Methylation of the Gpat2 promoter regulates transient expression during mouse spermatogenesis

Spermatogenesis is a highly regulated process that involves both mitotic and meiotic divisions, as well as cellular differentiation to yield mature spermatozoa from undifferentiated germinal stem cells. Although Gpat2 was originally annotated as a glycerol-3-phospate acyltransferase by sequence homology to Gpat1 , GPAT2 is highly expressed in testis but not in lipogenic tissues and is not up-regulated during adipocyte differentiation. New data show that GPAT2 is required for the synthesis of piRNAs, a group of small RNAs that protect the germ cell genome from retrotransposable elements. In order to understand the relationship between GPAT2 and its role in the testis, we focused on Gpat2 expression during the first wave of mouse spermatogenesis. Gpat2 expression was analyzed by qPCR, in situ hybridization, immunohistochemistry and Western blot. Gpat2 mRNA content and protein expression were maximal at 15 dpp and restricted to pachytene spermatocytes. To achieve this transient expression, both epigenetic mechanisms and trans-acting factors are involved. In vitro assays showed that Gpat2 expression correlates with DNA demethylation and histone acetylation and that it is up-regulated by retinoic acid. Epigenetic regulation by DNA methylation was confirmed in vivo in germ cells by bisulfite sequencing of the Gpat2 promoter. Consistent with the initiation of meiosis at 11 dpp, methylation decreased dramatically. Thus, Gpat2 is expressed at a specific stage of spermatogenesis, consistent with piRNA synthesis and meiosis I prophase, and its on-off expression pattern responds predominantly to epigenetic modifications.Fil: Garcia Fabiani, Maria Belen. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata ; ArgentinaFil: Montanaro, Mauro Aldo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata ; ArgentinaFil: Lacunza, Ezequiel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Médicas. Centro de Investigaciones Inmunológicas Básicas y Aplicadas; ArgentinaFil: Cattaneo, Elizabeth Renee. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata ; ArgentinaFil: Coleman, Rosalind A.. University of North Carolina; Estados UnidosFil: Pellon Maison, Magali. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata ; ArgentinaFil: Gonzalez-Baró; MR. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata ; Argentin

The Magic Flute

Program listing performers and works performe

Emergencies within hospital wards: An observational study of the non-technical skills of medical emergency teams

Background Medical emergency teams are essential in responding to acute deterioration of patients in hospitals, requiring both clinical and non-technical skills. This study aims to assess the non-technical skills of medical emergency teams during hospital ward emergencies and explore team members perceptions and experiences of the use non-technical skills during medical emergencies. Methods A multi-methods study was conducted in two phases. During phase one observation and assessment of non-technical skills used in medical emergencies using the Team Emergency Assessment Measure (TEAM™) was conducted; and in the phase two in-depth interviews were undertaken with medical emergency team members. Results Based on 20 observations, mean TEAM™ ratings for non-technical skill domains were: ‘leadership’ 5.0 out of 8 (±2.0); ‘teamwork’ 21.6 out of 28 (±3.6); and ‘task management’ 6.5 out of 8 (±1.4). The mean ‘global’ score was 7.5 out of 10 (±1.5). The qualitative findings identified three areas, ‘individual’, ‘team’ and ‘other’ contributing factors, which impacted upon the non-technical skills of medical emergency teams. Conclusion Non-technical skills of hospital medical emergency teams differ, and the impact of the skill mix on resuscitation outcomes was recognised by team members. These findings emphasize the importance non-technical skills in resuscitation training and well-developed processes for medical emergency teams

Glycerol-3-phosphate acyltransferase-2 is expressed in spermatic germ cells and incorporates arachidonic acid into triacylglycerols

Background: De novo glycerolipid synthesis begins with the acylation of glycerol-3 phosphate catalyzed by glycerol-3-phosphate acyltransferase (GPAT). In mammals, at least four GPAT isoforms have been described, differing in their cell and tissue locations and sensitivity to sulfhydryl reagents. In this work we show that mitochondrial GPAT2 overexpression in CHO-K1 cells increased TAG content and both GPAT and AGPAT activities 2-fold with arachidonoyl-CoA as a substrate, indicating specificity for this fatty acid. Methods and Results: Incubation of GPAT2-transfected CHO-K1 cells with [1-14C]arachidonate for 3 h increased incorporation of [14C]arachidonate into TAG by 40%. Consistently, arachidonic acid was present in the TAG fraction of cells that overexpressed GPAT2, but not in control cells, corroborating GPAT2's role in synthesizing TAG that is rich in arachidonic acid. In rat and mouse testis, Gpat2 mRNA was expressed only in primary spermatocytes; the protein was also detected in late stages of spermatogenesis. During rat sexual maturation, both the testicular TAG content and the arachidonic acid content in the TAG fraction peaked at 30 d, matching the highest expression of Gpat2 mRNA and protein. Conclusions: These results strongly suggest that GPAT2 expression is linked to arachidonoyl-CoA incorporation into TAG in spermatogenic germ cells.Facultad de Ciencias Médica

Methylation of the <i>Gpat2</i> promoter regulates transient expression during mouse spermatogenesis

Spermatogenesis is a highly regulated process that involves both mitotic and meiotic divisions, as well as cellular differentiation to yield mature spermatozoa from undifferentiated germinal stem cells. Although Gpat2 was originally annotated as a glycerol-3-phospate acyltransferase by sequence homology to Gpat1, GPAT2 is highly expressed in testis but not in lipogenic tissues and is not up-regulated during adipocyte differentiation. New data show that GPAT2 is required for the synthesis of piRNAs, a group of small RNAs that protect the germ cell genome from retrotransposable elements. In order to understand the relationship between GPAT2 and its role in the testis, we focused on Gpat2 expression during the first wave of mouse spermatogenesis. Gpat2 expression was analyzed by qPCR, in situ hybridization, immunohistochemistry and Western blot. Gpat2 mRNA content and protein expression were maximal at 15 dpp and restricted to pachytene spermatocytes. To achieve this transient expression, both epigenetic mechanisms and trans-acting factors are involved. In vitro assays showed that Gpat2 expression correlates with DNA demethylation and histone acetylation and that it is up-regulated by retinoic acid. Epigenetic regulation by DNA methylation was confirmed in vivo in germ cells by bisulfite sequencing of the Gpat2 promoter. Consistent with the initiation of meiosis at 11 dpp, methylation decreased dramatically. Thus, Gpat2 is expressed at a specific stage of spermatogenesis, consistent with piRNA synthesis and meiosis I prophase, and its on-off expression pattern responds predominantly to epigenetic modifications.Facultad de Ciencias MédicasInstituto de Investigaciones Bioquímicas de La PlataCentro de Investigaciones Inmunológicas Básicas y Aplicada

Glicerol-3-fosfato aciltransferasa 2: su relación con la espermatogénesis y la proliferación celular

La enzima glicerol-3-fosfato aciltransferasa (GPAT) cataliza el primer paso y limitante en la síntesis de novo de glicerolípidos celulares. En mamíferos han sido descriptas al menos cuatro isoformas con diferentes localizaciones celulares y parámetros cinéticos. La isoforma mitocondrial 1 (GPAT1) ha sido extensamente estudiada y se ha demostrado que participa en la síntesis de triacilglicéridos (TAG) en hígado de rata. La isoforma mitocondrial 2 (GPAT2) presenta homología de secuencia con GPAT1, pero se expresa preferencialmente en testículo de rata y no tiene preferencia por los ácidos grasos saturados, por lo cual hipotetizamos que estaría involucrada en la síntesis de TAG ricos en ácidos grasos insaturados, moléculas que actúan como reservorios de estos ácidos grasos necesarios para la espermatogénesis. Nuestro objetivo es determinar si la función de GPAT2 (isoforma mitocondrial 2) está asociada a la síntesis celular de TAG (triacilglicéridos), y si su expresión se correlaciona con el inicio de la espermatogénesis.Facultad de Ciencias Médica

Staying active, staying strong : Pilot evaluation of a once-weekly, community-based strength training program for older adults

Issue addressed: Little is known about the effectiveness of once-weekly strength training programs for older adults based in community settings. This pilot study evaluated such a program to assess changes in the functional fitness of participants. Methods: A pre-test/post-test within subjects study design was used with new participants in the 10-week Staying Active, Staying Strong (SASS) program (all aged 50+ years). The Seniors Fitness Test (SFT) and SF-36 were used to assess functional fitness and health-related quality of life respectively. Perception of physical ability was assessed using a study-specific questionnaire. Pre- and post-test SFT and SF-36 scores were compared using paired t-tests. Frequency of responses was used to describe participant perceptions. Results: 110 evaluation participants (mean age 68.2 years; 85% female), 49% of those who completed the pre-test, also completed the post-test. Evaluation participants significantly improved their strength (assessed using arm curls and sit-to-stand); endurance (two-minute step test); flexibility (sit and reach, back scratch); and agility/dynamic balance (eight-foot up and go). SF-36 physical-functioning domain scores also significantly improved. Most participants reported improved strength, fitness, mobility, general well-being and confidence in performing daily activities. Conclusion: Weekly, community-based strength training programs show promise in improving the functional capacity, including the strength, of older adults. More thorough evaluation is now required to confirm these findings

Glycerol-3-phosphate acyltranferase-2 behaves as a cancer testis gene and promotes growth and tumorigenicity of the breast cancer MDA-MB-231 cell line

The de novo synthesis of glycerolipids in mammalian cells begins with the acylation of glycerol-3-phosphate, catalyzed by glycerol-3-phosphate acyltransferase (GPAT). GPAT2 is a mitochondrial isoform primarily expressed in testis under physiological conditions. Because it is aberrantly expressed in multiple myeloma, it has been proposed as a novel cancer testis gene. Using a bioinformatics approach, we found that GPAT2 is highly expressed in melanoma, lung, prostate and breast cancer, and we validated GPAT2 expression at the protein level in breast cancer by immunohistochemistry. In this case GPAT2 expression correlated with a higher histological grade. 5-Aza-2′ deoxycytidine treatment of human cells lines induced GPAT2 expression suggesting epigenetic regulation of gene expression. In order to evaluate the contribution of GPAT2 to the tumor phenotype, we silenced its expression in MDA-MB-231 cells. GPAT2 knockdown diminished cell proliferation, anchorage independent growth, migration and tumorigenicity, and increased staurosporine-induced apoptosis. In contrast, GPAT2 over-expression increased cell proliferation rate and resistance to staurosporine-induced apoptosis. To understand the functional role of GPAT2, we performed a co-expression analysis in mouse and human testis and found a significant association with semantic terms involved in cell cycle, DNA integrity maintenance, piRNA biogenesis and epigenetic regulation. Overall, these results indicate the GPAT2 would be directly associated with the control of cell proliferation. In conclusion, we confirm GPAT2 as a cancer testis gene and that its expression contributes to the tumor phenotype of MDA-MB-231 cells.Instituto de Investigaciones Bioquímicas de La Plat

Glycerol-3-phosphate acyltransferase-2 is expressed in spermatic germ cells and incorporates arachidonic acid into triacylglycerols

Background: De novo glycerolipid synthesis begins with the acylation of glycerol-3 phosphate catalyzed by glycerol-3-phosphate acyltransferase (GPAT). In mammals, at least four GPAT isoforms have been described, differing in their cell and tissue locations and sensitivity to sulfhydryl reagents. In this work we show that mitochondrial GPAT2 overexpression in CHO-K1 cells increased TAG content and both GPAT and AGPAT activities 2-fold with arachidonoyl-CoA as a substrate, indicating specificity for this fatty acid. Methods and Results: Incubation of GPAT2-transfected CHO-K1 cells with [1-14C]arachidonate for 3 h increased incorporation of [14C]arachidonate into TAG by 40%. Consistently, arachidonic acid was present in the TAG fraction of cells that overexpressed GPAT2, but not in control cells, corroborating GPAT2's role in synthesizing TAG that is rich in arachidonic acid. In rat and mouse testis, Gpat2 mRNA was expressed only in primary spermatocytes; the protein was also detected in late stages of spermatogenesis. During rat sexual maturation, both the testicular TAG content and the arachidonic acid content in the TAG fraction peaked at 30 d, matching the highest expression of Gpat2 mRNA and protein. Conclusions: These results strongly suggest that GPAT2 expression is linked to arachidonoyl-CoA incorporation into TAG in spermatogenic germ cells.Facultad de Ciencias Médica

The Astropy Problem

The Astropy Project (http://astropy.org) is, in its own words, "a community effort to develop a single core package for Astronomy in Python and foster interoperability between Python astronomy packages." For five years this project has been managed, written, and operated as a grassroots, self-organized, almost entirely volunteer effort while the software is used by the majority of the astronomical community. Despite this, the project has always been and remains to this day effectively unfunded. Further, contributors receive little or no formal recognition for creating and supporting what is now critical software. This paper explores the problem in detail, outlines possible solutions to correct this, and presents a few suggestions on how to address the sustainability of general purpose astronomical software