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Transcriptome and small RNA landscapes of tree shrew testes and epididymides during reproductive and non-reproductive seasons

ABSTRACT: The treeshrew (Tupaia belangeri), as a species closely related to primates, exhibits significant seasonal reproductive traits in adult males, including cyclical changes in testicular morphology and spermatogenesis between the breeding and non-breeding seasons. However, comprehensive and systematic data resources on the molecular expression features associated with this physiological phenomenon are still lacking. In this study, we utilized the Illumina high-throughput sequencing platform to perform RNA-seq and small RNA-seq on testicular (Testis), caput epididymis (Caput), and caudal epididymis (Cauda) tissues collected from male tree shrews during both breeding (R) and non-breeding (NR) seasons. These analyses were performed to generate a paired transcriptomic and small RNA dataset covering multiple tissues and seasonal states. RNA-seq analysis identified 21,299 expressed genes, and weighted gene co-expression network analysis (WGCNA) was performed to identify modules and candidate genes significantly associated with reproductive status. The small RNA-seq data systematically characterized the composition and expression landscapes of miRNAs and piRNAs, leading to the identification of 263 novel miRNAs, 59,590 piRNAs, and 139 piRNA clusters. Differential expression analyses and computational predictions were further integrated to construct a miRNA–candidate gene–piRNA association network. The multi-omics data generated in this study provide a systematic and reusable molecular resource for deciphering the gene expression and small RNA characteristics associated with seasonal reproduction in male tree shrews. This dataset can serve as an important reference for investigating dynamic changes in mammalian reproductive tissues.

ORGANISM(S): Tupaia belangeri chinensis

PROVIDER: GSE335066 | GEO | 2026/06/16

REPOSITORIES: GEO

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Project description:The role of estrogen and testosterone in the regulation of gene expression in the proximal reproductive tract is not completely understood. To address this question, mice were treated with testosterone or estradiol and RNA from the efferent ducts and caput epididymis was processed and hybridized to Affymetrix MOE 430 2.0 microarrays. Analysis of array output identified probe sets in each tissue with altered levels in hormone treated versus control animals. Hormone treatment efficacy was confirmed by determination of serum hormone levels pre- and post-treatment and observed changes in transcript levels of previously reported hormone-responsive genes. Tissue-specific hormone sensitivity was observed with 2867 and 3197 probe sets changing significantly in the efferent ducts after estrogen and testosterone treatment, respectively. In the caput epididymis, 117 and 268 probe sets changed after estrogen and testosterone treatment, respectively, demonstrating a greater response to hormone in the efferent ducts than the caput epididymis. Transcripts sharing similar profiles in the intact and hormone-treated animals compared with castrated controls were also identified. Ontological analysis of probe sets revealed a significant number of hormone-regulated transcripts encode proteins associated with lipid metabolism, transcription and steroid metabolism in both tissues. Real-time RT-PCR was employed to confirm array data and investigate other potential hormone-responsive regulators of proximal reproductive tract function. The results of this work reveal previously unknown responses to estrogen in the caput epididymis and to testosterone in the efferent ducts as well as tissue specific hormone sensitivity in the proximal reproductive tract. Adult animals were castrated or sham-castrated, allowed to recover for 14 days, and then treated with 0.015 mg estradiol (castrated), 0.015 mg testosterone propionate (castrated), or vehicle (castrated and sham-castrated as biological controls) in duplicate. Efferent duct and caput epididymis was collected from each sample and analyzed. Duplicates are included in the provided data and numbered 1 or 2 for each treatment regimen.

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.

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Transcriptome and small RNA landscapes of tree shrew testes and epididymides during reproductive and non-reproductive seasons

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