HNF1 regulates critical functions of the human epididymis epithelium. [ChIP-Seq]
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE76376 | GEO | 2017/10/06
SECONDARY ACCESSION(S): PRJNA307118
REPOSITORIES: GEO
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Project description:HNF1α and HNF1β recognize the same DNA consensus sequence in the genome, to which they bind as homodimers or heterodimers. Both factors share a high degree of homology their DNA binding and dimerization (N-terminus) regions but have a more divergent C-terminal transactivation domain. HNF1β is essential for the generation of a functional male reproductive tract in mice and genital tract abnormalities are evident in humans with recessive mutations in HNF1β. The functions of HNF1α and HNF1β have been studied in epithelia from other several tissues (liver, kidney, intestine, and pancreas) but their role in the adult human epididymis epithelium (HEE) remains unexplored. We established that HNF1α/β are expressed in caput HEE cells and are predicted to occupy cis-regulatory elements in these cells. To investigate the contribution of HNF1 in controlling gene expression in caput cells we performed siRNA-mediated depletion of HNF1α and HNF1β together, followed by RNA-seq analysis. Three replicas of caput cells were transfected with the specific siRNAs or with a non-targeting control siRNA. RNA-seq after HNF1 depletion showed significant alterations in the expression of genes encoding ion channels and exchangers that are involved in controlling the luminal environment in the caput epididymis.
2017-10-06 | GSE76377 | GEO
Project description:Purpose: To compare the transcriptome profiles (RNA-seq) of cultured human epididymis cells and tissue from the caput, corpus and cauda regions of the human epididymis. Methods: Human epididymis tissue was obtained with Institutional Review Board approval from 3 patients (UC05, UC06, UC09, range: 22 - 36 years) undergoing inguinal radical orchiectomy for a clinical diagnosis of testicular cancer. None of the epididymides had extension of the testicular cancer. The three anatomical regions: caput, corpus and cauda, were separated and segments of each snap frozen. Adult human epididymis epithelial (HEE) cultures were also established from tissue. RNA was extracted from both tissue and cultured HEE cells and RNA-seq libraries prepared (TruSeq RNA Sample Preparation Kit v2, Low-Throughput protocol, Illumina). Libraries were sequenced on Illumina HiSeq2500 machines. Data were analyzed using TopHat and Cufflinks. Results: Libraries generated ~19-39 million reads per library from the cells (95-99% mapping to the human genome) and ~14-39 million reads from the tissue samples (84-99% mapped). Raw reads were aligned to the genome with Tophat and gene expression values were processed using Cufflinks as Fragments Per Kilobase per Million mapped fragments (FPKM). FPKM values were subject to principle component analysis, which revealed that though caput, corpus and cauda cell samples respectively from UC05, UC06 and UC09 clustered together. RNA-seq data from the 3 biological replicas (UC05, UC06 and UC09) of caput, corpus and cauda were pooled for further analysis. Cufflinks was used to determine differentially expressed genes (DEGs) between caput, corpus and cauda cells, combined from the 3 donors. The gene expression profiles of corpus and cauda are remarkably similar and both differ from the caput to a similar degree. We identified ~40 genes differentially expressed between corpus and cauda and more than 1600 DEGs between caput and cauda. The DEGs for each comparison (caput and corpus/cauda) were analysed using a gene ontology process enrichment analysis (DAVID, Huang et al., NAR 2009;37:1-13, Huang et al., 2009 Nat Prot 4:44-57). Conclusions: Here we describe an in depth analysis of the gene expression repertoire of primary cultures of epithelial cells and intact tissues from each region of the adult human epididymis. These data will be valuable to decipher pathways of normal epididymis function and aspects of epididymis disease that cause male infertility. RNA-seq was performed on libraries generated from caput, corpus and cauda-derived cultured cells (passage 2 or 3) from 3 donors and on caput, corpus and cauda tissue from 2 of the same donors. Donor age range: 22 - 36 years.
2016-01-04 | E-GEOD-72986 | biostudies-arrayexpress
Project description:Viral infections of the epididymis are associated with epididymitis, which damages the epithelium and impairs fertility. We showed previously that innate immune response genes were differentially expressed in the corpus and cauda region of the human epididymis in comparison to the caput. Here we investigate the antiviral defense response mechanisms of human epididymis epithelial (HEE) cells. Toll-like receptor (TLR) 3 and retinoic acid-inducible gene I (RIG-I)- like receptors (RLRs) are enriched in HEE cells from the corpus and cauda region. Furthermore, corpus and cauda HEE cells show an enhanced response to antiviral ligands (poly(I:C) and HSV-60), as shown by increased IFN-β mRNA expression and secretion of IFN-β. We also show that paired box 2 (PAX2), which was implicated in regulating antiviral response pathways is required for basal expression of the DNA sensor, Z-DNA binding protein (ZBP1) and type I interferon, in caput but not in cauda cells.
2019-10-11 | GSE104944 | GEO
Project description:Primary human epididymis epithelial (HEE) cells are valuable reagents for functional studies on the human epididymis. We used them previously to determine the transcriptional networks that establish cell identity along the length of the epididymis from caput, corpus and cauda. These studies on HEE cells and organoids derived from them revealed important cellular properties. However, similar to other primary cells, HEE cells undergo replicative senescence and de-differentiation in culture. A cocktail of small molecules was shown elsewhere to extend longevity of epithelial basal cells. The components included transforming growth factor β (TGF-β)/bone morphogenetic protein (BMP) antagonists, WNT agonist and Rho-associated and coiled-coil containing protein kinase (ROCK) inhibitor (ROCKi), which together prevented the senescence-related upregulation of TGF-β signaling pathway members. Here we treat HEE cells with the same cocktail and observed enhanced replicative potential and prolonged expression of markers of HEE differentiation. This treatment expands the differentiated HEE cell population available from individual epididymis tissue samples that can be used for molecular, cellular and functional studies.
2024-05-20 | GSE203535 | GEO
Project description:Purpose: To compare the transcriptome profiles (RNA-seq) of cultured human epididymis cells and tissue from the caput, corpus and cauda regions of the human epididymis. Methods: Human epididymis tissue was obtained with Institutional Review Board approval from 3 patients (UC05, UC06, UC09, range: 22 - 36 years) undergoing inguinal radical orchiectomy for a clinical diagnosis of testicular cancer. None of the epididymides had extension of the testicular cancer. The three anatomical regions: caput, corpus and cauda, were separated and segments of each snap frozen. Adult human epididymis epithelial (HEE) cultures were also established from tissue. RNA was extracted from both tissue and cultured HEE cells and RNA-seq libraries prepared (TruSeq RNA Sample Preparation Kit v2, Low-Throughput protocol, Illumina). Libraries were sequenced on Illumina HiSeq2500 machines. Data were analyzed using TopHat and Cufflinks. Results: Libraries generated ~19-39 million reads per library from the cells (95-99% mapping to the human genome) and ~14-39 million reads from the tissue samples (84-99% mapped). Raw reads were aligned to the genome with Tophat and gene expression values were processed using Cufflinks as Fragments Per Kilobase per Million mapped fragments (FPKM). FPKM values were subject to principle component analysis, which revealed that though caput, corpus and cauda cell samples respectively from UC05, UC06 and UC09 clustered together. RNA-seq data from the 3 biological replicas (UC05, UC06 and UC09) of caput, corpus and cauda were pooled for further analysis. Cufflinks was used to determine differentially expressed genes (DEGs) between caput, corpus and cauda cells, combined from the 3 donors. The gene expression profiles of corpus and cauda are remarkably similar and both differ from the caput to a similar degree. We identified ~40 genes differentially expressed between corpus and cauda and more than 1600 DEGs between caput and cauda. The DEGs for each comparison (caput and corpus/cauda) were analysed using a gene ontology process enrichment analysis (DAVID, Huang et al., NAR 2009;37:1-13, Huang et al., 2009 Nat Prot 4:44-57). Conclusions: Here we describe an in depth analysis of the gene expression repertoire of primary cultures of epithelial cells and intact tissues from each region of the adult human epididymis. These data will be valuable to decipher pathways of normal epididymis function and aspects of epididymis disease that cause male infertility.
2016-01-04 | GSE72986 | GEO
Project description:The epithelium lining the epididymis in the male reproductive tract maintains a luminal environment that promotes sperm cell maturation. This process is dependent on the coordinated expression of many genes that encode proteins with a role in epithelial transport. We previously generated genome-wide maps of open chromatin in primary human fetal epididymis epithelial cells to identify potential regulatory elements controlling coordinated gene expression in the epididymis epithelium. Subsequent in silico analysis identified transcription factor binding sites (TFBS) that were over-represented in the HEE open chromatin, include the motif for paired box 2 (PAX2). PAX2 is a critical transcriptional regulator of urogenital tract development, which is well studied in the kidney but is unexplored in the epididymis. Due to the limited lifespan of primary HEE cells in culture we investigated the role of PAX2 in an immortalized HEE cell line (REP). First, REP cells were evaluated by DNase-seq and their open chromatin map overlapped that of primary HEE cells at ~ 65% of sites. Moreover, the PAX2-binding motif was again identified as an overrepresented TFBS within intergenic open chromatin, though on fewer chromosomes than in the primary HEE cells. To identify PAX2-target genes in REP cells, RNA-seq analysis was performed after siRNA-mediated depletion of PAX2 in comparison to a non-targeting siRNA. In response to PAX2-represssion, 3142 transcripts were differentially expressed (1334 up-regulated and 1808 down-regulated). Novel PAX2 targets included multiple genes encoding proteins with a predicted function in the epididymis epithelium. examination of open chromatin region in REP cells with 2 replicates
2015-01-07 | E-GEOD-59782 | biostudies-arrayexpress
Project description:The epithelium lining the epididymis in the male reproductive tract maintains a luminal environment that promotes sperm cell maturation. This process is dependent on the coordinated expression of many genes that encode proteins with a role in epithelial transport. We previously generated genome-wide maps of open chromatin in primary human fetal epididymis epithelial cells to identify potential regulatory elements controlling coordinated gene expression in the epididymis epithelium. Subsequent in silico analysis identified transcription factor binding sites (TFBS) that were over-represented in the HEE open chromatin, include the motif for paired box 2 (PAX2). PAX2 is a critical transcriptional regulator of urogenital tract development, which is well studied in the kidney but is unexplored in the epididymis. Due to the limited lifespan of primary HEE cells in culture we investigated the role of PAX2 in an immortalized HEE cell line (REP). First, REP cells were evaluated by DNase-seq and their open chromatin map overlapped that of primary HEE cells at ~ 65% of sites. Moreover, the PAX2-binding motif was again identified as an overrepresented TFBS within intergenic open chromatin, though on fewer chromosomes than in the primary HEE cells. To identify PAX2-target genes in REP cells, RNA-seq analysis was performed after siRNA-mediated depletion of PAX2 in comparison to a non-targeting siRNA. In response to PAX2-represssion, 3142 transcripts were differentially expressed (1334 up-regulated and 1808 down-regulated). Novel PAX2 targets included multiple genes encoding proteins with a predicted function in the epididymis epithelium. mRNA profile of control and PAX2 knockdown REP cells
2015-01-07 | E-GEOD-59781 | biostudies-arrayexpress
Project description:The epithelium lining the epididymis in the male reproductive tract maintains a luminal environment that promotes sperm cell maturation. This process is dependent on the coordinated expression of many genes that encode proteins with a role in epithelial transport. We previously generated genome-wide maps of open chromatin in primary human fetal epididymis epithelial cells to identify potential regulatory elements controlling coordinated gene expression in the epididymis epithelium. Subsequent in silico analysis identified transcription factor binding sites (TFBS) that were over-represented in the HEE open chromatin, include the motif for paired box 2 (PAX2). PAX2 is a critical transcriptional regulator of urogenital tract development, which is well studied in the kidney but is unexplored in the epididymis. Due to the limited lifespan of primary HEE cells in culture we investigated the role of PAX2 in an immortalized HEE cell line (REP). First, REP cells were evaluated by DNase-seq and their open chromatin map overlapped that of primary HEE cells at ~ 65% of sites. Moreover, the PAX2-binding motif was again identified as an overrepresented TFBS within intergenic open chromatin, though on fewer chromosomes than in the primary HEE cells. To identify PAX2-target genes in REP cells, RNA-seq analysis was performed after siRNA-mediated depletion of PAX2 in comparison to a non-targeting siRNA. In response to PAX2-represssion, 3142 transcripts were differentially expressed (1334 up-regulated and 1808 down-regulated). Novel PAX2 targets included multiple genes encoding proteins with a predicted function in the epididymis epithelium. Gene expression in REP cells was measured at 16 hours after exposure to 1nM R1881 or vehicle control.
2015-01-07 | E-GEOD-60381 | biostudies-arrayexpress
Project description:Objectives Here we used single cell RNA-seq technologies to elucidate the identity of cells within the human efferent ducts and compared them to caput epididymis cells. We also compared the cellularity of primary tissues with those of 2D and 3D (organoid) culture models used for functional studies. Materials and Methods Human epididymis tissue was dissected to separate different anatomical regions and digested to release single cells for processing on the 10X Genomics Chromium platform. Primary human epididymis epithelial (HEE) cells and HEE organoids were grown as described previously and subjected to single cell (sc) RNA-seq. ScRNA-seq data were processed by standard bioinformatics pipelines and used for comparative analysis. Results We define the cell types in the efferent ducts which include specialized epithelial cells, connective tissue stromal cells, vascular endothelial cells, smooth muscle cells and immune cells, but lack basal cells that are seen in the caput epididymis. Furthermore, we identify a sub-population of epithelial cells which have marker genes found in the bladder and urothelium. Comparative genomics analysis of the 2D and 3D culture models shows cellular identities adapted to the culture environment while still maintaining similarity to the primary tissue. Discussion Our data suggest that efferent ducts are lined with a transitional epithelium, which like the urothelium is able to stretch and contract depending on luminal volume. This is consistent with its primary role in seminal fluid resorption and sperm concentration. Moreover, we describe the cellularity of models to study the human epididymis epithelium in vitro. Conclusion Single cell RNA-seq data from the human epididymis make a valuable contribution to our understanding of this highly specialized organ.
2023-06-21 | GSE235009 | GEO
Project description:The androgen receptor (AR) has a pivotal role in regulating gene expression in the male reproductive system. Due to the involvement of AR in prostate cancer, its role is best studied in the prostate gland epithelium and prostate cancer cell lines. Here we investigate the transcriptional program of AR in normal human epididymis epithelial (HEE) cells. After AR stimulation of caput HEE cells with the synthetic androgen R1881, AR targets were revealed with RNA-sequencing. Next, AR occupancy genome-wide was determined in control or R1881-stimulated HEE cells by chromatin immunoprecipitation and deep sequencing (ChIP-seq). The results identify about 200 genes that are differentially expressed (DEGs) in HEE cells after AR activation. Some of these DEGs show occupancy of AR at their promoters or cis-regulatory elements suggesting direct regulation. However there is little overlap in AR-associated DEGs between HEE and prostate epithelial cells. Inspection of over-represented motifs in AR ChIP-seq peaks identified CAAT-enhancer binding protein beta (CEBPB) and Runt-related transcription factor 1 (RUNX1) as potential co-factors, with no evidence for FOXA1, which is an important co-factor in the prostate epithelium. CEBPB and RUNX1 ChIP-seq in HEE cells showed that both these factors often occupied AR-binding sites, though rarely simultaneously. Further analysis at a single AR-regulated locus (FK506-binding protein 5, FKPB5) suggests that RUNX1 may inhibit AR occupancy, while CEBP appears to be a co-activator. These data suggest a novel AR transcriptional network governs differentiated functions of the human epididymis epithelium.
2018-08-21 | GSE109061 | GEO