Project description:GATA3 is a basic and essential transcription factor that regulates many pathophysiological processes that is required for the development of mammary luminal epithelial cells. Loss-of-function GATA3 alterations in breast cancer are associated with poor prognosis. However, its regulation and function in miRNAs expression has been less explored.
Project description:We performed ChIPseq for GATA3 on MCF-7 cells treated with vehicle (DMSO), GDC-9545 or GNE-858. GNE-858 is a non-ER degrader (SERM) from the same chemical series as GDC-9545.
Project description:CAKUT (congenital anomalies of the kidney and the urinary tract) is one of the most prevalent birth defects, affecting around 1% of newborns. However, the genetic factors underlying the majority of CAKUT are unknown. Drosha, the microRNA-processing ribonuclease, has been reported to regulate the nephric cap meschyme development, and its mutations are implicated in Wilms tumor, a pediatric kidney cancer. Here, we investigated the role of Drosha in mesangial cells and its impact on glomerular capillary tuft formation. Deletion of Drosha in mesangial cells (DroshacKO) disrupted the glomerular capillary tufts formation, leading to dysplastic glomeruli, proteinuria, oliguria, reduced looping of glomerular capillaries and capillary dilation. Drosha-knockdown (KD) in mesangial cells (SV40 MES 13) leads to decreased cell proliferation and reduced Gata3 protein level. Transcriptome analysis by RNA-seq shows decreased level of ribosome protein gene (RPG)s but unaltered mRNA level of major genes affecting kidney development, including Gata3, Pax2, Atn1, Wt1 etc. in Drosha-KD compared to Control SV40 MES13 cells. Further analysis indicates Drosha regulates the translation of Gata3 in mesangial cells via regulating RPG transcription. Our work, for the first time, reveals that Drosha in mesangial cells orchestrates the formation of glomerular capillary tufts by regulating Gata3 translation. It identifies the critical role of DROSHA in nephric development and proposes DROSHA as a novel potential causal gene for CAKUT. [miRNA-Seq] This study investigates the role of DROSHA in microRNA biogenesis using the mouse renal mesangial cell line ME13 sv40. Small RNA sequencing was performed to compare microRNA expression profiles between DROSHA knockdown cells and control cells. The results aim to elucidate how DROSHA depletion alters microRNA expression patterns, which may provide insights into post-transcriptional regulation in renal mesangial cells.
Project description:The GATA3 transcription factor is one of the most frequently mutated genes in breast cancer. Heterozygous mutations, largely frameshifting, are seen in 15% of estrogen receptor positive breast cancers, the subtype in which these mutations are almost exclusively found. Mouse studies have shown that Gata3 is critical for breast development and that GATA3 gene dosage affects breast tumor progression. Human patient data have shown that high Gata3 expression, a feature of luminal subtype breast cancers, is associated with a better prognosis. Although the frequency of GATA3 mutation suggests an important role in breast cancer development or progression, there is little understanding of how mutations in GATA3 affect its function in luminal breast epithelial cells and what gene expression changes result as a consequence of the mutations. Here, using gene editing, we have created two sets of isogenic human luminal breast cancer cell lines with and without a truncating GATA3 mutation. GATA3 mutation enhanced tumor growth in vivo but did not affect sensitivity to clinically used hormonal therapies or chemotherapeutic agents. We identified genes upregulated and downregulated in GATA3 mutant cells, a subset of which was concordantly differentially expressed in GATA3 mutant primary luminal breast cancers. Addback of mutant GATA3 recapitulated mutation-specific gene expression changes and enhanced soft agar colony formation, suggesting a gain of function for the mutant protein.
Project description:Transcription factor GATA3 is essential for the specification and maintenance of the luminal cell differentiation in the mammary gland, and its expression is progressively lost during luminal breast cancer progression. However, how loss-of-function of GATA3 contributes to the development of breast cancer is still poorly understood. Here, we report that GATA3 nucleates a transcription repression program composed of G9A and MTA3-, but not MTA1- or MTA2-, constituted NuRD complex. Genome-wide analysis of the GATA3/G9A/NuRD(MTA3) targets identified a cohort of genes that are critically involved in epithelial-to-mesenchymal transition and cell invasion.
Project description:Purpose: We aimed to investigate in depth the regulation of microRNA expression by hypoxia in the breast cancer cell line MCF-7, establish the relationship between microRNA expression and HIF binding sites, pri-miRNA transcription and microRNA processing gene expression. Methods: microRNA sequencing data and gene expression microarray data were generated from MCF-7 cells submitted to an hypoxia timecourse (16h, 32h and 48h at 1% Oxygen). Data was integrated to 500 published high-stringency HIF binding sites identified in MCF-7 cells. Results: We identified 41 microRNAs significantly up- and 28 down- regulated, of which 38 mature and 20 star forms are reported in conjunction with hypoxia for the first time. HIF-1α and HIF-2α binding sites within 50kb distance of microRNA loci were found by integration of HIF ChIP-seq data, showing overall association between binding sites and up-regulation. Gene expression profiling analysis showed no full coordination between pri-miRNA and microRNA expression, pointing towards additional levels of regulation. Several transcripts playing a role in microRNA processing were found regulated by hypoxia, of which two were HIF dependent. Conclusions: The data support the hypothesis that microRNA expression under hypoxia is regulated at transcriptional and post-transcriptional level. HIF is involved at both levels, regulating the transcription of certain microRNAs and also the expression of key elements of the microRNA processing pathway.
Project description:Purpose: We aimed to investigate in depth the regulation of microRNA expression by hypoxia in the breast cancer cell line MCF-7, establish the relationship between microRNA expression and HIF binding sites, pri-miRNA transcription and microRNA processing gene expression. Methods: microRNA sequencing data and gene expression microarray data were generated from MCF-7 cells submitted to an hypoxia timecourse (16h, 32h and 48h at 1% Oxygen). Data was integrated to 500 published high-stringency HIF binding sites identified in MCF-7 cells. Results: We identified 41 microRNAs significantly up- and 28 down- regulated, of which 38 mature and 20 star forms are reported in conjunction with hypoxia for the first time. HIF-1α and HIF-2α binding sites within 50kb distance of microRNA loci were found by integration of HIF ChIP-seq data, showing overall association between binding sites and up-regulation. Gene expression profiling analysis showed no full coordination between pri-miRNA and microRNA expression, pointing towards additional levels of regulation. Several transcripts playing a role in microRNA processing were found regulated by hypoxia, of which two were HIF dependent. Conclusions: The data support the hypothesis that microRNA expression under hypoxia is regulated at transcriptional and post-transcriptional level. HIF is involved at both levels, regulating the transcription of certain microRNAs and also the expression of key elements of the microRNA processing pathway.
Project description:Purpose: We aimed to investigate in depth the regulation of microRNA expression by hypoxia in the breast cancer cell line MCF-7, establish the relationship between microRNA expression and HIF binding sites, pri-miRNA transcription and microRNA processing gene expression. Methods: microRNA sequencing data and gene expression microarray data were generated from MCF-7 cells submitted to an hypoxia timecourse (16h, 32h and 48h at 1% Oxygen). Data was integrated to 500 published high-stringency HIF binding sites identified in MCF-7 cells. Results: We identified 41 microRNAs significantly up- and 28 down- regulated, of which 38 mature and 20 star forms are reported in conjunction with hypoxia for the first time. HIF-1α and HIF-2α binding sites within 50kb distance of microRNA loci were found by integration of HIF ChIP-seq data, showing overall association between binding sites and up-regulation. Gene expression profiling analysis showed no full coordination between pri-miRNA and microRNA expression, pointing towards additional levels of regulation. Several transcripts playing a role in microRNA processing were found regulated by hypoxia, of which two were HIF dependent. Conclusions: The data support the hypothesis that microRNA expression under hypoxia is regulated at transcriptional and post-transcriptional level. HIF is involved at both levels, regulating the transcription of certain microRNAs and also the expression of key elements of the microRNA processing pathway.
Project description:SIRT1 is a nuclear NAD+-dependent protein deacetylase. Expression microarray analysis was used to study the effect of SIRT1 knockdown on gene expression in MCF-7 breast cancer cells.
Project description:NAMPT is an enzyme in the mammalian NAD+ salvage pathway. Expression microarray analysis was used to study the effect of NAMPT knockdown on gene expression in MCF-7 breast cancer cells.