006) with the polymorphic marker D1S188. Our data demonstrate that STGD and fundus flavimaculatus are the same disorder clinically and genetically and provide further evidence for genetic homogeneity of this phenotype. Analysis of recombination on disease chromosomes placed the STGD gene within a 4-cM interval between markers D1S435 and D1S236. A physical map was constructed of a YAC contig flanking STGD, from markers D1S500 to D1S495, and includes the critical interval delineated by historical recombinants. This contig spans {approximately}31 cM, with one gap (3-5 cM) that is outside the 4-cM critical region. Localization of STGD to a single YAC contig will facilitate its positional cloning. 75 refs., 3 figs., 21 tabs.

  19. An endogenous promoter for conditional gene expression in Acremonium chrysogenum: the xylan and xylose inducible promoter xyl1(P.).

    PubMed

    Blatzer, Michael; Gsaller, Fabio; Abt, Beate; Schrettl, Markus; Specht, Thomas; Haas, Hubertus

    2014-01-01

    Acremonium chrysogenum is the natural producer of the beta-lactam antibiotic cephalosporin C and therefore of significant biotechnological importance. Here we identified and characterized the xylanase-encoding xyl1 gene and demonstrate that its promoter, xyl1(P), is suitable for conditional expression of heterologous genes in A. chrysogenum. This was shown by xylose and xylan-inducible xyl1(P)-driven expression of genes encoding green fluorescence protein and phleomycin resistance. Moreover, we demonstrate the potential of the xyl1(P) promoter for selection marker recycling. Taken together, these finding will help to overcome the limitation in genetic tools in this important filamentous fungus.

  20. New cross-bridged cyclam derivative CB-TE1K1P, an improved bifunctional chelator for copper radionuclides.

    PubMed

    Zeng, Dexing; Ouyang, Qin; Cai, Zhengxin; Xie, Xiang-Qun; Anderson, Carolyn J

    2014-01-04

    A new cross-bridged cyclam chelator, CB-TE1K1P, was developed for copper-based radiopharmaceuticals, and this chelator can be labelled with (64)Cu under mild conditions in high specific activity. DBCO-PEG4-CB-TE1K1P was synthesized for conjugation to proteins, while Dde-CB-TE1K1P((t)Bu2)-OH was synthesized for solid-phase peptide synthesis. Examples of the conjugation chemistry, radiolabelling and serum stability of each are presented.

  1. TVENT1P user's manual, a computer code for analyzing tornado-induced gas-dynamic transients in flow networks

    SciTech Connect

    Andrae, R.W.; Tang, P.K.; Gregory, W.S.

    1984-09-01

    TVENT1P is a revised version of the TVENT computer code, which was designed to predict the flows and pressures in a ventilation system subjected to a tornado. TVENT1P is essentially the same code, but we have added a material transport algorithm and features for turning blowers off and on, changing blower speeds, and changing the resistance of dampers and filters. These features make it possible to depict a sequence of events during a single run. Other features have been added to make the code more versatile. Example problems are included to demonstrate applications for TVENT1P.

  2. 78 FR 22361 - Social Security Ruling, SSR 13-1p; Titles II and XVI: Agency Processes for Addressing Allegations...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-04-15

    ... From the Federal Register Online via the Government Publishing Office SOCIAL SECURITY ADMINISTRATION Social Security Ruling, SSR 13-1p; Titles II and XVI: Agency Processes for Addressing Allegations of Unfairness, Prejudice, Partiality, Bias, Misconduct, or Discrimination by Administrative...

  3. 78 FR 9987 - Social Security Ruling, SSR 13-1p; Titles II and XVI: Agency Processes for Addressing Allegations...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-12

    ... From the Federal Register Online via the Government Publishing Office SOCIAL SECURITY ADMINISTRATION Social Security Ruling, SSR 13-1p; Titles II and XVI: Agency Processes for Addressing Allegations of Unfairness, Prejudice, Partiality, Bias, Misconduct, or Discrimination by Administrative...

  4. 78 FR 8217 - Social Security Ruling, SSR 13-1p; Titles II and XVI: Agency Processes for Addressing Allegations...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-05

    ... From the Federal Register Online via the Government Publishing Office SOCIAL SECURITY ADMINISTRATION Social Security Ruling, SSR 13-1p; Titles II and XVI: Agency Processes for Addressing Allegations of Unfairness, Prejudice, Partiality, Bias, Misconduct, or Discrimination by Administrative...

  5. Modeling the neutral gas and dust coma of Comet 1P/Halley

    NASA Astrophysics Data System (ADS)

    Rubin, Martin; Tenishev, Valeriy M.; Combi, Michael R.; Hansen, Kenneth C.; Gombosi, Tamas I.; Altwegg, Kathrin; Balsiger, Hans

    2010-05-01

    The neutral gas environment of a comet is largely influenced by dissociation of parent molecules created at the surface of the comet and collisions of all the involved species. We compare the results from a kinetic model of the neutral cometary environment with measurements from the Neutral Mass Spectrometer (NMS) and the Dust Impact Detection System (DIDSY) onboard the Giotto spacecraft which flew-by at comet 1P/Halley in 1986. We further show that our model is in good agreement to measurements obtained by the International Ultraviolet Explorer (IUE), sounding rocket experiments, and the International Halley Watch (IHW). The model solves the Boltzmann equation with a Direct Simulation Monte Carlo technique [Tenishev et al. (2008, Astrophys. J., 685, 659-677)] by tracking trajectories of gas molecules and dust grains under the influence of the comet's weak gravity field with momentum exchange among particles modeled in a probabilistic manner. The cometary nucleus is considered to be the source of dust and the parent species (in our model: H2O, CO, H2CO, CO2, CH3OH, C2H6, C2H4, C2H2, HCN, NH3, and CH4) in the coma. Subsequently our model also tracks the corresponding dissociation products (H, H2, O, OH, C, CH, CH2, CH3, N, NH, NH2, C2, C2H, C2H5, CN, and HCO). This work has been supported by JPL subcontract 1266313 under NASA grant NMO710889, NASA planetary atmospheres program grant NNX09AB59G, grant AST-0707283 from the NSF Planetary Astronomy program, and the Swiss National Science Foundation.

  6. SQSTM1/p62 interacts with HDAC6 and regulates deacetylase activity.

    PubMed

    Yan, Jin; Seibenhener, Michael Lamar; Calderilla-Barbosa, Luis; Diaz-Meco, Maria-Theresa; Moscat, Jorge; Jiang, Jianxiong; Wooten, Marie W; Wooten, Michael C

    2013-01-01

    Protein aggregates can form in the cytoplasm of the cell and are accumulated at aggresomes localized to the microtubule organizing center (MTOC) where they are subsequently degraded by autophagy. In this process, aggregates are engulfed into autophagosomes which subsequently fuse with lysosomes for protein degradation. A member of the class II histone deacetylase family, histone deacetylase 6(HDAC6) has been shown to be involved in both aggresome formation and the fusion of autophagosomes with lysosomes making it an attractive target to regulate protein aggregation. The scaffolding protein sequestosome 1(SQSTM1)/p62 has also been shown to regulate accumulation and autophagic clearance of protein aggregates. Recent studies have revealed colocalization of HDAC6 and p62 to ubiquitinated mitochondria, as well as, ubiquitinated protein aggregates associated with the E3 ubiquitin ligase TRIM50. HDAC6 deacetylase activity is required for aggresome formation and can be regulated by protein interaction with HDAC6. Due to their colocalization at ubiquitinated protein aggregates, we sought to examine if p62 specifically interacted with HDAC6 and if so, if this interaction had any effect on HDAC6 activity and/or the physiological function of cortactin-F-actin assembly. We succeeded in identifying and mapping the direct interaction between HDAC6 and p62. We further show that this interaction regulates HDAC6 deacetylase activity. Data are presented demonstrating that the absence of p62 results in hyperactivation of HDAC6 and deacetylation of α-tubulin and cortactin. Further, upon induction of protein misfolding we show that p62 is required for perinuclear co-localization of cortactin-F-actin assemblies. Thus, our findings indicate that p62 plays a key role in regulating the recruitment of F-actin network assemblies to the MTOC, a critical cellular function that is required for successful autophagic clearance of protein aggregates.

  7. HIV-1 p6 - a structured to flexible multifunctional membrane-interacting protein.

    PubMed

    Solbak, Sara Marie Øie; Reksten, Tove Ragna; Hahn, Friedrich; Wray, Victor; Henklein, Petra; Henklein, Peter; Halskau, Øyvind; Schubert, Ulrich; Fossen, Torgils

    2013-02-01

    The human immunodeficiency virus type 1 (HIV-1) p6 protein has recently been recognized as a docking site for several cellular and viral binding partners and is important for the formation of infectious viruses. Most of its known functions are suggested to occur under hydrophobic conditions near the cytoplasmic membrane, where the protein is presumed to exist in its most structured state. Although p6 is involved in manifold specific interactions, the protein has previously been considered to possess a random structure in aqueous solution. We show that p6 exhibits a defined structure with N- and C-terminal helical domains, connected by a flexible hinge region in 100mM dodecylphosphocholine micelle solution at pH 7 devoid of any organic co-solvents, indicating that this is a genuine limiting structural feature of the molecule in a hydrophobic environment. Furthermore, we show that p6 directly interacts with a cytoplasmic model membrane through both N-terminal and C-terminal regions by use of surface plasmon resonance (SPR) spectroscopy. Phosphorylation of Ser-40 located in the center of the C-terminal α-helix does not alter the secondary structure of the protein but amplifies the interaction with membranes significantly, indicating that p6 binds to the polar head groups at the surface of the cytoplasmic membrane. The increased hydrophobic membrane interaction of p6(23-52) S40F correlated with the observed increased amount of the polyprotein Gag in the RIPA insoluble fraction when Ser40 of p6 was mutated with Phe indicating that p6 modulates the membrane interactions of HIV-1 Gag.

  8. Identification of Risk and Age-at-Onset Genes on Chromosome 1p in Parkinson Disease

    PubMed Central

    Oliveira, Sofia A.; Li, Yi-Ju; Noureddine, Maher A.; Züchner, Stephan; Qin, Xuejun; Pericak-Vance, Margaret A.; Vance, Jeffery M.

    2005-01-01

    We previously reported a linkage region on chromosome 1p (LOD = 3.41) for genes controlling age at onset (AAO) in Parkinson disease (PD). This region overlaps with the previously reported PARK10 locus. To identify the gene(s) associated with AAO and risk of PD in this region, we first applied a genomic convergence approach that combined gene expression and linkage data. No significant results were found. Second, we performed association mapping across a 19.2-Mb region centered under the AAO linkage peak. An iterative association mapping approach was done by initially genotyping single-nucleotide polymorphisms at an average distance of 100 kb apart and then by increasing the density of markers as needed. Using the overall data set of 267 multiplex families, we identified six associated genes in the region, but further screening of a subset of 83 families linked to the chromosome 1 locus identified only two genes significantly associated with AAO in PD: the γ subunit of the translation initiation factor EIF2B gene (EIF2B3), which was more significant in the linked subset and the ubiquitin-specific protease 24 gene (USP24). Unexpectedly, the human immunodeficiency virus enhancer-binding protein 3 gene (HIVEP3) was found to be associated with risk for susceptibility to PD. We used several criteria to define significant results in the presence of multiple testing, including criteria derived from a novel cluster approach. The known or putative functions of these genes fit well with the current suspected pathogenic mechanisms of PD and thus show great potential as candidates for the PARK10 locus. PMID:15986317

  9. A Chemogenomic Screen Reveals Novel Snf1p/AMPK Independent Regulators of Acetyl-CoA Carboxylase.

    PubMed

    Bozaquel-Morais, Bruno L; Madeira, Juliana B; Venâncio, Thiago M; Pacheco-Rosa, Thiago; Masuda, Claudio A; Montero-Lomeli, Monica

    2017-01-01

    Acetyl-CoA carboxylase (Acc1p) is a key enzyme in fatty acid biosynthesis and is essential for cell viability. To discover new regulators of its activity, we screened a Saccharomyces cerevisiae deletion library for increased sensitivity to soraphen A, a potent Acc1p inhibitor. The hits identified in the screen (118 hits) were filtered using a chemical-phenotype map to exclude those associated with pleiotropic drug resistance. This enabled the identification of 82 ORFs that are genetic interactors of Acc1p. The main functional clusters represented by these hits were "transcriptional regulation", "protein post-translational modifications" and "lipid metabolism". Further investigation of the "transcriptional regulation" cluster revealed that soraphen A sensitivity is poorly correlated with ACC1 transcript levels. We also studied the three top unknown ORFs that affected soraphen A sensitivity: SOR1 (YDL129W), SOR2 (YIL092W) and SOR3 (YJR039W). Since the C18/C16 ratio of lipid acyl lengths reflects Acc1p activity levels, we evaluated this ratio in the three mutants. Deletion of SOR2 and SOR3 led to reduced acyl lengths, suggesting that Acc1p is indeed down-regulated in these strains. Also, these mutants showed no differences in Snf1p/AMPK activation status and deletion of SNF1 in these backgrounds did not revert soraphen A sensitivity completely. Furthermore, plasmid maintenance was reduced in sor2Δ strain and this trait was shared with 18 other soraphen A sensitive hits. In summary, our screen uncovered novel Acc1p Snf1p/AMPK-independent regulators.

  10. Albumin modulates S1P delivery from red blood cells in perfused microvessels: mechanism of the protein effect.

    PubMed

    Adamson, R H; Clark, J F; Radeva, M; Kheirolomoom, A; Ferrara, K W; Curry, F E

    2014-04-01

    Removal of plasma proteins from perfusates increases vascular permeability. The common interpretation of the action of albumin is that it forms part of the permeability barrier by electrostatic binding to the endothelial glycocalyx. We tested the alternate hypothesis that removal of perfusate albumin in rat venular microvessels decreased the availability of sphingosine-1-phosphate (S1P), which is normally carried in plasma bound to albumin and lipoproteins and is required to maintain stable baseline endothelial barriers (Am J Physiol Heart Circ Physiol 303: H825-H834, 2012). Red blood cells (RBCs) are a primary source of S1P in the normal circulation. We compared apparent albumin permeability coefficients [solute permeability (Ps)] measured using perfusates containing albumin (10 mg/ml, control) and conditioned by 20-min exposure to rat RBCs with Ps when test perfusates were in RBC-conditioned protein-free Ringer solution. The control perfusate S1P concentration (439 ± 46 nM) was near the normal plasma value at 37 °C and established a stable baseline Ps (0.9 ± 0.4 × 10(-6) cm/s). Ringer solution perfusate contained 52 ± 8 nM S1P and increased Ps more than 10-fold (16.1 ± 3.9 × 10(-6) cm/s). Consistent with albumin-dependent transport of S1P from RBCs, S1P concentrations in RBC-conditioned solutions decreased as albumin concentration, hematocrit, and temperature decreased. Protein-free Ringer solution perfusates that used liposomes instead of RBCs as flow markers failed to maintain normal permeability, reproducing the "albumin effect" in these mammalian microvessels. We conclude that the albumin effect depends on the action of albumin to facilitate the release and transport of S1P from RBCs that normally provide a significant amount of S1P to the endothelium.

  11. SCFCdc4-mediated Degradation of the Hac1p Transcription Factor Regulates the Unfolded Protein Response in Saccharomyces cerevisiae

    PubMed Central

    Pal, Bhupinder; Chan, Nickie C.; Helfenbaum, Leon; Tan, Kaeling; Tansey, William P.

    2007-01-01

    The Saccharomyces cerevisiae basic leucine zipper transcription factor Hac1p is synthesized in response to the accumulation of unfolded polypeptides in the lumen of the endoplasmic reticulum (ER), and it is responsible for up-regulation of ∼5% of all yeast genes, including ER-resident chaperones and protein-folding catalysts. Hac1p is one of the most short-lived yeast proteins, having a half-life of ∼1.5 min. Here, we have shown that Hac1p harbors a functional PEST degron and that degradation of Hac1p by the proteasome involves the E2 ubiquitin-conjugating enzyme Ubc3/Cdc34p and the SCFCdc4 E3 complex. Consistent with the known nuclear localization of Cdc4p, rapid degradation of Hac1p requires the presence of a functional nuclear localization sequence, which we demonstrated to involve basic residues in the sequence 29RKRAKTK35. Two-hybrid analysis demonstrated that the PEST-dependent interaction of Hac1p with Cdc4p requires Ser146 and Ser149. Turnover of Hac1p may be dependent on transcription because it is inhibited in cell mutants lacking Srb10 kinase, a component of the SRB/mediator module of the RNA polymerase II holoenzyme. Stabilization of Hac1p by point mutation or deletion, or as the consequence of defects in components of the degradation pathway, results in increased unfolded protein response element-dependent transcription and improved cell viability under ER stress conditions. PMID:17108329

  12. A Chemogenomic Screen Reveals Novel Snf1p/AMPK Independent Regulators of Acetyl-CoA Carboxylase

    PubMed Central

    Bozaquel-Morais, Bruno L.; Madeira, Juliana B.; Venâncio, Thiago M.; Pacheco-Rosa, Thiago; Masuda, Claudio A.; Montero-Lomeli, Monica

    2017-01-01

    Acetyl-CoA carboxylase (Acc1p) is a key enzyme in fatty acid biosynthesis and is essential for cell viability. To discover new regulators of its activity, we screened a Saccharomyces cerevisiae deletion library for increased sensitivity to soraphen A, a potent Acc1p inhibitor. The hits identified in the screen (118 hits) were filtered using a chemical-phenotype map to exclude those associated with pleiotropic drug resistance. This enabled the identification of 82 ORFs that are genetic interactors of Acc1p. The main functional clusters represented by these hits were “transcriptional regulation”, “protein post-translational modifications” and “lipid metabolism”. Further investigation of the “transcriptional regulation” cluster revealed that soraphen A sensitivity is poorly correlated with ACC1 transcript levels. We also studied the three top unknown ORFs that affected soraphen A sensitivity: SOR1 (YDL129W), SOR2 (YIL092W) and SOR3 (YJR039W). Since the C18/C16 ratio of lipid acyl lengths reflects Acc1p activity levels, we evaluated this ratio in the three mutants. Deletion of SOR2 and SOR3 led to reduced acyl lengths, suggesting that Acc1p is indeed down-regulated in these strains. Also, these mutants showed no differences in Snf1p/AMPK activation status and deletion of SNF1 in these backgrounds did not revert soraphen A sensitivity completely. Furthermore, plasmid maintenance was reduced in sor2Δ strain and this trait was shared with 18 other soraphen A sensitive hits. In summary, our screen uncovered novel Acc1p Snf1p/AMPK-independent regulators. PMID:28076367

  13. 1p36.32 rearrangements and the role of PI-PLC η2 in nervous tumours.

    PubMed

    Lo Vasco, Vincenza Rita

    2011-07-01

    Deletions in the distal region of the short arm of chromosome 1 (1p36) are widely diffuse, both in congenital 1p36 Deletion Syndrome and as somatic abnormalities in tumours. Rearrangements in 1p36 have been described in a broad spectrum of human neoplasias in addition to other chromosomal abnormalities. In neuroblastomas, wide hemizygous deletions in 1p36.23-1p36.32 have been described suggesting that the 1p36 region contains a tumour-suppressor gene involved in malignancy. A role for phosphoinositide (PI)-specific phospholipase C (PLC) η2, whose gene maps on 1p36.32, was suggested. PI-PLC η2 belongs to a family of enzymes related to the phosphoinositide signalling pathway, which provide an important intracellular signalling system involved in a variety of cell functions such as hormone secretion, neurotransmitter signal transduction, cell growth, membrane trafficking, ion channel activity, regulation of the cytoskeleton, cell cycle control and apoptosis. Expression of PI-PLC η2 occurs after birth and continues throughout the life. Synapse formation occurs during a short period of postnatal development. Thus, it is likely that PI-PLC η2 acts in formation and maintenance of the neuronal network in the brain. The fact that PI-PLC η2, a highly neuron-specific isozyme, is abundantly expressed in the postnatal brain suggests the importance of PI-PLC η2 in formation and maintenance of the neuronal network in the postnatal brain. Further studies are required to verify the possible involvement of PI-PLC η2 mutation/deletion in central nervous tumour tissues presenting abnormalities of the 1p36 chromosomal band.

  14. Conformational analysis on the wild type and mutated forms of human ORF1p: a molecular dynamics study.

    PubMed

    Muthukumaran, Rajagopalan; Sangeetha, Balasubramanian; Amutha, Ramaswamy

    2015-07-01

    The protein ORF1p, encoded by the LINE-1 retrotransposon, is responsible for the packaging and transposition of its RNA transcript and is reported to be involved in various genetic disorders. The three domains of ORF1p co-ordinate together to facilitate the transposition, and the mechanism of nucleic acid binding is not yet clear. The C-terminal domain of ORF1p adopts a lifted, twisted or rested state, which is regulated by several inter- and intra-domain interactions that are explored in this study. The residues, Glu147, Asp151, Lys154, Arg261 and Tyr282, are majorly involved in mediating the functional dynamics of ORF1p by forming H-bonds and π-interactions. The importance of these residues was elucidated by performing molecular dynamics simulations on both native as well as mutated ORF1p. The Q147A-D151A-K154A mutant expressed unique dynamics featuring the lifting motion of the CTD core alone, while the R261A mutant resulted in the oscillatory motion of CTD. In both cases, the CTDs were held in place by Tyr282 and in its absence, the structural stability of CTDs in the trimeric unit was significantly affected. Additional interactions responsible for stabilizing the trimeric ORF1p to express its native dynamics were extracted in this study. The central role of Tyr282 in maintaining the functional state of ORF1p to facilitate nucleic acid binding and formation of ribonucleoprotein complex is well highlighted. The knowledge gained from this study forms the basis for understanding the nucleic acid binding mechanism of ORF1p, which could further provide additional support in exploring various genetic disorders.

  15. SCFCdc4-mediated degradation of the Hac1p transcription factor regulates the unfolded protein response in Saccharomyces cerevisiae.

    PubMed

    Pal, Bhupinder; Chan, Nickie C; Helfenbaum, Leon; Tan, Kaeling; Tansey, William P; Gething, Mary-Jane

    2007-02-01

    The Saccharomyces cerevisiae basic leucine zipper transcription factor Hac1p is synthesized in response to the accumulation of unfolded polypeptides in the lumen of the endoplasmic reticulum (ER), and it is responsible for up-regulation of approximately 5% of all yeast genes, including ER-resident chaperones and protein-folding catalysts. Hac1p is one of the most short-lived yeast proteins, having a half-life of approximately 1.5 min. Here, we have shown that Hac1p harbors a functional PEST degron and that degradation of Hac1p by the proteasome involves the E2 ubiquitin-conjugating enzyme Ubc3/Cdc34p and the SCF(Cdc4) E3 complex. Consistent with the known nuclear localization of Cdc4p, rapid degradation of Hac1p requires the presence of a functional nuclear localization sequence, which we demonstrated to involve basic residues in the sequence (29)RKRAKTK(35). Two-hybrid analysis demonstrated that the PEST-dependent interaction of Hac1p with Cdc4p requires Ser146 and Ser149. Turnover of Hac1p may be dependent on transcription because it is inhibited in cell mutants lacking Srb10 kinase, a component of the SRB/mediator module of the RNA polymerase II holoenzyme. Stabilization of Hac1p by point mutation or deletion, or as the consequence of defects in components of the degradation pathway, results in increased unfolded protein response element-dependent transcription and improved cell viability under ER stress conditions.

  16. Anti-S1P antibody as a novel therapeutic strategy for VEGFR TKI resistant renal cancer

    PubMed Central

    Bullock, Andrea J.; Callea, Marcella; Shah, Harleen; Song, Jiaxi; Moreno, Kelli; Visentin, Barbara; Deutschman, Douglas; Alsop, David C.; Atkins, Michael B.; Mier, James W.; Signoretti, Sabina; Bhasin, Manoj; Sabbadini, Roger A.; Bhatt, Rupal S.

    2015-01-01

    Purpose VEGFR2 tyrosine kinase inhibition (TKI) is a valuable treatment approach for patients with metastatic RCC. However, resistance to treatment is inevitable. Identification of novel targets could lead to better treatment for both patients with TKI naïve or resistant RCC. Experimental design In this study, we performed transcriptome analysis of VEGFR TKI resistant tumors in a murine model and discovered that the SPHK/S1P pathway is upregulated at the time of resistance. We tested S1P pathway inhibition using an anti-S1P mAb (sphingomab), in two mouse xenograft models of RCC, and assessed tumor SPHK expression and S1P plasma levels in patients with metastatic RCC. Results Resistant tumors expressed several hypoxia regulated genes. The SPHK1 pathway was among the most highly upregulated pathways that accompanied resistance to VEGFR TKI therapy. SPHK1 was expressed in human RCC, and the product of SPHK1 activity, S1P, was elevated in patients with metastatic RCC suggesting that human RCC behavior could, in part, be due to over-production of S1P. Sphingomab neutralization of extracellular S1P slowed tumor growth in both mouse models. Mice bearing tumors that had developed resistance to sunitinib treatment also exhibited tumor growth suppression with sphingomab. Sphingomab treatment led to a reduction in tumor blood flow as measured by MRI. Conclusions Our findings suggest that S1P inhibition may be a novel therapeutic strategy in patients with treatment naïve RCC and also in the setting of resistance to VEGFR TKI therapy. PMID:25589614

  17. Effect of multidrug resistance 1/P-glycoprotein on the hypoxia-induced multidrug resistance of human laryngeal cancer cells.

    PubMed

    Li, Dawei; Zhou, Liang; Huang, Jiameng; Xiao, Xiyan

    2016-08-01

    In a previous study, it was demonstrated that hypoxia upregulated the multidrug resistance (MDR) of laryngeal cancer cells to chemotherapeutic drugs, with multidrug resistance 1 (MDR1)/P-glycoprotein (P-gp) expression also being upregulated. The present study aimed to investigate the role and mechanism of MDR1/P-gp on hypoxia-induced MDR in human laryngeal carcinoma cells. The sensitivity of laryngeal cancer cells to multiple drugs and cisplatin-induced apoptosis was determined by CCK-8 assay and Annexin-V/propidium iodide staining analysis, respectively. The accumulation of rhodamine 123 (Rh123) in the cells served as an estimate of drug accumulation and was evaluated by flow cytometry (FCM). MDR1/P-gp expression was inhibited using interference RNA, and the expression of the MDR1 gene was analyzed using reverse transcription-quantitative polymerase chain reaction and western blotting. As a result, the sensitivity to multiple chemotherapeutic agents and the apoptosis rate of the hypoxic laryngeal carcinoma cells increased following a decrease in MDR1/P-gp expression (P<0.05). Additionally, FCM analysis of fluorescence intensity indicated that the downregulated expression of MDR1/P-gp markedly increased intracellular Rh123 accumulation (P<0.05). Such results suggest that MDR1/P-gp serves an important role in regulating hypoxia-induced MDR in human laryngeal carcinoma cells through a decrease in intracellular drug accumulation.

  18. Chl1p, a DNA helicase-like protein in budding yeast, functions in sister-chromatid cohesion.

    PubMed Central

    Skibbens, Robert V

    2004-01-01

    From the time of DNA replication until anaphase onset, sister chromatids remain tightly paired along their length. Ctf7p/Eco1p is essential to establish sister-chromatid pairing during S-phase and associates with DNA replication components. DNA helicases precede the DNA replication fork and thus will first encounter chromatin sites destined for cohesion. In this study, I provide the first evidence that a DNA helicase is required for proper sister-chromatid cohesion. Characterizations of chl1 mutant cells reveal that CHL1 interacts genetically with both CTF7/ECO1 and CTF18/CHL12, two genes that function in sister-chromatid cohesion. Consistent with genetic interactions, Chl1p physically associates with Ctf7p/Eco1p both in vivo and in vitro. Finally, a functional assay reveals that Chl1p is critical for sister-chromatid cohesion. Within the budding yeast genome, Chl1p exhibits the highest degree of sequence similarity to human CHL1 isoforms and BACH1. Previous studies revealed that human CHLR1 exhibits DNA helicase-like activities and that BACH1 is a helicase-like protein that associates with the tumor suppressor BRCA1 to maintain genome integrity. Our findings document a novel role for Chl1p in sister-chromatid cohesion and provide new insights into the possible mechanisms through which DNA helicases may contribute to cancer progression when mutated. PMID:15020404

  19. 1p36 deletion syndrome confirmed by fluorescence in situ hybridization and array-comparative genomic hybridization analysis

    PubMed Central

    Kang, Dong Soo; Shin, Eunsim

    2016-01-01

    Pediatric epilepsy can be caused by various conditions, including specific syndromes. 1p36 deletion syndrome is reported in 1 in 5,000–10,000 newborns, and its characteristic clinical features include developmental delay, mental retardation, hypotonia, congenital heart defects, seizure, and facial dysmorphism. However, detection of the terminal deletion in chromosome 1p by conventional G-banded karyotyping is difficult. Here we present a case of epilepsy with profound developmental delay and characteristic phenotypes. A 7-year- and 6-month-old boy experienced afebrile generalized seizure at the age of 5 years and 3 months. He had recurrent febrile seizures since 12 months of age and showed severe global developmental delay, remarkable hypotonia, short stature, and dysmorphic features such as microcephaly; small, low-set ears; dark, straight eyebrows; deep-set eyes; flat nasal bridge; midface hypoplasia; and a small, pointed chin. Previous diagnostic work-up, including conventional chromosomal analysis, revealed no definite causes. However, array-comparative genomic hybridization analysis revealed 1p36 deletion syndrome with a 9.15-Mb copy loss of the 1p36.33-1p36.22 region, and fluorescence in situ hybridization analysis (FISH) confirmed this diagnosis. This case highlights the need to consider detailed chromosomal study for patients with delayed development and epilepsy. Furthermore, 1p36 deletion syndrome should be considered for patients presenting seizure and moderate-to-severe developmental delay, particularly if the patient exhibits dysmorphic features, short stature, and hypotonia. PMID:28018437

  20. Extending the phenotype of monosomy 1p36 syndrome and mapping of a critical region for obesity and hyperphagia.

    PubMed

    D'Angelo, Carla S; Kohl, Ilana; Varela, Monica Castro; de Castro, Cláudia I E; Kim, Chong A; Bertola, Débora R; Lourenço, Charles M; Koiffmann, Célia P

    2010-01-01

    Rearrangements of 1p36 are the most frequently detected abnormalities in diagnostic testing for chromosomal cryptic imbalances and include variably sized simple terminal deletions, derivative chromosomes, interstitial deletions, and complex rearrangements. These rearrangements result in the specific pattern of malformation and neurodevelopmental disabilities that characterizes monosomy 1p36 syndrome. Thus far, no individual gene within this region has been conclusively determined to be causative of any component of the phenotype. Nor is it known if the rearrangements convey phenotypes via a haploinsufficiency mechanism or through a position effect. We have used multiplex ligation-dependent probe amplification to screen for deletions of 1p36 in a group of 154 hyperphagic and overweight/obese, PWS negative individuals, and in a separate group of 83 patients initially sent to investigate a variety of other conditions. The strategy allowed the identification and delineation of rearrangements in nine subjects with a wide spectrum of clinical presentations. Our work reinforces the association of monosomy 1p36 and obesity and hyperphagia, and further suggests that these features may be associated with non-classical manifestations of this disorder in addition to a submicroscopic deletion of approximately 2-3 Mb in size. Multiplex ligation probe amplification using the monosomy 1p36 syndrome-specific kit coupled to the subtelomeric kit is an effective approach to identify and delineate rearrangements at 1p36.

  1. 1p36 deletion syndrome confirmed by fluorescence in situ hybridization and array-comparative genomic hybridization analysis.

    PubMed

    Kang, Dong Soo; Shin, Eunsim; Yu, Jeesuk

    2016-11-01

    Pediatric epilepsy can be caused by various conditions, including specific syndromes. 1p36 deletion syndrome is reported in 1 in 5,000-10,000 newborns, and its characteristic clinical features include developmental delay, mental retardation, hypotonia, congenital heart defects, seizure, and facial dysmorphism. However, detection of the terminal deletion in chromosome 1p by conventional G-banded karyotyping is difficult. Here we present a case of epilepsy with profound developmental delay and characteristic phenotypes. A 7-year- and 6-month-old boy experienced afebrile generalized seizure at the age of 5 years and 3 months. He had recurrent febrile seizures since 12 months of age and showed severe global developmental delay, remarkable hypotonia, short stature, and dysmorphic features such as microcephaly; small, low-set ears; dark, straight eyebrows; deep-set eyes; flat nasal bridge; midface hypoplasia; and a small, pointed chin. Previous diagnostic work-up, including conventional chromosomal analysis, revealed no definite causes. However, array-comparative genomic hybridization analysis revealed 1p36 deletion syndrome with a 9.15-Mb copy loss of the 1p36.33-1p36.22 region, and fluorescence in situ hybridization analysis (FISH) confirmed this diagnosis. This case highlights the need to consider detailed chromosomal study for patients with delayed development and epilepsy. Furthermore, 1p36 deletion syndrome should be considered for patients presenting seizure and moderate-to-severe developmental delay, particularly if the patient exhibits dysmorphic features, short stature, and hypotonia.

  2. [Effects of lead and selenium on telomere binding protein Rap1p, telomerase and telomeric DNA in Saccharomyces cerevisiae].

    PubMed

    Cui, Qing-Hua; Tang, Chia-Chun; Huang, You-Guo

    2002-03-01

    The effects on S.cerevisiae telomere binding protein Rap1p, telomerase and telomeric DNA by the lead (Pb), the selenium (Se) and Pb + Se were tested respectively in this study. Compared with the control S.cerevisiae after 100 gene rations, the mean telomere length shortened, Rap1p concentration was significantly lower and the secondary structure of Rap1p was disturbed, the telomerase activity was reduced in Pb treated cells. In Se treated cells, telomere length was significantly longer, and telomerase activity expressed higher. The concentration and secondary structure of Rap1p were similar to that of the control. Further more, the viability of Pb treated cells were significantly reduced while cells undergone other three treatments were similar and normal. These results suggest that Pb could damage Rap1p, reduce telomerase activity, resulting in the telomer length shortening and cell death. On the other hand, Se could protect and repair the damage in Rap1p and telomere caused by Pb to some extent.

  3. Role of JAK-STAT pathway in reducing cardiomyocytes hypoxia/reoxygenation injury induced by S1P postconditioning.

    PubMed

    Wang, Yuqing; Wang, Dongfei; Zhang, Lizhi; Ye, Fangyu; Li, Mengmeng; Wen, Ke

    2016-08-05

    This experiment was designed to explore the protection of sphingosine1-phosphate (S1P) postconditioning on rat myocardial cells injured by hypoxia/reoxygenation acting via the Janus kinase-signal transducer and activator of transcription (JAK-STAT) signal pathway. The data showed that S1P could significantly increase cell viability, lower the rate of apoptosis, decrease the content of lactate dehydrogenase (LDH) and caspase3 activity in the culture medium, increase the activity of total superoxide dismutase (T-SOD) and manganese superoxide dismutase (Mn-SOD), reduce the loss of mitochondrial membrane potential and the fluorescence intensity of intracellular calcium, as well as increase the phosphorylation of JAK2 and STAT3 in comparison with the H/R group. When the JAK inhibitor AG490 or the STAT inhibitor stattic were added, the effects of S1P were inhibited. Our date shows that S1P protects H9c2 cells from hypoxia/reoxygenation injury and that the protection by S1P was inhibited by AG490 and stattic. Therefore S1P protects H9c2 cells against hypoxia/reoxygenation injury via the JAK-STAT pathway.

  4. Cdc42p regulation of the yeast formin Bni1p mediated by the effector Gic2p

    PubMed Central

    Chen, Hsin; Kuo, Chun-Chen; Kang, Hui; Howell, Audrey S.; Zyla, Trevin R.; Jin, Michelle; Lew, Daniel J.

    2012-01-01

    Actin filaments are dynamically reorganized to accommodate ever-changing cellular needs for intracellular transport, morphogenesis, and migration. Formins, a major family of actin nucleators, are believed to function as direct effectors of Rho GTPases, such as the polarity regulator Cdc42p. However, the presence of extensive redundancy has made it difficult to assess the in vivo significance of the low-affinity Rho GTPase–formin interaction and specifically whether Cdc42p polarizes the actin cytoskeleton via direct formin binding. Here we exploit a synthetically rewired budding yeast strain to eliminate the redundancy, making regulation of the formin Bni1p by Cdc42p essential for viability. Surprisingly, we find that direct Cdc42p–Bni1p interaction is dispensable for Bni1p regulation. Alternative paths linking Cdc42p and Bni1p via “polarisome” components Spa2p and Bud6p are also collectively dispensable. We identify a novel regulatory input to Bni1p acting through the Cdc42p effector, Gic2p. This pathway is sufficient to localize Bni1p to the sites of Cdc42p action and promotes a polarized actin organization in both rewired and wild-type contexts. We suggest that an indirect mechanism linking Rho GTPases and formins via Rho effectors may provide finer spatiotemporal control for the formin-nucleated actin cytoskeleton. PMID:22918946

  5. The Saccharomyces cerevisiae v-SNARE Vti1p Is Required for Multiple Membrane Transport Pathways to the Vacuole

    PubMed Central

    von Mollard, Gabriele Fischer; Stevens, Tom H.

    1999-01-01

    The interaction between v-SNAREs on transport vesicles and t-SNAREs on target membranes is required for membrane traffic in eukaryotic cells. Here we identify Vti1p as the first v-SNARE protein found to be required for biosynthetic traffic into the yeast vacuole, the equivalent of the mammalian lysosome. Certain vti1-ts yeast mutants are defective in alkaline phosphatase transport from the Golgi to the vacuole and in targeting of aminopeptidase I from the cytosol to the vacuole. VTI1 interacts genetically with the vacuolar t-SNARE VAM3, which is required for transport of both alkaline phosphatase and aminopeptidase I to the vacuole. The v-SNARE Nyv1p forms a SNARE complex with Vam3p in homotypic vacuolar fusion; however, we find that Nyv1p is not required for any of the three biosynthetic pathways to the vacuole. v-SNAREs were thought to ensure specificity in membrane traffic. However, Vti1p also functions in two additional membrane traffic pathways: Vti1p interacts with the t-SNAREs Pep12p in traffic from the TGN to the prevacuolar compartment and with Sed5p in retrograde traffic to the cis-Golgi. The ability of Vti1p to mediate multiple fusion steps requires additional proteins to ensure specificity in membrane traffic. PMID:10359592

  6. The modification of Gat1p in nitrogen catabolite repression to enhance non-preferred nitrogen utilization in Saccharomyces cerevisiae

    PubMed Central

    Zhao, Xinrui; Zou, Huijun; Chen, Jian; Du, Guocheng; Zhou, Jingwen

    2016-01-01

    In Saccharomyces cerevisiae, when preferred nitrogen sources are present, the metabolism of non-preferred nitrogen is repressed. Previous work showed that this metabolic regulation is primarily controlled by nitrogen catabolite repression (NCR) related regulators. Among these regulators, two positive regulators (Gln3p and Gat1p) could be phosphorylated and sequestered in the cytoplasm leading to the transcription of non-preferred nitrogen metabolic genes being repressed. The nuclear localization signals (NLSs) and nuclear localization regulatory signals (NLRSs) in Gln3p and Gat1p play essential roles in the regulation of their localization in cells. However, compared with Gln3p, the information of NLS and NLRS for Gat1p remains unknown. In this study, residues 348–375 and 366–510 were identified as the NLS and NLRS of Gat1p firstly. In addition, the modifications of Gat1p (mutations on the NLS and truncation on the NLRS) were attempted to enhance the transcription of non-preferred nitrogen metabolic genes. Quantitative real-time PCR showed that the transcriptional levels of 15 non-preferred nitrogen metabolic genes increased. Furthermore, during the shaking-flask culture tests, the utilization of urea, proline and allantoine was significantly increased. Based on these results, the genetic engineering on Gat1p has a great potential in enhancing non-preferred nitrogen metabolism in S. cerevisiae. PMID:26899143

  7. Pheromone responsiveness is regulated by components of the Gpr1p-mediated glucose sensing pathway in Saccharomyces cerevisiae.

    PubMed

    Willhite, D Grant; Brigati, Jennifer R; Selcer, Katie E; Denny, Joshua E; Duck, Zachary A; Wright, Stephen E

    2014-09-01

    Many fungi have evolved mechanisms to assess environmental nutrient availability prior to the energy-intensive process of mating. In this study, we examined one such system in Saccharomyces cerevisiae, involving a glucose-sensing pathway mediated by Gpr1p and the pheromone-induced mating pathway. Initially we observed that the mating pathway in MATa cells is sensitive to environmental glucose depletion. This phenomenon can be partially reversed with a high glucose spike, but not with the addition of low levels of glucose. Deletion of the low-affinity glucose receptor, Gpr1p, eliminated this glucose-induced recovery of pheromone responsiveness. We then determined the impact of GPR1 deletion on the mating pathway and observed that, in all end points studied, the mating pathway response to pheromone is reduced in the absence of Gpr1p. Similarly, elimination of the Gα for Gpr1p, Gpa2p, resulted in reduction in pheromone sensitivity in all assays studied. The negative effect of removing Gpr1p on mating pathway activation could be recovered by overexpressing the mating receptor, Ste2p. Furthermore, Ste2p levels are reduced in the absence of glucose and GPR1. These data suggest that activity of the GPCR-mediated mating pathway in S. cerevisiae is modulated by extracellular glucose concentrations through the only other GPCR in MATa cells, Gpr1p.

  8. Birth of Archaeal Cells: Molecular Phylogenetic Analyses of G1P Dehydrogenase, G3P Dehydrogenases, and Glycerol Kinase Suggest Derived Features of Archaeal Membranes Having G1P Polar Lipids

    PubMed Central

    2016-01-01

    Bacteria and Eukarya have cell membranes with sn-glycerol-3-phosphate (G3P), whereas archaeal membranes contain sn-glycerol-1-phosphate (G1P). Determining the time at which cells with either G3P-lipid membranes or G1P-lipid membranes appeared is important for understanding the early evolution of terrestrial life. To clarify this issue, we reconstructed molecular phylogenetic trees of G1PDH (G1P dehydrogenase; EgsA/AraM) which is responsible for G1P synthesis and G3PDHs (G3P dehydrogenase; GpsA and GlpA/GlpD) and glycerol kinase (GlpK) which is responsible for G3P synthesis. Together with the distribution of these protein-encoding genes among archaeal and bacterial groups, our phylogenetic analyses suggested that GlpA/GlpD in the Commonote (the last universal common ancestor of all extant life with a cellular form, Commonote commonote) acquired EgsA (G1PDH) from the archaeal common ancestor (Commonote archaea) and acquired GpsA and GlpK from a bacterial common ancestor (Commonote bacteria). In our scenario based on this study, the Commonote probably possessed a G3P-lipid membrane synthesized enzymatically, after which the archaeal lineage acquired G1PDH followed by the replacement of a G3P-lipid membrane with a G1P-lipid membrane. PMID:27774041

  9. Association of Constitutive Hyperphosphorylation of Hsf1p with a Defective Ethanol Stress Response in Saccharomyces cerevisiae Sake Yeast Strains

    PubMed Central

    Noguchi, Chiemi; Watanabe, Daisuke; Zhou, Yan; Akao, Takeshi

    2012-01-01

    Modern sake yeast strains, which produce high concentrations of ethanol, are unexpectedly sensitive to environmental stress during sake brewing. To reveal the underlying mechanism, we investigated a well-characterized yeast stress response mediated by a heat shock element (HSE) and heat shock transcription factor Hsf1p in Saccharomyces cerevisiae sake yeast. The HSE-lacZ activity of sake yeast during sake fermentation and under acute ethanol stress was severely impaired compared to that of laboratory yeast. Moreover, the Hsf1p of modern sake yeast was highly and constitutively hyperphosphorylated, irrespective of the extracellular stress. Since HSF1 allele replacement did not significantly affect the HSE-mediated ethanol stress response or Hsf1p phosphorylation patterns in either sake or laboratory yeast, the regulatory machinery of Hsf1p is presumed to function differently between these types of yeast. To identify phosphatases whose loss affected the control of Hsf1p, we screened a series of phosphatase gene deletion mutants in a laboratory strain background. Among the 29 mutants, a Δppt1 mutant exhibited constitutive hyperphosphorylation of Hsf1p, similarly to the modern sake yeast strains, which lack the entire PPT1 gene locus. We confirmed that the expression of laboratory yeast-derived functional PPT1 recovered the HSE-mediated stress response of sake yeast. In addition, deletion of PPT1 in laboratory yeast resulted in enhanced fermentation ability. Taken together, these data demonstrate that hyperphosphorylation of Hsf1p caused by loss of the PPT1 gene at least partly accounts for the defective stress response and high ethanol productivity of modern sake yeast strains. PMID:22057870

  10. Oxidized LDL-induced angiogenesis involves sphingosine 1-phosphate: prevention by anti-S1P antibody

    PubMed Central

    Camaré, Caroline; Trayssac, Magali; Garmy-Susini, Barbara; Mucher, Elodie; Sabbadini, Roger; Salvayre, Robert; Negre-Salvayre, Anne

    2015-01-01

    BACKGROUND AND PURPOSE Neovascularization occurring in atherosclerotic lesions may promote plaque expansion, intraplaque haemorrhage and rupture. Oxidized LDL (oxLDL) are atherogenic, but their angiogenic effect is controversial; both angiogenic and anti-angiogenic effects have been reported. The angiogenic mechanism of oxLDL is partly understood, but the role of the angiogenic sphingolipid, sphingosine 1-phosphate (S1P), in this process is not known. Thus, we investigated whether S1P is involved in the oxLDL-induced angiogenesis and whether an anti-S1P monoclonal antibody can prevent this effect. EXPERIMENTAL APPROACH Angiogenesis was assessed by capillary tube formation by human microvascular endothelial cells (HMEC-1) cultured on Matrigel and in vivo by the Matrigel plug assay in C57BL/6 mice. KEY RESULTS Human oxLDL exhibited a biphasic angiogenic effect on HMEC-1; low concentrations were angiogenic, higher concentrations were cytotoxic. The angiogenic response to oxLDL was blocked by the sphingosine kinase (SPHK) inhibitor, dimethylsphingosine, by SPHK1-siRNA and by an anti-S1P monoclonal antibody. Moreover, inhibition of oxLDL uptake and subsequent redox signalling by anti-CD36 and anti-LOX-1 receptor antibodies and by N-acetylcysteine, respectively, blocked SPHK1 activation and tube formation. In vivo, in the Matrigel plug assay, low concentrations of human oxLDL or murine oxVLDL also triggered angiogenesis, which was prevented by i.p. injection of the anti-S1P antibody. CONCLUSION AND IMPLICATIONS These data highlight the role of S1P in angiogenesis induced by oxLDL both in HMEC-1 cultured on Matrigel and in vivo in the Matrigel plug model in mice, and demonstrate that the anti-S1P antibody effectively blocks the angiogenic effect of oxLDL. PMID:25176316

  11. S1P prophylaxis mitigates acute hypobaric hypoxia-induced molecular, biochemical, and metabolic disturbances: A preclinical report.

    PubMed

    Chawla, Sonam; Rahar, Babita; Saxena, Shweta

    2016-05-01

    Sphingosine-1-phosphate (S1P) is emerging to have hypoxic preconditioning potential in various preclinical studies. The study aims to evaluate the preclinical preconditioning efficacy of exogenously administered S1P against acute hypobaric hypoxia (HH)-induced pathological disturbances. Male Sprague Dawley rats (200 ± 20 g) were preconditioned with 1, 10, and 100 μg/kg body weight (b.w.) S1P (i.v.) for three consecutive days. On the third day, S1P preconditioned animals, along with hypoxia control animals, were exposed to HH equivalent to 7,620 m (280 mm Hg) for 6 h. Postexposure status of cardiac energy production, circulatory vasoactive mediators, pulmonary and cerebral oxidative damage, and inflammation were assessed. HH exposure led to cardiac energy deficit indicated by low ATP levels and pronounced AMPK activation levels, raised circulatory levels of brain natriuretic peptide and endothelin-1 with respect to total nitrate (NOx), redox imbalance, inflammation, and alterations in NOx levels in the pulmonary and cerebral tissues. These pathological precursors have been routinely reported to be coincident with high-altitude diseases. Preconditioning with S1P, especially 1 µg/kg b.w. dose, was seen to reverse the manifestation of these pathological disturbances. The protective efficacy could be attributed, at least in part, to enhanced activity of cardioprotective protein kinase C and activation of small GTPase Rac1, which led to further induction of hypoxia-adaptive molecular mediators: hypoxia-inducible factor (HIF)-1α and Hsp70. This is a first such report, to the best of our knowledge, elucidating the mechanism of exogenous S1P-mediated HIF-1α/Hsp70 induction. Conclusively, systemic preconditioning with 1 μg/kg b.w. S1P in rats protects against acute HH-induced pathological disturbances. © 2016 IUBMB Life 68(5):365-375, 2016.

  12. Association of constitutive hyperphosphorylation of Hsf1p with a defective ethanol stress response in Saccharomyces cerevisiae sake yeast strains.

    PubMed

    Noguchi, Chiemi; Watanabe, Daisuke; Zhou, Yan; Akao, Takeshi; Shimoi, Hitoshi

    2012-01-01

    Modern sake yeast strains, which produce high concentrations of ethanol, are unexpectedly sensitive to environmental stress during sake brewing. To reveal the underlying mechanism, we investigated a well-characterized yeast stress response mediated by a heat shock element (HSE) and heat shock transcription factor Hsf1p in Saccharomyces cerevisiae sake yeast. The HSE-lacZ activity of sake yeast during sake fermentation and under acute ethanol stress was severely impaired compared to that of laboratory yeast. Moreover, the Hsf1p of modern sake yeast was highly and constitutively hyperphosphorylated, irrespective of the extracellular stress. Since HSF1 allele replacement did not significantly affect the HSE-mediated ethanol stress response or Hsf1p phosphorylation patterns in either sake or laboratory yeast, the regulatory machinery of Hsf1p is presumed to function differently between these types of yeast. To identify phosphatases whose loss affected the control of Hsf1p, we screened a series of phosphatase gene deletion mutants in a laboratory strain background. Among the 29 mutants, a Δppt1 mutant exhibited constitutive hyperphosphorylation of Hsf1p, similarly to the modern sake yeast strains, which lack the entire PPT1 gene locus. We confirmed that the expression of laboratory yeast-derived functional PPT1 recovered the HSE-mediated stress response of sake yeast. In addition, deletion of PPT1 in laboratory yeast resulted in enhanced fermentation ability. Taken together, these data demonstrate that hyperphosphorylation of Hsf1p caused by loss of the PPT1 gene at least partly accounts for the defective stress response and high ethanol productivity of modern sake yeast strains.

  13. Genetic evidence for the roles of the bud-site-selection genes BUD5 and BUD2 in control of the Rsr1p (Bud1p) GTPase in yeast.

    PubMed Central

    Bender, A

    1993-01-01

    Yeast cells normally display either an axial (for MATa or MAT alpha cells) or bipolar (for MATa/alpha cells) pattern of bud-site selection. The RSR1 gene, which was previously identified as a multicopy suppressor of Ts- mutations in the bud-emergence gene CDC24, encodes a GTPase of the Ras family that is required for both budding patterns. Mutations in Rsr1p that presumably block its ability to bind or hydrolyze GTP cause a randomized budding phenotype, suggesting that regulators of Rsr1p will prove to be required for proper bud positioning. The BUD5 gene product is required for proper bud-site selection and contains similarity to GDP-dissociation stimulators (GDS) for Ras-type proteins, suggesting that Bud5p may be a GDS for Rsr1p. Here I report that BUD5 is required for wild-type RSR1, but not for mutationally activated rsr1val12, to serve as a multicopy suppressor of cdc24, indicating that Bud5p functions as a GDS for Rsr1p in vivo. To identify the GAP (GTPase-activating protein) for Rsr1p, a genetic selection was designed based on the observation that mutationally activated rsr1val12, but not wild-type RSR1, can serve as a multicopy suppressor of yeast RAS2(Ts) mutants. Mutants were selected that allowed wild-type RSR1 to act as a multicopy suppressor of RAS2(Ts). Two such mutations proved to be in the BUD2 gene, suggesting that Bud2p functions as a GAP for Rsr1p in vivo. Images Fig. 1 Fig. 2 Fig. 3 PMID:8234337

  14. Chromosome 1p31.1p31.3 Deletion in a Patient with Craniosynostosis, Central Nervous System and Renal Malformation: Case Report and Review of the Literature.

    PubMed

    Rivera-Pedroza, Carlos I; Barraza-García, Jimena; Paumard-Hernández, Beatriz; Nevado, Julian; Orbea-Gallardo, Carlos; Sánchez Del Pozo, Jaime; Heath, Karen E

    2017-01-01

    Interstitial deletions in the short arm of chromosome 1 are infrequent. We report a female with a 1p31.1p31.3 deletion and cloverleaf skull, who presented with renal and central nervous system malformations, cleft palate, severe ocular anomalies, and cutis laxa, in addition to the previously described clinical data present in other cases with deletions encompassing this region, such as developmental delay, seizures, round face with a prominent nose, micro/retrognathia, half-opened mouth, short neck, hand/foot malformations, hernia, congenital heart malformations, and abnormal external genitalia. The deletion spanned ∼18.6 Mb and included a total of 68 OMIM protein coding genes. We have reviewed 17 cases previously described in the literature and in DECIPHER involving the chromosomal region 1p31.1p31.3. Only 3 of these affect the whole region, 9 are partial deletions of this region, and 5 are much smaller deletions. Taking into account the MORBID ID and the haploinsufficiency score of the genes, we go on to propose which genes may explain particular clinical features observed in the patient. IL23R may be responsible for the craniosynostosis, FOXD2 for the renal anomalies, LHX8 for closure defects of the palate, and ST6GALNAC3 for skin anomalies. In summary, we have identified a chromosome 1p31.1p31.3 deletion in a patient with an atypical presentation of craniosynostosis amongst other more typical features observed in individuals with similar deletions.

  15. Vitamin D3 protects against Aβ peptide cytotoxicity in differentiated human neuroblastoma SH- SY5Y cells: A role for S1P1/p38MAPK/ATF4 axis.

    PubMed

    Pierucci, Federica; Garcia-Gil, Mercedes; Frati, Alessia; Bini, Francesca; Martinesi, Maria; Vannini, Eleonora; Mainardi, Marco; Luzzati, Federico; Peretto, Paolo; Caleo, Matteo; Meacci, Elisabetta

    2017-04-01

    Besides its classical function of bone metabolism regulation, 1alpha, 25-dihydroxyvitamin D3 (1,25(OH)2D3), acts on a variety of tissues including the nervous system, where the hormone plays an important role as neuroprotective, antiproliferating and differentiating agent. Sphingolipids are bioactive lipids that play critical and complex roles in regulating cell fate. In the present paper we have investigated whether sphingolipids are involved in the protective action of 1,25(OH)2D3. We have found that 1,25(OH)2D3 prevents amyloid-β peptide (Aβ(1-42)) cytotoxicity both in differentiated SH-SY5Y human neuroblastoma cells and in vivo. In differentiated SH-SY5Y cells, Aβ(1-42) strongly reduces the sphingosine-1-phosphate (S1P)/ceramide (Cer) ratio while 1,25(OH)2D3 partially reverts this effect. 1,25(OH)2D3 reverts also the Aβ(1-42)-induced reduction of sphingosine kinase activity. We have also studied the crosstalk between 1,25(OH)2D3 and S1P signaling pathways downstream to the activation of S1P receptor subtype S1P1. Notably, we found that 1,25(OH)2D3 prevents the reduction of S1P1 expression promoted by Aβ(1-42) and thereby it modulates the downstream signaling leading to ER stress damage (p38MAPK/ATF4). Similar effects were observed by using ZK191784. In addition, chronic treatment with 1,25(OH)2D3 protects from aggregated Aβ(1-42)-induced damage in the CA1 region of the rat hippocampus and promotes cell proliferation in the hippocampal dentate gyrus of adult mice. In conclusion, these results represent the first evidence of the role of 1,25(OH)2D3 and its structural analogue ZK191784 in counteracting the Aβ(1-42) peptide-induced toxicity through the modulation of S1P/S1P1/p38MAPK/ATF4 pathway in differentiated SH-SY5Y cells.

  16. Study of di-pion Bottomonium Transitions and Search for the h_b(1P) State

    SciTech Connect

    Lees, J.P.; Poireau, V.; Tisserand, V.; Garra Tico, J.; Grauges, E.; Martinelli, M.; Milanes, D.A.; Palano, A.; Pappagallo, M.; Eigen, G.; Stugu, B.; Sun, L.; Brown, D.N.; Kerth, L.T.; Kolomensky, Yu.G.; Lynch, G.; Koch, H.; Schroeder, T.; Asgeirsson, D.J.; Hearty, C.; Mattison, T.S.; /British Columbia U. /Brunel U. /Novosibirsk, IYF /UC, Irvine /UC, Riverside /UC, Santa Barbara /UC, Santa Cruz /Caltech /Cincinnati U. /Colorado U. /Colorado State U. /Dortmund U. /Dresden, Tech. U. /Ecole Polytechnique /Edinburgh U. /INFN, Ferrara /INFN, Ferrara /Ferrara U. /INFN, Ferrara /Frascati /INFN, Genoa /Genoa U. /INFN, Genoa /INFN, Genoa /Genoa U. /INFN, Genoa /Indian Inst. Tech., Guwahati /Harvard U. /Harvey Mudd Coll. /Heidelberg U. /Humboldt U., Berlin /Imperial Coll., London /Iowa State U. /Iowa State U. /Johns Hopkins U. /Paris U., VI-VII /LLNL, Livermore /Liverpool U. /Queen Mary, U. of London /Royal Holloway, U. of London /Royal Holloway, U. of London /Louisville U. /Mainz U., Inst. Kernphys. /Manchester U. /Maryland U. /Massachusetts U., Amherst /MIT /McGill U. /INFN, Milan /Milan U. /INFN, Milan /INFN, Milan /Milan U. /Mississippi U. /Montreal U. /INFN, Naples /Naples U. /NIKHEF, Amsterdam /NIKHEF, Amsterdam /Notre Dame U. /Ohio State U. /Oregon U. /INFN, Padua /Padua U. /INFN, Padua /INFN, Padua /Padua U. /Paris U., VI-VII /INFN, Perugia /Perugia U. /INFN, Pisa /Pisa U. /INFN, Pisa /Pisa, Scuola Normale Superiore /INFN, Pisa /Pisa U. /INFN, Pisa /INFN, Pisa /Pisa U. /INFN, Pisa /Princeton U. /INFN, Rome /INFN, Rome /Rome U. /INFN, Rome /INFN, Rome /Rome U. /INFN, Rome /Rostock U. /Rutherford /DAPNIA, Saclay /SLAC /South Carolina U. /Southern Methodist U. /Stanford U., Phys. Dept. /SUNY, Albany /Tel Aviv U. /Tennessee U. /Texas Nuclear Corp., Austin /Texas U., Dallas /INFN, Turin /Turin U. /INFN, Trieste /Trieste U. /Valencia U. /Victoria U. /Warwick U. /Wisconsin U., Madison

    2011-12-09

    We study inclusive di-pion decays using a sample of 108 x 10{sup 6} {Upsilon}(3S) events recorded with the BABAR detector. We search for the decay mode {Upsilon}(3S) {yields} {pi}{sup +}{pi}{sup -}h{sub b} (1P) and find no evidence for the bottomonium spin-singlet state h{sub b}(1P) in the invariant mass distribution recoiling against the {pi}{sup +}{pi}{sup -} system. Assuming the h{sub b}(1P) mass to be 9.900 GeV/c{sup 2}, we measure the upper limit on the branching fraction {Beta}[{Upsilon}(3S) {yields} {pi}{sup +}{pi}{sup -}h{sub b}(1P)] < 1.2 x 10{sup -4}, at 90% confidence level. We also investigate the {chi}{sub bJ}(2P) {yields} {pi}{sup +}{pi}{sup -} {chi}{sub bJ}(1P), {Upsilon}(3S) {yields} {pi}{sup +}{pi}{sup -}{Upsilon}(2S), and {Upsilon}(2S) {yields} {pi}{sup +}{pi}{sup -}{Upsilon}(1S) di-pion transitions and present an improved measurement of the branching fraction of the {Upsilon}(3S) {yields} {pi}{sup +}{pi}{sup -}{Upsilon}(2S) decay and of the {Upsilon}(3S) - {Upsilon}(2S) mass difference.

  17. Monosomy1p36.3 and trisomy 19p13.3 in a child with periventricular nodular heterotopia.

    PubMed

    Descartes, Maria; Mikhail, Fady M; Franklin, Judith C; McGrath, Tony M; Bebin, Martina

    2011-10-01

    Monosomy 1p36 is a clinically recognizable syndrome that is considered to be the most common terminal deletion syndrome. It has characteristic clinical features that include craniofacial dysmorphism, congenital anomalies, hearing deficits, developmental delay, mental retardation, hypotonia, seizures, and brain anomalies. Brain anomalies in patients with 1p36 deletion are frequent but inconsistent. To date, 2 cases with monosomy 1p36 associated with periventricular nodular heterotopia (PNH) have been reported. We report a 2-month-old boy with multiple congenital anomalies; brain magnetic resonance imaging revealed PNH. The first 2 described cases were pure terminal deletions, whereas our patient carried unbalanced translocation due to an adjacent 1 segregation of a balanced maternal translocation, resulting in monosomy 1p36.3 and trisomy 19p13.3 identified by whole-genome array comparative genomic hybridization analysis. Our patient, with a smaller deletion that the 2 previously reported cases, can help narrow the critical region for PNH in association with the 1p36 deletion. Several potential candidate genes are discussed.

  18. Behavioral effects of SQSTM1/p62 overexpression in mice: support for a mitochondrial role in depression and anxiety.

    PubMed

    Seibenhener, M Lamar; Zhao, Ting; Du, Yifeng; Calderilla-Barbosa, Luis; Yan, Jin; Jiang, Jianxiong; Wooten, Marie W; Wooten, Michael C

    2013-07-01

    Affective spectrum and anxiety disorders have come to be recognized as the most prevalently diagnosed psychiatric disorders. Among a suite of potential causes, changes in mitochondrial energy metabolism and function have been associated with such disorders. Thus, proteins that specifically change mitochondrial functionality could be identified as molecular targets for drugs related to treatment for affective spectrum disorders. Here, we report generation of transgenic mice overexpressing the scaffolding and mitophagy related protein Sequestosome1 (SQSTM1/p62) or a single point mutant (P392L) in the UBA domain of SQSTM1/p62. We show that overexpression of SQSTM1/p62 increases mitochondrial energy output and improves transcription factor import into the mitochondrial matrix. These elevated levels of mitochondrial functionality correlate directly with discernible improvements in mouse behaviors related to affective spectrum and anxiety disorders. We also describe how overexpression of SQSTM1/p62 improves spatial learning and long term memory formation in these transgenic mice. These results suggest that SQSTM1/p62 provides an attractive target for therapeutic agents potentially suitable for the treatment of anxiety and affective spectrum disorders.

  19. Refinement of causative genes in monosomy 1p36 through clinical and molecular cytogenetic characterization of small interstitial deletions.

    PubMed

    Rosenfeld, Jill A; Crolla, John A; Tomkins, Susan; Bader, Patricia; Morrow, Bernice; Gorski, Jerome; Troxell, Robin; Forster-Gibson, Cynthia; Cilliers, Deirdre; Hislop, R Gordon; Lamb, Allen; Torchia, Beth; Ballif, Blake C; Shaffer, Lisa G

    2010-08-01

    Monosomy 1p36 is the most common terminal deletion syndrome seen in humans, occurring in approximately 1 in 5,000 live births. Common features include mental retardation, characteristic dysmorphic features, hypotonia, seizures, hearing loss, heart defects, cardiomyopathy, and behavior abnormalities. Similar phenotypes are seen among patients with a variety of deletion sizes, including terminal and interstitial deletions, complex rearrangements, and unbalanced translocations. Consequently, critical regions harboring causative genes for each of these features have been difficult to identify. Here we report on five individuals with 200-823 kb overlapping deletions of proximal 1p36.33, four of which are apparently de novo. They present with features of monosomy 1p36, including developmental delay and mental retardation, dysmorphic features, hypotonia, behavioral abnormalities including hyperphagia, and seizures. The smallest region of deletion overlap is 174 kb and contains five genes; these genes are likely candidates for some of the phenotypic features in monosomy 1p36. Other genes deleted in a subset of the patients likely play a contributory role in the phenotypes, including GABRD and seizures, PRKCZ and neurologic features, and SKI and dysmorphic and neurologic features. Characterization of small deletions is important for narrowing critical intervals and for the identification of causative or candidate genes for features of monosomy 1p36 syndrome.

  20. Monosomy 1p36 - a multifaceted and still enigmatic syndrome: four clinically diverse cases with shared white matter abnormalities.

    PubMed

    Õiglane-Shlik, Eve; Puusepp, Sanna; Talvik, Inga; Vaher, Ulvi; Rein, Reet; Tammur, Pille; Reimand, Tiia; Teek, Rita; Žilina, Olga; Tomberg, Tiiu; Õunap, Katrin

    2014-05-01

    Monosomy 1p36 is the most common subtelomeric deletion syndrome seen in humans. Uniform features of the syndrome include early developmental delay and consequent intellectual disability, muscular hypotonia, and characteristic dysmorphic facial features. The gene-rich nature of the chromosomal band, inconsistent deletion sizes and overlapping clinical features have complicated relevant genotype-phenotype correlations. We describe four patients with isolated chromosome 1p36 deletions. All patients shared white matter abnormalities, allowing us to narrow the critical region for white matter involvement to the deletion size of up to 2.5 Mb from the telomere. We hypothesise that there might be a gene(s) responsible for myelin development in the 1p36 subtelomeric region. Other significant clinical findings were progressive spastic paraparesis, epileptic encephalopathy, various skeletal anomalies, Prader-Willi-like phenotype, neoplastic changes - a haemangioma and a benign skin tumour, and in one case, sleep myoclonus, a clinical entity not previously described in association with 1p36 monosomy. Combined with prior studies, our results suggest that the clinical features seen in monosomy 1p36 have more complex causes than a classical contiguous gene deletion syndrome.

  1. The lipid droplet enzyme Tgl1p hydrolyzes both steryl esters and triglycerides in the yeast, Saccharomyces cerevisiae.

    PubMed

    Jandrositz, Anita; Petschnigg, Julia; Zimmermann, Robert; Natter, Klaus; Scholze, Hubert; Hermetter, Albin; Kohlwein, Sepp D; Leber, Regina

    2005-06-15

    Based on sequence homology to mammalian acid lipases, yeast reading frame YKL140w was predicted to encode a triacylglycerol (TAG) lipase in yeast and was hence named as TGL1, triglyceride lipase 1. A deletion of TGL1, however, resulted in an increase of the cellular steryl ester content. Fluorescently labeled lipid analogs that become covalently linked to the enzyme active site upon catalysis were used to discriminate between the lipase and esterase activities of Tgl1p. Tgl1p preferred single-chain esterase inhibitors over lipase inhibitors in vitro. Under assay conditions optimal for acid lipases, Tgl1p exhibited steryl esterase activity only and lacked any triglyceride lipase activity. In contrast, at pH 7.4, Tgl1p also exhibited TAG lipase activity; however, steryl ester hydrolase activity was still predominant. Tgl1p localized exclusively to lipid droplets which are the intracellular storage compartment of steryl esters and triacylglycerols in the yeast S. cerevisiae. In a tgl1 deletion mutant, the mobilization of steryl esters in vivo was delayed, but not abolished, suggesting the existence of additional enzymes involved in steryl ester mobilization.

  2. Dual-function sRNA encoded peptide SR1P modulates moonlighting activity of B. subtilis GapA

    PubMed Central

    Gimpel, Matthias; Brantl, Sabine

    2016-01-01

    ABSTRACT SR1 is a dual-function sRNA from B. subtilis that acts as a base-pairing regulatory RNA and as a peptide-encoding mRNA. Both functions of SR1 are highly conserved. Previously, we uncovered that the SR1 encoded peptide SR1P binds the glycolytic enzyme GapA resulting in stabilization of gapA mRNA. Here, we demonstrate that GapA interacts with RNases Y and J1, and this interaction was RNA-independent. About 1% of GapA molecules purified from B. subtilis carry RNase J1 and about 2% RNase Y. In contrast to the GapA/RNase Y interaction, the GapA/RNaseJ1 interaction was stronger in the presence of SR1P. GapA/SR1P-J1/Y displayed in vitro RNase activity on known RNase J1 substrates. Moreover, the RNase J1 substrate SR5 has altered half-lives in a ΔgapA strain and a Δsr1 strain, suggesting in vivo functions of the GapA/SR1P/J1 interaction. Our results demonstrate that the metabolic enzyme GapA moonlights in recruiting RNases while GapA bound SR1P promotes binding of RNase J1 and enhances its activity. PMID:27449348

  3. Identification of the Yeast R-SNARE Nyv1p as a Novel Longin Domain-containing Protein

    PubMed Central

    Wen, Wenyu; Chen, Lu; Wu, Hao; Sun, Xin; Zhang, Mingjie

    2006-01-01

    Using nuclear magnetic resonance spectroscopy, we establish that the N-terminal domain of the yeast vacuolar R-SNARE Nyv1p adopts a longin-like fold similar to those of Sec22b and Ykt6p. Nyv1p is sorted to the limiting membrane of the vacuole via the adaptor protein (AP)3 adaptin pathway, and we show that its longin domain is sufficient to direct transport to this location. In contrast, we found that the longin domains of Sec22p and Ykt6p were not sufficient to direct their localization. A YXXΦ-like adaptin-dependent sorting signal (Y31GTI34) unique to the longin domain of Nyv1p mediates interactions with the AP3 complex in vivo and in vitro. We show that amino acid substitutions to Y31GTI34 (Y31Q;I34Q) resulted in mislocalization of Nyv1p as well as reduced binding of the mutant protein to the AP3 complex. Although the sorting of Nyv1p to the limiting membrane of the vacuole is dependent upon the Y31GTI34 motif, and Y31 in particular, our findings with structure-based amino acid substitutions in the mu chain (Apm3p) of yeast AP3 suggest a mechanistically distinct role for this subunit in the recognition of YXXΦ-like sorting signals. PMID:16855025

  4. Coupling between the DEAD-box RNA helicases Ded1p and eIF4A

    PubMed Central

    Gao, Zhaofeng; Putnam, Andrea A; Bowers, Heath A; Guenther, Ulf-Peter; Ye, Xuan; Kindsfather, Audrey; Hilliker, Angela K; Jankowsky, Eckhard

    2016-01-01

    Eukaryotic translation initiation involves two conserved DEAD-box RNA helicases, eIF4A and Ded1p. Here we show that S. cerevisiae eIF4A and Ded1p directly interact with each other and simultaneously with the scaffolding protein eIF4G. We delineate a comprehensive thermodynamic framework for the interactions between Ded1p, eIF4A, eIF4G, RNA and ATP, which indicates that eIF4A, with and without eIF4G, acts as a modulator for activity and substrate preferences of Ded1p, which is the RNA remodeling unit in all complexes. Our results reveal and characterize an unexpected interdependence between the two RNA helicases and eIF4G, and suggest that Ded1p is an integral part of eIF4F, the complex comprising eIF4G, eIF4A, and eIF4E. DOI: http://dx.doi.org/10.7554/eLife.16408.001 PMID:27494274

  5. S1P-Dependent Trafficking of Intracellular Yersinia pestis through Lymph Nodes Establishes Buboes and Systemic Infection

    PubMed Central

    St. John, Ashley L.; Ang, W.X. Gladys; Huang, Min-Nung; Kunder, Christian A.; Chan, Elizabeth W.; Gunn, Michael D.; Abraham, Soman N.

    2015-01-01

    SUMMARY Pathologically swollen lymph nodes (LNs), or buboes, characterize Yersinia pestis infection, yet how they form and function is unknown. We report that colonization of the draining LN (dLN) occurred due to trafficking of infected dendritic cells and monocytes in temporally distinct waves in response to redundant chemotactic signals, including through CCR7, CCR2, and sphingosine-1-phospate (S1P) receptors. Retention of multiple subsets of phagocytes within peripheral LNs using the S1P receptor agonist FTY720 or S1P1-specific agonist SEW2871 increased survival, reduced colonization of downstream LNs, and limited progression to transmission-associated septicemic or pneumonic disease states. Conditional deletion of S1P1 in mononuclear phagocytes abolished node-to-node trafficking of infected cells. Thus, Y. pestis-orchestrated LN remodeling promoted its dissemination via host cells through the lymphatic system but can be blocked by prevention of leukocyte egress from DLNs. These findings define a novel trafficking route of mononuclear phagocytes and identify S1P as a therapeutic target during infection. PMID:25238098

  6. Photoaffinity labeling of Ras converting enzyme 1 (Rce1p) using a benzophenone-containing peptide substrate.

    PubMed

    Kyro, Kelly; Manandhar, Surya P; Mullen, Daniel; Schmidt, Walter K; Distefano, Mark D

    2010-08-01

    Isoprenylation is a post-translational modification that increases protein hydrophobicity and helps target certain proteins to membranes. Ras converting enzyme 1 (Rce1p) is an endoprotease that catalyzes the removal of a three residue fragment from the C-terminus of isoprenylated proteins. To obtain structural information about this membrane protein, photoaffinity labeling agents are being prepared and employed. Here, we describe the synthesis of a benzophenone-containing peptide substrate analogue for Rce1p. Using a continuous spectrofluorometric assay, this peptide was shown to be a substrate for Rce1p. Mass spectrometry was performed to confirm the site of cleavage and structure of the processed probe. Photolysis of the biotinylated compound in the presence of membranes containing Rce1p followed by streptavidin pull-down and Western blot analysis indicated that Rce1p had been labeled by the probe. Photolysis in the presence of both the biotinylated, benzophenone-containing probe and a farnesylated peptide competitor reduced the extent of labeling, suggesting that labeling is occurring in the active site.

  7. Photoaffinity Labeling of Ras Converting Enzyme 1 (Rce1p) using a Benzophenone-Containing Peptide Substrate

    PubMed Central

    Kyro, Kelly; Manandhar, Surya P.; Mullen, Daniel; Schmidt, Walter K.; Distefano, Mark D.

    2010-01-01

    Isoprenylation is a post-translational modification that increases protein hydrophobicity and helps target certain proteins to membranes. Ras Converting Enzyme 1 (Rce1p) is an endoprotease that catalyzes the removal of a three residue fragment from the C-terminus of isoprenylated proteins. To obtain structural information about this membrane protein, photoaffinity labeling agents are being prepared and employed. Here, we describe the synthesis of a benzophenone-containing peptide substrate analogue for Rce1p. Using a continuous spectrofluorometric assay, this peptide was shown to be a substrate for Rce1p. Mass spectrometry was performed to confirm the site of cleavage and structure of the processed probe. Photolysis of the biotinylated compound in the presence of membranes containing Rce1p followed by streptavidin pull-down and Western blot analysis indicated that Rce1p had been labeled by the probe. Photolysis in the presence of both the biotinylated, benzophenone-containing probe and a farnesylated peptide competitor reduced the extent of labeling, suggesting that labeling is occurring in the active site. PMID:20619662

  8. GPI anchor attachment is required for Gas1p transport from the endoplasmic reticulum in COP II vesicles.

    PubMed Central

    Doering, T L; Schekman, R

    1996-01-01

    Inositol starvation of auxotrophic yeast interrupts glycolipid biosynthesis and prevents lipid modification of a normally glycosyl phosphatidylinositol (GPI)-linked protein, Gas1p. The unanchored Gas1p precursor undergoes progressive modification in the endoplasmic reticulum (ER), but is not modified by Golgi-specific glycosylation. Starvation-induced defects in anchor assembly and protein processing are rapid, and occur without altered maturation of other proteins. Cells remain competent to manufacture anchor components and to process Gas1p efficiently once inositol is restored. Newly synthesized Gas1p is packaged into vesicles formed in vitro from perforated yeast spheroplasts incubated with either yeast cytosol or the purified Sec proteins (COP II) required for vesicle budding from the ER. In vitro synthesized vesicles produced by inositol-starved membranes do not contain detectable Gas1p. These studies demonstrate that COP II components fulfill the soluble protein requirements for packaging a GPI-anchored protein into ER-derived transport vesicles. However, GPI anchor attachment is required for this packaging to occur. Images PMID:8598201

  9. Behavioral Effects of SQSTM1/p62 Overexpression in Mice: Support for a Mitochondrial Role in Depression and Anxiety

    PubMed Central

    Seibenhener, M. Lamar; Zhao, Ting; Du, Yifeng; Calderilla-Barbosa, Luis; Yan, Jin; Jiang, Jianxiong; Wooten, Marie W.; Wooten, Michael C.

    2013-01-01

    Affective spectrum and anxiety disorders have come to be recognized as the most prevalently diagnosed psychiatric disorders. Among a suite of potential causes, changes in mitochondrial energy metabolism and function have been associated with such disorders. Thus, proteins that specifically change mitochondrial functionality could be identified as molecular targets for drugs related to treatment for affective spectrum disorders. Here, we report generation of transgenic mice overexpressing the scaffolding and mitophagy related protein Sequestosome1 (SQSTM1/p62) or a single point mutant (P392L) in the UBA domain of SQSTM1/p62. We show that overexpression of SQSTM1/p62 increases mitochondrial energy output and improves transcription factor import into the mitochondrial matrix. These elevated levels of mitochondrial functionality correlate directly with discernible improvements in mouse behaviors related to affective spectrum and anxiety disorders. We also describe how overexpression of SQSTM1/p62 improves spatial learning and long term memory formation in these transgenic mice. These results suggest that SQSTM1/p62 provides an attractive target for therapeutic agents potentially suitable for the treatment of anxiety and affective spectrum disorders. PMID:23591541

  10. An Escherichia coli system for assay of F1p site-specific recombination on substrate plasmids.

    PubMed

    Snaith, M R; Kilby, N J; Murray, J A

    1996-11-21

    We have developed an Escherichia coli system for testing the behaviour of plasmids carrying target sites for the F1p site-specific recombinase. The E. coli strain BL-FLP is described, which carries a chromosomally integrated bacteriophage T7 RNA polymerase gene expressed from a lac promoter, and harbours the plasmid pMS40.pMS40 has the features: (i) it carries the FLP recombinase gene under the control of a bacteriophage T7 promoter, (ii) it confers kanamycin resistance, and (iii) it uses an R6K origin of replication; these two latter features make it compatible with most conventional cloning vectors. Substrate plasmids carrying F1p-recognition targets (FRT) are transformed into BL-FLP, and the consequences of F1p-mediated recombination can be analysed after subsequent extraction of plasmid DNA. We show that this system is capable of base-perfect F1p-mediated recombination on plasmid substrates. We also present a corrected sequence of the commonly used F1p substrate plasmid, pNEO beta GAL (O'Gorman et al. (1991) Science 251, 1351-1355).

  11. The Reticulon and Dp1/Yop1p Proteins Form Immobile Oligomers in the Tubular Endoplasmic Reticulum*S⃞

    PubMed Central

    Shibata, Yoko; Voss, Christiane; Rist, Julia M.; Hu, Junjie; Rapoport, Tom A.; Prinz, William A.; Voeltz, Gia K.

    2008-01-01

    We recently identified a class of membrane proteins, the reticulons and DP1/Yop1p, which shape the tubular endoplasmic reticulum (ER) in yeast and mammalian cells. These proteins are highly enriched in the tubular portions of the ER and virtually excluded from other regions. To understand how they promote tubule formation, we characterized their behavior in cellular membranes and addressed how their localization in the ER is determined. Using fluorescence recovery after photobleaching, we found that yeast Rtn1p and Yop1p are less mobile in the membrane than normal ER proteins. Sucrose gradient centrifugation and cross-linking analyses show that they form oligomers. Mutants of yeast Rtn1p, which no longer localize exclusively to the tubular ER or are even totally inactive in inducing ER tubules, are more mobile and oligomerize less extensively. The mammalian reticulons and DP1 are also relatively immobile and can form oligomers. The conserved reticulon homology domain that includes the two membrane-embedded segments is sufficient for the localization of the reticulons to the tubular ER, as well as for their diffusional immobility and oligomerization. Finally, ATP depletion in both yeast and mammalian cells further decreases the mobilities of the reticulons and DP1. We propose that oligomerization of the reticulons and DP1/Yop1p is important for both their localization to the tubular domains of the ER and for their ability to form tubules. PMID:18442980

  12. Nuclear Drosophila CerS Schlank regulates lipid homeostasis via the homeodomain, independent of the lag1p motif.

    PubMed

    Voelzmann, André; Wulf, Anna-Lena; Eckardt, Franka; Thielisch, Melanie; Brondolin, Mirco; Pesch, Yanina-Yasmin; Sociale, Mariangela; Bauer, Reinhard; Hoch, Michael

    2016-04-01

    Drosophila Ceramide Synthase (CerS) Schlank regulates both ceramide synthesis and fat metabolism. Schlank contains a catalytic lag1p motif and, like many CerS in other species, a homeodomain of unknown function. Here, we show that the Drosophila CerS Schlank is imported into the nucleus and requires two nuclear localization signals (NLSs) within its homeodomain and functional Importin-β import machinery. Expression of Schlank variants containing the homeodomain without functional lag1p motif rescued the fat metabolism phenotype of schlank mutants whereas a variant with a mutated NLS site did not rescue. Thus, the homeodomain of Schlank is involved in the regulation of lipid metabolism independent of the catalytic lag1p motif.

  13. FTH1P3, a Novel H-Ferritin Pseudogene Transcriptionally Active, Is Ubiquitously Expressed and Regulated during Cell Differentiation

    PubMed Central

    Di Sanzo, Maddalena; Aversa, Ilenia; Santamaria, Gianluca; Gagliardi, Monica; Panebianco, Mariafranca; Biamonte, Flavia; Zolea, Fabiana; Faniello, Maria Concetta

    2016-01-01

    Ferritin, the major iron storage protein, performs its essential functions in the cytoplasm, nucleus and mitochondria. The variable assembly of 24 subunits of the Heavy (H) and Light (L) type composes the cytoplasmic molecule. In humans, two distinct genes code these subunits, both belonging to complex multigene families. Until now, one H gene has been identified with the coding sequence interrupted by three introns and more than 20 intronless copies widely dispersed on different chromosomes. Two of the intronless genes are actively transcribed in a tissue-specific manner. Herein, we report that FTH1P3, another intronless pseudogene, is transcribed. FTH1P3 transcript was detected in several cell lines and tissues, suggesting that its transcription is ubiquitary, as it happens for the parental ferritin H gene. Moreover, FTH1P3 expression is positively regulated during the cell differentiation process. PMID:26982978

  14. A rationally engineered yeast pyruvyltransferase Pvg1p introduces sialylation-like properties in neo-human-type complex oligosaccharide.

    PubMed

    Higuchi, Yujiro; Yoshinaga, Sho; Yoritsune, Ken-Ichi; Tateno, Hiroaki; Hirabayashi, Jun; Nakakita, Shin-Ichi; Kanekiyo, Miho; Kakuta, Yoshimitsu; Takegawa, Kaoru

    2016-05-19

    Pyruvylation onto the terminus of oligosaccharide, widely seen from prokaryote to eukaryote, confers negative charges on the cell surface and seems to be functionally similar to sialylation, which is found at the end of human-type complex oligosaccharide. However, detailed molecular mechanisms underlying pyruvylation have not been clarified well. Here, we first determined the crystal structure of fission yeast pyruvyltransferase Pvg1p at a resolution of 2.46 Å. Subsequently, by combining molecular modeling with mutational analysis of active site residues, we obtained a Pvg1p mutant (Pvg1p(H168C)) that efficiently transferred pyruvyl moiety onto a human-type complex glycopeptide. The resultant pyruvylated human-type complex glycopeptide recognized similar lectins on lectin arrays as the α2,6-sialyl glycopeptides. This newly-generated pyruvylation of human-type complex oligosaccharides would provide a novel method for glyco-bioengineering.

  15. A rationally engineered yeast pyruvyltransferase Pvg1p introduces sialylation-like properties in neo-human-type complex oligosaccharide

    PubMed Central

    Higuchi, Yujiro; Yoshinaga, Sho; Yoritsune, Ken-ichi; Tateno, Hiroaki; Hirabayashi, Jun; Nakakita, Shin-ichi; Kanekiyo, Miho; Kakuta, Yoshimitsu; Takegawa, Kaoru

    2016-01-01

    Pyruvylation onto the terminus of oligosaccharide, widely seen from prokaryote to eukaryote, confers negative charges on the cell surface and seems to be functionally similar to sialylation, which is found at the end of human-type complex oligosaccharide. However, detailed molecular mechanisms underlying pyruvylation have not been clarified well. Here, we first determined the crystal structure of fission yeast pyruvyltransferase Pvg1p at a resolution of 2.46 Å. Subsequently, by combining molecular modeling with mutational analysis of active site residues, we obtained a Pvg1p mutant (Pvg1pH168C) that efficiently transferred pyruvyl moiety onto a human-type complex glycopeptide. The resultant pyruvylated human-type complex glycopeptide recognized similar lectins on lectin arrays as the α2,6-sialyl glycopeptides. This newly-generated pyruvylation of human-type complex oligosaccharides would provide a novel method for glyco-bioengineering. PMID:27194449

  16. FTH1P3, a Novel H-Ferritin Pseudogene Transcriptionally Active, Is Ubiquitously Expressed and Regulated during Cell Differentiation.

    PubMed

    Di Sanzo, Maddalena; Aversa, Ilenia; Santamaria, Gianluca; Gagliardi, Monica; Panebianco, Mariafranca; Biamonte, Flavia; Zolea, Fabiana; Faniello, Maria Concetta; Cuda, Giovanni; Costanzo, Francesco

    2016-01-01

    Ferritin, the major iron storage protein, performs its essential functions in the cytoplasm, nucleus and mitochondria. The variable assembly of 24 subunits of the Heavy (H) and Light (L) type composes the cytoplasmic molecule. In humans, two distinct genes code these subunits, both belonging to complex multigene families. Until now, one H gene has been identified with the coding sequence interrupted by three introns and more than 20 intronless copies widely dispersed on different chromosomes. Two of the intronless genes are actively transcribed in a tissue-specific manner. Herein, we report that FTH1P3, another intronless pseudogene, is transcribed. FTH1P3 transcript was detected in several cell lines and tissues, suggesting that its transcription is ubiquitary, as it happens for the parental ferritin H gene. Moreover, FTH1P3 expression is positively regulated during the cell differentiation process.

  17. Cell Surface Display and Characterization of Rhizopus oryzae Lipase in Pichia pastoris Using Sed1p as an Anchor Protein.

    PubMed

    Li, Wenqian; Shi, Hao; Ding, Huaihai; Wang, Liangliang; Zhang, Yu; Li, Xun; Wang, Fei

    2015-07-01

    It has been investigated to conduct the surface displaying of lipase from Rhizopus oryzae onto the cells of Pichia pastoris yeast using Sed1p as an anchor protein. A yeast cell surface display plasmid pPICZαA-rol-histag-sed1p was constructed by fusing rol and sed1p gene fragments into the plasmid pPICZαA, followed by introducing recombinant plasmid into P. pastoris cells. Surface display levels were monitored by Western Blot and immunofluorescence microscopy. The activity of displaying lipase obtained from recombinant mutS reached at 60 U/g-dry cell. In addition, the displaying lipase was stable in broad ranges of temperatures and pH, with the optimum temperature at 40 °C and pH 7.5. These results indicate that the P. pastoris displaying lipase may have potential in whole-cell biocatalyst.

  18. Sfh1p, a component of a novel chromatin-remodeling complex, is required for cell cycle progression.

    PubMed Central

    Cao, Y; Cairns, B R; Kornberg, R D; Laurent, B C

    1997-01-01

    Several eukaryotic multiprotein complexes, including the Saccharomyces cerevisiae Snf/Swi complex, remodel chromatin for transcription. In contrast to the Snf/Swi proteins, Sfh1p, a new Snf5p paralog, is essential for viability. The evolutionarily conserved domain of Sfh1p is sufficient for normal function, and Sfh1p interacts functionally and physically with an essential Snf2p paralog in a novel nucleosome-restructuring complex called RSC (for remodels the structure of chromatin). A temperature-sensitive sfh1 allele arrests cells in the G2/M phase of the cell cycle, and the Sfh1 protein is specifically phosphorylated in the G1 phase. Together, these results demonstrate a link between chromatin remodeling and progression through the cell division cycle, providing genetic clues to possible targets for RSC function. PMID:9154831

  19. Mechanism of Folding and Activation of Subtilisin Kexin Isozyme-1 (SKI-1)/Site-1 Protease (S1P).

    PubMed

    da Palma, Joel Ramos; Cendron, Laura; Seidah, Nabil Georges; Pasquato, Antonella; Kunz, Stefan

    2016-01-29

    The proprotein convertase subtilisin kexin isozyme-1 (SKI-1)/site-1 protease (S1P) is implicated in lipid homeostasis, the unfolded protein response, and lysosome biogenesis. The protease is further hijacked by highly pathogenic emerging viruses for the processing of their envelope glycoproteins. Zymogen activation of SKI-1/S1P requires removal of an N-terminal prodomain, by a multistep process, generating the mature enzyme. Here, we uncover a modular structure of the human SKI-1/S1P prodomain and define its function in folding and activation. We provide evidence that the N-terminal AB fragment of the prodomain represents an autonomous structural and functional unit that is necessary and sufficient for folding and partial activation. In contrast, the C-terminal BC fragment lacks a defined structure but is crucial for autoprocessing and full catalytic activity. Phylogenetic analysis revealed that the sequence of the AB domain is highly conserved, whereas the BC fragment shows considerable variation and seems even absent in some species. Notably, SKI-1/S1P of arthropods, like the fruit fly Drosophila melanogaster, contains a shorter prodomain comprised of full-length AB and truncated BC regions. Swapping the prodomain fragments between fly and human resulted in a fully mature and active SKI-1/S1P chimera. Our study suggests that primordial SKI-1/S1P likely contained a simpler prodomain consisting of the highly conserved AB fragment that represents an independent folding unit. The BC region appears as a later evolutionary acquisition, possibly allowing more subtle fine-tuning of the maturation process.

  20. Mechanism of Folding and Activation of Subtilisin Kexin Isozyme-1 (SKI-1)/Site-1 Protease (S1P)*

    PubMed Central

    da Palma, Joel Ramos; Cendron, Laura; Seidah, Nabil Georges; Pasquato, Antonella; Kunz, Stefan

    2016-01-01

    The proprotein convertase subtilisin kexin isozyme-1 (SKI-1)/site-1 protease (S1P) is implicated in lipid homeostasis, the unfolded protein response, and lysosome biogenesis. The protease is further hijacked by highly pathogenic emerging viruses for the processing of their envelope glycoproteins. Zymogen activation of SKI-1/S1P requires removal of an N-terminal prodomain, by a multistep process, generating the mature enzyme. Here, we uncover a modular structure of the human SKI-1/S1P prodomain and define its function in folding and activation. We provide evidence that the N-terminal AB fragment of the prodomain represents an autonomous structural and functional unit that is necessary and sufficient for folding and partial activation. In contrast, the C-terminal BC fragment lacks a defined structure but is crucial for autoprocessing and full catalytic activity. Phylogenetic analysis revealed that the sequence of the AB domain is highly conserved, whereas the BC fragment shows considerable variation and seems even absent in some species. Notably, SKI-1/S1P of arthropods, like the fruit fly Drosophila melanogaster, contains a shorter prodomain comprised of full-length AB and truncated BC regions. Swapping the prodomain fragments between fly and human resulted in a fully mature and active SKI-1/S1P chimera. Our study suggests that primordial SKI-1/S1P likely contained a simpler prodomain consisting of the highly conserved AB fragment that represents an independent folding unit. The BC region appears as a later evolutionary acquisition, possibly allowing more subtle fine-tuning of the maturation process. PMID:26645686

  1. Hypoxia augments outgrowth endothelial cell (OEC) sprouting and directed migration in response to sphingosine-1-phosphate (S1P).

    PubMed

    Williams, Priscilla A; Stilhano, Roberta S; To, Vivian P; Tran, Lyndon; Wong, Kevin; Silva, Eduardo A

    2015-01-01

    Therapeutic angiogenesis provides a promising approach to treat ischemic cardiovascular diseases through the delivery of proangiogenic cells and/or molecules. Outgrowth endothelial cells (OECs) are vascular progenitor cells that are especially suited for therapeutic strategies given their ease of noninvasive isolation from umbilical cord or adult peripheral blood and their potent ability to enhance tissue neovascularization. These cells are recruited to sites of vascular injury or tissue ischemia and directly incorporate within native vascular endothelium to participate in neovessel formation. A better understanding of how OEC activity may be boosted under hypoxia with external stimulation by proangiogenic molecules remains a challenge to improving their therapeutic potential. While vascular endothelial growth factor (VEGF) is widely established as a critical factor for initiating angiogenesis, sphingosine-1-phosphate (S1P), a bioactive lysophospholipid, has recently gained great enthusiasm as a potential mediator in neovascularization strategies. This study tests the hypothesis that hypoxia and the presence of VEGF impact the angiogenic response of OECs to S1P stimulation in vitro. We found that hypoxia altered the dynamically regulated S1P receptor 1 (S1PR1) expression on OECs in the presence of S1P (1.0 μM) and/or VEGF (1.3 nM). The combined stimuli of S1P and VEGF together promoted OEC angiogenic activity as assessed by proliferation, wound healing, 3D sprouting, and directed migration under both normoxia and hypoxia. Hypoxia substantially augmented the response to S1P alone, resulting in ~6.5-fold and ~25-fold increases in sprouting and directed migration, respectively. Overall, this report highlights the importance of establishing hypoxic conditions in vitro when studying ischemia-related angiogenic strategies employing vascular progenitor cells.

  2. Na(+) transport, and the E(1)P-E(2)P conformational transition of the Na(+)/K(+)-ATPase.

    PubMed Central

    Babes, A; Fendler, K

    2000-01-01

    We have used admittance analysis together with the black lipid membrane technique to analyze electrogenic reactions within the Na(+) branch of the reaction cycle of the Na(+)/K(+)-ATPase. ATP release by flash photolysis of caged ATP induced changes in the admittance of the compound membrane system that are associated with partial reactions of the Na(+)/K(+)-ATPase. Frequency spectra and the Na(+) dependence of the capacitive signal are consistent with an electrogenic or electroneutral E(1)P <--> E(2)P conformational transition which is rate limiting for a faster electrogenic Na(+) dissociation reaction. We determine the relaxation rate of the rate-limiting reaction and the equilibrium constants for both reactions at pH 6.2-8.5. The relaxation rate has a maximum value at pH 7.4 (approximately 320 s(-1)), which drops to acidic (approximately 190 s(-1)) and basic (approximately 110 s(-1)) pH. The E(1)P <--> E(2)P equilibrium is approximately at a midpoint position at pH 6.2 (equilibrium constant approximately 0.8) but moves more to the E(1)P side at basic pH 8.5 (equilibrium constant approximately 0.4). The Na(+) affinity at the extracellular binding site decreases from approximately 900 mM at pH 6.2 to approximately 200 mM at pH 8.5. The results suggest that during Na(+) transport the free energy supplied by the hydrolysis of ATP is mainly used for the generation of a low-affinity extracellular Na(+) discharge site. Ionic strength and lyotropic anions both decrease the relaxation rate. However, while ionic strength does not change the position of the conformational equilibrium E(1)P <--> E(2)P, lyotropic anions shift it to E(1)P. PMID:11053130

  3. Chimeric CYP21A1P/CYP21A2 genes identified in Czech patients with congenital adrenal hyperplasia.

    PubMed

    Vrzalová, Zuzana; Hrubá, Zuzana; Hrabincová, Eva Sťahlová; Vrábelová, Slávka; Votava, Felix; Koloušková, Stanislava; Fajkusová, Lenka

    2011-01-01

    Congenital adrenal hyperplasia (CAH) comprises a group of autosomal recessive disorders caused by an enzymatic deficiency which impairs the biosynthesis of cortisol and, in the majority of severe cases, also the biosynthesis of aldosterone. Approximately 95% of all CAH cases are caused by mutations in the steroid 21-hydroxylase gene (CYP21A2). The CYP21A2 gene and its inactive pseudogene (CYP21A1P) are located within the HLA class III region of the major histocompatibility complex (MHC) locus on chromosome 6p21.3. In this study, we describe chimeric CYP21A1P/CYP21A2 genes detected in our patients with 21-hydroxylase deficiency (21OHD). Chimeric CYP21A1P/CYP21A2 genes were present in 171 out of 508 mutated CYP21A2 alleles (33.8%). We detected four types of chimeric CYP21A1P/CYP21A2 genes: three of them have been described previously as CH-1, CH-3, CH-4, and one type is novel. The novel chimeric gene, termed CH-7, was detected in 21.4% of the mutant alleles. Possible causes of CYP21A1P/CYP21A2 formation are associated with 1) high recombination rate in the MHC locus, 2) high recombination rate between highly homologous genes and pseudogenes in the CYP21 gene area, and 3) the existence of chi-like sequences and repetitive minisatellite consensus sequences in CYP21A2 and CYP21A1P which play a role in promoting genetic recombination.

  4. CDC28 phosphorylates Cac1p and regulates the association of chromatin assembly factor I with chromatin.

    PubMed

    Jeffery, Daniel C B; Kakusho, Naoko; You, Zhiying; Gharib, Marlene; Wyse, Brandon; Drury, Erin; Weinreich, Michael; Thibault, Pierre; Verreault, Alain; Masai, Hisao; Yankulov, Krassimir

    2015-01-01

    Chromatin Assembly Factor I (CAF-I) plays a key role in the replication-coupled assembly of nucleosomes. It is expected that its function is linked to the regulation of the cell cycle, but little detail is available. Current models suggest that CAF-I is recruited to replication forks and to chromatin via an interaction between its Cac1p subunit and the replication sliding clamp, PCNA, and that this interaction is stimulated by the kinase CDC7. Here we show that another kinase, CDC28, phosphorylates Cac1p on serines 94 and 515 in early S phase and regulates its association with chromatin, but not its association with PCNA. Mutations in the Cac1p-phosphorylation sites of CDC28 but not of CDC7 substantially reduce the in vivo phosphorylation of Cac1p. However, mutations in the putative CDC7 target sites on Cac1p reduce its stability. The association of CAF-I with chromatin is impaired in a cdc28-1 mutant and to a lesser extent in a cdc7-1 mutant. In addition, mutations in the Cac1p-phosphorylation sites by both CDC28 and CDC7 reduce gene silencing at the telomeres. We propose that this phosphorylation represents a regulatory step in the recruitment of CAF-I to chromatin in early S phase that is distinct from the association of CAF-I with PCNA. Hence, we implicate CDC28 in the regulation of chromatin reassembly during DNA replication. These findings provide novel mechanistic insights on the links between cell-cycle regulation, DNA replication and chromatin reassembly.

  5. CDC28 phosphorylates Cac1p and regulates the association of chromatin assembly factor i with chromatin

    PubMed Central

    Jeffery, Daniel CB; Kakusho, Naoko; You, Zhiying; Gharib, Marlene; Wyse, Brandon; Drury, Erin; Weinreich, Michael; Thibault, Pierre; Verreault, Alain; Masai, Hisao; Yankulov, Krassimir

    2015-01-01

    Chromatin Assembly Factor I (CAF-I) plays a key role in the replication-coupled assembly of nucleosomes. It is expected that its function is linked to the regulation of the cell cycle, but little detail is available. Current models suggest that CAF-I is recruited to replication forks and to chromatin via an interaction between its Cac1p subunit and the replication sliding clamp, PCNA, and that this interaction is stimulated by the kinase CDC7. Here we show that another kinase, CDC28, phosphorylates Cac1p on serines 94 and 515 in early S phase and regulates its association with chromatin, but not its association with PCNA. Mutations in the Cac1p-phosphorylation sites of CDC28 but not of CDC7 substantially reduce the in vivo phosphorylation of Cac1p. However, mutations in the putative CDC7 target sites on Cac1p reduce its stability. The association of CAF-I with chromatin is impaired in a cdc28–1 mutant and to a lesser extent in a cdc7–1 mutant. In addition, mutations in the Cac1p-phosphorylation sites by both CDC28 and CDC7 reduce gene silencing at the telomeres. We propose that this phosphorylation represents a regulatory step in the recruitment of CAF-I to chromatin in early S phase that is distinct from the association of CAF-I with PCNA. Hence, we implicate CDC28 in the regulation of chromatin reassembly during DNA replication. These findings provide novel mechanistic insights on the links between cell-cycle regulation, DNA replication and chromatin reassembly. PMID:25602519

  6. Regulation of mitochondrial translation of the ATP8/ATP6 mRNA by Smt1p.

    PubMed

    Rak, Malgorzata; Su, Chen Hsien; Xu, Jonathan Tong; Azpiroz, Ricardo; Singh, Angela Mohan; Tzagoloff, Alexander

    2016-03-15

    Expression of the mitochondrially encoded ATP6 and ATP8 genes is translationally regulated by F1 ATPase. We report a translational repressor (Smt1p) of the ATP6/8 mRNA that, when mutated, restores translation of the encoded Atp6p and Atp8p subunits of the ATP synthase. Heterozygous smt1 mutants fail to rescue the translation defect, indicating that the mutations are recessive. Smt1p is an intrinsic inner membrane protein, which, based on its sedimentation, has a native size twice that of the monomer. Affinity purification of tagged Smt1p followed by reverse transcription of the associated RNA and PCR amplification of the resultant cDNA with gene-specific primers demonstrated the presence in mitochondria of Smt1p-ATP8/ATP6 and Smt1p-COB mRNA complexes. These results indicate that Smt1p is likely to be involved in translational regulation of both mRNAs. Applying Occam's principle, we favor a mechanistic model in which translation of the ATP8/ATP6 bicistronic mRNA is coupled to the availability of F1 for subsequent assembly of the Atp6p and Atp8p products into the ATP synthase. The mechanism of this regulatory pathway is proposed to entail a displacement of the repressor from the translationally mute Smt1-ATP8/ATP6 complex by F1, thereby permitting the Atp22p activator to interact with and promote translation of the mRNA.

  7. Reduced TORC1 signaling abolishes mitochondrial dysfunctions and shortened chronological lifespan of Isc1p-deficient cells

    PubMed Central

    Teixeira, Vitor; Medeiros, Tânia C.; Vilaça, Rita; Moradas-Ferreira, Pedro; Costa, Vítor

    2014-01-01

    The target of rapamycin (TOR) is an important signaling pathway on a hierarchical network of interacting pathways regulating central biological processes, such as cell growth, stress response and aging. Several lines of evidence suggest a functional link between TOR signaling and sphingolipid metabolism. Here, we report that the TORC1-Sch9p pathway is activated in cells lacking Isc1p, the yeast orthologue of mammalian neutral sphingomyelinase 2. The deletion of TOR1 or SCH9 abolishes the premature aging, oxidative stress sensitivity and mitochondrial dysfunctions displayed by isc1Δ cells and this is correlated with the suppression of the autophagic flux defect exhibited by the mutant strain. The protective effect of TOR1 deletion, as opposed to that of SCH9 deletion, is not associated with the attenuation of Hog1p hyperphosphorylation, which was previously implicated in isc1Δ phenotypes. Our data support a model in which Isc1p regulates mitochondrial