Identification of a novel transcriptional variant in the mouse Sry locus by long-read RNA-seq and conventional RNA-seq
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE151472 | GEO | 2020/10/01
REPOSITORIES: GEO
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Project description:A major event in mammalian male sex determination is the induction of the testis determining factor Sry and its downstream gene Sox9. The current study provides one of the first genome wide analyses of the downstream gene binding targets for SRY and SOX9 to help elucidate the molecular control of Sertoli cell differentiation and testis development. A modified ChIP-Chip analysis using a comparative hybridization was used to identify 71 direct downstream binding targets for SRY and 109 binding targets for SOX9. Interestingly, only 5 gene targets overlapped between SRY and SOX9. In addition to the direct response element binding gene targets, a large number of atypical binding gene targets were identified for both SRY and SOX9. Bioinformatic analysis of the downstream binding targets identified gene networks and cellular pathways potentially involved in the induction of Sertoli cell differentiation and testis development. The specific DNA sequence binding site motifs for both SRY and SOX9 were identified. Observations provide insights into the molecular control of male gonadal sex determination. The current study provides one of the first genome wide analyses of the downstream gene binding targets for SRY and SOX9 to help elucidate the molecular control of Sertoli cell differentiation and testis development. At embryonic day 13 (E13) of pregnancy rats were euthanized and embryonic gonads were collected for chromatin. A modified ChIP-Chip analysis using a comparative hybridization was used to identify direct downstream binding targets for SRY and for SOX9. Then, bioinformatic analysis of the downstream binding targets was done to identify gene networks and cellular pathways that are potentially involved in the induction of Sertoli cell differentiation and testis development.
2014-12-05 | E-GEOD-63854 | biostudies-arrayexpress
Project description:The cascade of molecular events involved in mammalian sex determination has been shown to involve the SRY gene, but specific downstream events have eluded researchers for decades. The current study identifies one of the first direct downstream targets of the male sex-determining factor SRY as the basic-helix-loop-helix (bHLH) transcription factor TCF21. SRY was found to directly associate with the Tcf21 promoter SRY/SOX9 response element both in vitro and in vivo during male sex determination. TCF21 was found to promote an in vitro sex reversal of embryonic ovarian cells to promote precursor Sertoli cell differentiation. Therefore, SRY acts directly on the Tcf21 promoter to, in part, initiate a cascade of events associated with Sertoli cell differentiation and embryonic testis development.
2011-04-25 | GSE24666 | GEO
Project description:The mammalian sex-determining gene Sry induces male development. Through analyzing transcriptome of pre-Sertoli cells, we identified a novel sequence transcribed within palindromic sequence surrounding Sry. To examine whether this sequence contains transcription start site, Cap analysis gene expression (CAGE) -seq analysis was performed, in which putative transcriptional start sites can be identified by sequencing the 3’ end of cDNA corresponding to 5’ end of RNA. CAGE seq revealed that Cap site was absent within the novel sequence, whereas Sry transcription start site found to locate in minus strand at chrY:2,663,800-2,663,900. Combining the result from long-read RNA seq, we concluded that this sequence is the second exon of mouse Sry.
2020-10-01 | GSE151473 | GEO
Project description:The cascade of molecular events involved in mammalian sex determination has been shown to involve the SRY gene, but specific downstream events have eluded researchers for decades. The current study identifies one of the first direct downstream targets of the male sex-determining factor SRY as the basic-helix-loop-helix (bHLH) transcription factor TCF21. SRY was found to directly associate with the Tcf21 promoter SRY/SOX9 response element both in vitro and in vivo during male sex determination. TCF21 was found to promote an in vitro sex reversal of embryonic ovarian cells to promote precursor Sertoli cell differentiation. Therefore, SRY acts directly on the Tcf21 promoter to, in part, initiate a cascade of events associated with Sertoli cell differentiation and embryonic testis development. We used microarrays to determine genes whose expression was stimulated in rat E13 ovarian cell sub-cultures in the presence of a pCMV-myc-expression plasmid over-expressing the Sry, Tsf21, and/or Tcf12 (Reb-alfa) genes. RNA samples from the control group (untreated E13 rat ovarian cells) are compared to RNA from 4 groups of treated E13 rat ovarian cells: 1) Sry overexpressing, 2) Tcf21 overexpressing, 3) Tcf12 (Reb alfa) overexpressing, and 4) Tcf21 + Tcf12 (Reb-alfa) overexpressing. Untreated E13 testis cell sub-cultures were also analyzed. 3 biological replicates each group.
2011-04-25 | E-GEOD-24666 | biostudies-arrayexpress
Project description:A major event in mammalian male sex determination is the induction of the testis determining factor Sry and its downstream gene Sox9. The current study provides one of the first genome wide analyses of the downstream gene binding targets for SRY and SOX9 to help elucidate the molecular control of Sertoli cell differentiation and testis development. A modified ChIP-Chip analysis using a comparative hybridization was used to identify 71 direct downstream binding targets for SRY and 109 binding targets for SOX9. Interestingly, only 5 gene targets overlapped between SRY and SOX9. In addition to the direct response element binding gene targets, a large number of atypical binding gene targets were identified for both SRY and SOX9. Bioinformatic analysis of the downstream binding targets identified gene networks and cellular pathways potentially involved in the induction of Sertoli cell differentiation and testis development. The specific DNA sequence binding site motifs for both SRY and SOX9 were identified. Observations provide insights into the molecular control of male gonadal sex determination. The current study provides one of the first genome wide analyses of the downstream gene binding targets for SRY and SOX9 to help elucidate the molecular control of Sertoli cell differentiation and testis development. At embryonic day 13 (E13) of pregnancy rats were euthanized and embryonic gonads were collected for chromatin. A modified ChIP-Chip analysis using a comparative hybridization was used to identify direct downstream binding targets for SRY and for SOX9. Then, bioinformatic analysis of the downstream binding targets was done to identify gene networks and cellular pathways that are potentially involved in the induction of Sertoli cell differentiation and testis development.
2014-12-05 | E-GEOD-63855 | biostudies-arrayexpress
Project description:A critical transcription factor required for mammalian male sex determination is SRY (sex determining region on the Y chromosome). The expression of SRY in precursor Sertoli cells is one of the initial events in testis development. The current study was designed to determine the impact of environmentally induced epigenetic transgenerational inheritance on SRY during gonadal sex determination in the male. The agricultural fungicide vinclozolin and vehicle control (DMSO) exposed gestating females (F0 generation) during gonadal sex determination promoted the transgenerational inheritance of differential DNA methylation in sperm of the F3 generation (great grand-offspring). The fetal gonads in F3 generation males were used to identify potential alterations in SRY binding sites in the developing Sertoli cells. Chromatin immunoprecipitation with an SRY antibody followed by genome-wide promoter tiling array (ChIP-Chip) was used to identify alterations in SRY binding. A total of 81 adjacent oligonucleotide sites and 173 single oligo SRY binding sites were identified to be altered transgenerationally in the Sertoli cell vinclozolin lineage F3 generation males. Observations demonstrate the majority of the previously identified normal SRY binding sites were not altered and the altered SRY binding sites were novel and new additional sites. The chromosomal locations, gene associations and potentially modified cellular pathways were investigated. In summary, environmentally induces epigenetic transgenerational inheritance of germline epimutations appears to alter the cellular differentiation and development of the precursor Sertoli cell SRY binding during gonadal sex determination that influence the developmental origins of adult onset testis disease. The experimental design involved the intraperitoneal exposure of gestating female rats to vinclozolin or a vehicle control (dimethyl sulfoxide, DMSO) transiently from embryonic days E8-E14. Sister littermates were divided into control and vinclozolin treatment groups and mated to similar males to minimize the genetic variation between the control and vinclozolin lineages. Sufficient females were used so no inbreeding (sibling or cousin) occurred in any generation. The F1 generation was bred within the lineage to generate the F2 generation and these F2 generation bred to generate the F3 generation. The only exposure was the F0 generation female. The F3 generation control and vinclozolin lineage embryonic day 13 (E13) embryos were collected and the gonads micro-dissected and then sexed with an SRY PCR protocol. The male gonads were pooled from a minimum of three different litters and the pools used to collect DNA. Three different experiments were performed to collect 3 control and vinclozolin E13 F3 generation testis pools, each with different animals (n=25 gonads/pool). The chromatin DNA (not denatured) from each pool was fragmented and used in an SRY chromatin immunoprecipitation (ChIP) procedure for each pool separately. The control and vinclozolin SRY ChIP DNA were paired for a competitive hybridization on a genome-wide promoter tiling array (ChIP-Chip) assay. The hybridization data obtained was used to identify the SRY binding sites that were different in the F3 generation vinclozolin versus control lineage in E13 testis.
2015-08-28 | E-GEOD-72469 | biostudies-arrayexpress
Project description:A critical transcription factor required for mammalian male sex determination is SRY (sex determining region on the Y chromosome). The expression of SRY in precursor Sertoli cells is one of the initial events in testis development. The current study was designed to determine the impact of environmentally induced epigenetic transgenerational inheritance on SRY during gonadal sex determination in the male. The agricultural fungicide vinclozolin and vehicle control (DMSO) exposed gestating females (F0 generation) during gonadal sex determination promoted the transgenerational inheritance of differential DNA methylation in sperm of the F3 generation (great grand-offspring). The fetal gonads in F3 generation males were used to identify potential alterations in SRY binding sites in the developing Sertoli cells. Chromatin immunoprecipitation with an SRY antibody followed by genome-wide promoter tiling array (ChIP-Chip) was used to identify alterations in SRY binding. A total of 81 adjacent oligonucleotide sites and 173 single oligo SRY binding sites were identified to be altered transgenerationally in the Sertoli cell vinclozolin lineage F3 generation males. Observations demonstrate the majority of the previously identified normal SRY binding sites were not altered and the altered SRY binding sites were novel and new additional sites. The chromosomal locations, gene associations and potentially modified cellular pathways were investigated. In summary, environmentally induces epigenetic transgenerational inheritance of germline epimutations appears to alter the cellular differentiation and development of the precursor Sertoli cell SRY binding during gonadal sex determination that influence the developmental origins of adult onset testis disease.
2015-08-28 | GSE72469 | GEO
Project description:A major event in mammalian male sex determination is the induction of the testis determining factor Sry and its downstream gene Sox9. The current study provides one of the first genome wide analyses of the downstream gene binding targets for SRY and SOX9 to help elucidate the molecular control of Sertoli cell differentiation and testis development. A modified ChIP-Chip analysis using a comparative hybridization was used to identify 71 direct downstream binding targets for SRY and 109 binding targets for SOX9. Interestingly, only 5 gene targets overlapped between SRY and SOX9. In addition to the direct response element binding gene targets, a large number of atypical binding gene targets were identified for both SRY and SOX9. Bioinformatic analysis of the downstream binding targets identified gene networks and cellular pathways potentially involved in the induction of Sertoli cell differentiation and testis development. The specific DNA sequence binding site motifs for both SRY and SOX9 were identified. Observations provide insights into the molecular control of male gonadal sex determination.
2014-12-05 | GSE63854 | GEO
Project description:Sry is sufficient to induce testis formation and subsequent male development of internal and external genitalia in chromosomally female mice and humans. In XX sex-reversed males such as XX/Sry-transgenic (XX/Sry) mice, however, testicular germ cells always disappear soon after birth due to germ cell autonomous defects. Therefore, it remains unclear whether or not Sry alone is sufficient to induce a fully functional testicular soma capable of supporting complete spermatogenesis in the XX body. Here we demonstrated that the testicular somatic environment of XX/Sry males is defective in the later phases of spermatogenesis. Spermatogonial transplantation analyses using XX/Sry male mice revealed that donor XY spermatogonia are capable of proliferating, entering meiosis and differentiating into the round spermatid stage. XY donor-derived round spermatids, however, were frequently detached from the XX/Sry seminiferous epithelia and underwent cell death, thereby preventing further progress beyond the elongated spermatid stage. In contrast, immature XY seminiferous tubule segments transplanted under XX/Sry testis capsules clearly displayed proper differentiation into elongated spermatids in the transplanted XY donor tubules. Microarray analysis of seminiferous tubules isolated from XX/Sry testes confirmed missing expression of several Y-linked genes and alterations in the expression profile of genes associated with spermatogenesis. Therefore, our findings indicate dysfunction of the somatic tubule components, probably Sertoli cells, of XX/Sry testes, supporting our hypothesis that Sry alone is insufficient to induce a fully functional Sertoli cell in XX mice. Experiment Overall Design: Whole testes and seminiferous tubules of XX/Sry and W/Wv males were used for microarray expression analysis using the Affymetrix GeneChip system (Affymetrix, CA). In order to isolate the seminiferous tubules, the tunica was carefully removed from the testes which were then incubated in the medium with 5 mg/ml collagnease at 37oC for 40 min. The remaining seminiferous tubules were washed several times with PBS using a 70-ºm cell strainer to remove interstitial cells. After total RNA was extracted using a RNeasy Mini Kit (Qiagen, Germantown, MD), double-stranded cDNA and biotin-labeled cRNA were synthesized using One-Cycle cDNA Synthesis and IVT Labeling kits (Affymetrix, CA), respectively. Twenty micrograms of fragmented biotin-labeled cRNA was hybridized to the Affymetrix Mouse Expression Array MOE 430A for 16 hr at 45oC. The chips were washed, stained, and then scanned with the GeneArray Scanner (Hewlett Packard, CA) in accordance with the manufacturer's standard protocols. Finally, the microarray data were analyzed using Microarray Suite ver. 5.0 (Affymetrix). Differential expression was defined as a difference of 2-fold or more in both whole testis and seminiferous tubule samples between two recipient males. Mouse 430A Affymetrix Genome Array IDs were used to query the NetAffx data mining tool for gene annotations.
2008-06-13 | E-GEOD-5319 | biostudies-arrayexpress
Project description:Sry is sufficient to induce testis formation and subsequent male development of internal and external genitalia in chromosomally female mice and humans. In XX sex-reversed males such as XX/Sry-transgenic (XX/Sry) mice, however, testicular germ cells always disappear soon after birth due to germ cell autonomous defects. Therefore, it remains unclear whether or not Sry alone is sufficient to induce a fully functional testicular soma capable of supporting complete spermatogenesis in the XX body. Here we demonstrated that the testicular somatic environment of XX/Sry males is defective in the later phases of spermatogenesis. Spermatogonial transplantation analyses using XX/Sry male mice revealed that donor XY spermatogonia are capable of proliferating, entering meiosis and differentiating into the round spermatid stage. XY donor-derived round spermatids, however, were frequently detached from the XX/Sry seminiferous epithelia and underwent cell death, thereby preventing further progress beyond the elongated spermatid stage. In contrast, immature XY seminiferous tubule segments transplanted under XX/Sry testis capsules clearly displayed proper differentiation into elongated spermatids in the transplanted XY donor tubules. Microarray analysis of seminiferous tubules isolated from XX/Sry testes confirmed missing expression of several Y-linked genes and alterations in the expression profile of genes associated with spermatogenesis. Therefore, our findings indicate dysfunction of the somatic tubule components, probably Sertoli cells, of XX/Sry testes, supporting our hypothesis that Sry alone is insufficient to induce a fully functional Sertoli cell in XX mice. Keywords: comparative genomic hybridization
2007-07-15 | GSE5319 | GEO