Identifying splicing targets of CLAMP by mRNA-sequencing
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA909765 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR22561728_1.fastq.gz | Fastqsanger.gz | |||
| SRR22561728_2.fastq.gz | Fastqsanger.gz | |||
| SRR22561729_1.fastq.gz | Fastqsanger.gz | |||
| SRR22561729_2.fastq.gz | Fastqsanger.gz | |||
| SRR22561730_1.fastq.gz | Fastqsanger.gz |
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Project description:CLAMP is a GA-repeat motif binding transcription factor that regulates gene expression. We hypothesized that CLAMP also regulates RNA processing, specifically alternative splicing that occurs co-transcriptionally because CLAMP is bound to intronic regions that are rich in polypyrimidine tracts which contain GA-rich sequences. Furthermore, GA-rich repeat sequences are thought to have evolved from polypyrimidine tracts that regulate splicing. Also, MALDI-mass spectrometry data identifying putative CLAMP interactors found association with 33 RNA binding proteins, including 6 that regulate alternative splicing. Thus, to test our hypothesis that CLAMP shapes transcriptome diversity by regulating RNA transcript splicing, we performed mRNA-sequencing in the presence and absence of CLAMP in Kc (female) and S2 (male) cell lines and used a SUPPA based pipeline called time2splice (https://github.com/ashleymaeconard/time2splice) to analyze the sequencing data. Using both cell lines helped us to identify female (Kc) and male (S2) specific splicing events. We identified 452 genes differentially spliced beetween Kc and S2 cells. There are 46 and 113 CLAMP-dependent splicing events in Kc and S2 cells, respectively. Interestingly, 45 CLAMP-dependent events (belonging to 42 genes) are specific to Kc cells, i.e female-specific, whereas 112 CLAMP dependent events (belonging to 100 genes) in S2 cells are specific to S2 cells, i.e male-specific. Therefore, we identified a new role for the transcription factor CLAMP in regulated sex-specific splicing events in Kc and S2 cells.
2023-08-02 | GSE220439 | GEO
Project description:CLAMP transcription factor is a GA binding protein that recruits the male dosage compensation complex MSL to male X-chromosome, thus upregulating transcription level twice that of female X-chromosomes. Interestingly, CLAMP is essential for survival of both male and female individuals. However, what function it plays at the autosomes, especially in females is not known. It is well establised that GA-rich sequences to which CLAMP bind have evolved from polypyrimidine tracts that regulate splicing. Also, MALDI-MS data shows that CLAMP binds to RNA binding proteins involved in sex-specific splicing like, Squid. Furthermore, as part of MSL complex CLAMP interacts with MLE, a RNA helicase which is a known component of spliceosome complex conserved from flies to humans. Thus, we hypothesized that CLAMP plays a role in sex-spcific splicing. Given that CLAMP is a transcription factor, understanding mechanism of how it regulates splicing will add to our understanding about how regulators of gene expression shape the transcriptome differentially between males and females. We tested our hypothesis by identifying CLAMP dependent female and male sex-specific splicing events by analysing RNA-seq data from control as well as clampnull mutant male and female third instar larvae (L3) using suppa based time2splice pipeline (https://github.com/ashleymaeconard/time2splice). We identified 189 and 211 CLAMP dependent splicing events in female and male L3. Out of these 139 were female sex-specific splicing whereas 161 was male-sex-specific splicing. 50 splicing events identified as CLAMP dependent were non-specific to any particular sex, and were affect in both the sexes. Interestingly, we identified splicing of master regulator of sex-determination sxl affected by absence of CLAMP in females whereas splicing of one of the sex-determination pathway effectors, tra regulated by CLAMP in males. These findings indicate CLAMP plays important role in determining sex-specific transcript diversity in Drosophila.
2023-08-02 | GSE220455 | GEO
Project description:CLAMP is a maternally deposited transcription factor binding to GA-rich genomic region. It recruits male-specific MSL (Male sex-lethal) complex on male X-chromosome that regulates dosage compensation in males, and competes with other GA-binding proteins like GAF, acting both synergistically as well as antagonistically in a context dependent manner. Considering CLAMP's male specific role in dosage compensation but ubiquitous expression in both males and females, what regulates differential binding of CLAMP is still unknown. Furthermore, ChiP-seq analysis has being long used to study protein-DNA interactions, however, since it uses physical force to shear DNA after protein capture, it is prone to both false negative and false positive peaks being called. To minimise these discrepancies, a recent CutnRun MNase based technique have being developed (Heinkoff Lab) which captures the DNA fragments bound to protein of interest. Due to higher accuracy in identifying the size and sequence of the protein bound DNA region, one can also identify monomeric or multimeric protein binding on the chromatin. We have used this technique to identify differential and shared CLAMP binding sites in male and female cells. We also compared CLAMP DNA binding peaks with CLAMP RNA binding peaks (iCLIP data) in male and female cells to understand sex-biased co-transcriptional RNA processing mechanism.
2023-12-31 | GSE220053 | GEO
Project description:Deciphering the role of alternative splicing in developmental processes relies on the identification of key genes whose expression is controlled by splicing regulators throughout growth of a whole organism. Targeting expression of five SR proteins in the developing eye of Drosophila allowed us to show that these splicing factors induce various phenotypic alterations concerning eye organogenesis and viability. Although both dASF/SF2 and B52 caused defects in ommatidia structure, only B52 impairs normal photoreceptor axons projection and neurogenesis in visual ganglia. Consistently, microarray analyses revealed that many of the B52 targets are involved in brain organogenesis and we show that their splicing profile is altered both in B52 loss and gain of function. Conversely, a large proportion of dASF/SF2 targets are involved in eye development. This differential effect argues that SR proteins confer accuracy to developmental gene-expression programs, thus ensuring tissue identity and supporting cell-lineage decisions. Keywords: genetic modification Experiment aimed at identifying B52 potential mRNA targets in the Drosophila developing eye Control: GMR X GFP-NLS trangenic larvae The 2 IP- GFP cy3 vs B52 cy5 rep.1 and IP- GFP cy5 vs B52 cy3 rep.1 samples correspond to dye-swap experiments. Two replicates (rep1 and rep2) are included.
2007-09-17 | E-GEOD-5858 | biostudies-arrayexpress
Project description:Identifying transcriptional targets of ONECUT3 by RNAseq
| PRJNA837005 | ENA
Project description:Identifying miR-139 targets in motor neurons
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Project description:ChIP-seq and mRNA-seq experiments were performed to understand the role of the CLAMP protein in dosage compensation ChIP-seq experiments compared the binding profiles of CLAMP in male and female cells and mRNA-seq data to define the role of CLAMP in regulating genes on the X-chromosome
2013-07-15 | E-GEOD-39271 | biostudies-arrayexpress
Project description:Deciphering the role of alternative splicing in developmental processes relies on the identification of key genes whose expression is controlled by splicing regulators throughout growth of a whole organism. Targeting expression of five SR proteins in the developing eye of Drosophila allowed us to show that these splicing factors induce various phenotypic alterations concerning eye organogenesis and viability. Although both dASF/SF2 and B52 caused defects in ommatidia structure, only B52 impairs normal photoreceptor axons projection and neurogenesis in visual ganglia. Consistently, microarray analyses revealed that many of the B52 targets are involved in brain organogenesis and we show that their splicing profile is altered both in B52 loss and gain of function. Conversely, a large proportion of dASF/SF2 targets are involved in eye development. This differential effect argues that SR proteins confer accuracy to developmental gene-expression programs, thus ensuring tissue identity and supporting cell-lineage decisions. Keywords: genetic modification Experiment aimed at identifying dASF/SF2 potential mRNA targets in the Drosophila developing eye Control: GMR X GFP-NLS trangenic larvae The 2 IP- GFP cy3 vs ASF cy5 rep.1 and IP- GFP cy5 vs ASF cy3 rep.1 samples correspond to dye-swap experiments. Two replicates (rep1 and rep2) are included.
2007-09-17 | E-GEOD-5857 | biostudies-arrayexpress
Project description:Identifying transcriptional targets of ONECUT3 by ChIP-seq
| PRJNA836928 | ENA
Project description:Investigated the DNA binding specificity of the C-clamp of dnTCF1E Used a doxycycline inducible system to investigate the role of the C-clamp in the genome-wide binding of dnTCF1E. Induced dnTCF1EWT and dnTCF1Emut (C-clamp mutant that is null for C-clamp DNA binding) over 2 time points (2 hours and 9 hours). A 0 hour (no doxycycline, uninduced) control was also included.
2014-12-04 | E-GEOD-53536 | biostudies-arrayexpress