Abstract
BackgroundThe formation of matured and individual sperm involves a series of molecular and spectacular morphological changes of the developing cysts in Drosophila melanogaster testis. Recent advances in RNA Sequencing (RNA-Seq) technology help us to understand the complexity of eukaryotic transcriptomes by dissecting different tissues and developmental stages of organisms. To gain a better understanding of cellular differentiation of spermatogenesis, we applied RNA-Seq to analyse the testis-specific transcriptome, including coding and non-coding genes.ResultsWe isolated three different parts of the wild-type testis by dissecting and cutting the different regions: 1.) the apical region, which contains stem cells and developing spermatocytes 2.) the middle region, with enrichment of meiotic cysts 3.) the basal region, which contains elongated post-meiotic cysts with spermatids. Total RNA was isolated from each region and analysed by next-generation sequencing. We collected data from the annotated 17412 Drosophila genes and identified 5381 genes with significant transcript accumulation differences between the regions, representing the main stages of spermatogenesis. We demonstrated for the first time the presence and region specific distribution of 2061 lncRNAs in testis, with 203 significant differences. Using the available modENCODE RNA-Seq data, we determined the tissue specificity indices of Drosophila genes. Combining the indices with our results, we identified genes with region-specific enrichment in testis.ConclusionBy multiple analyses of our results and integrating existing knowledge about Drosophila melanogaster spermatogenesis to our dataset, we were able to describe transcript composition of different regions of Drosophila testis, including several stage-specific transcripts. We present searchable visualizations that can facilitate the identification of new components that play role in the organisation and composition of different stages of spermatogenesis, including the less known, but complex regulation of post-meiotic stages.
Highlights
The formation of matured and individual sperm involves a series of molecular and spectacular morphological changes of the developing cysts in Drosophila melanogaster testis
Transcriptome analysis of testis using RNA Sequencing (RNA-Seq) To gain a better understanding of cellular differentiation during spermatogenesis, we decided to compare transcript composition of different parts of the Drosophila testis
The apical region contains the spermatogonial stages represented by mitotically dividing cells; the middle region of the testis is enriched with meiotic spermatocytes and the basal region is filled with transcriptionally inactive elongated spermatids
Summary
The formation of matured and individual sperm involves a series of molecular and spectacular morphological changes of the developing cysts in Drosophila melanogaster testis. To gain a better understanding of cellular differentiation of spermatogenesis, we applied RNA-Seq to analyse the testis-specific transcriptome, including coding and non-coding genes. A Drosophila testis is a blind-ended tube, where consecutive stages of spermatogenesis are presented in the developmental order from the apical end to the basal end. Two somatic cyst cells support the germline cells throughout the entire spermatogenesis process. The Nebenkern unfurls and establishes two mitochondrial derivatives which elongate in the entire 1.8 mm length of the spermatid tail near the axoneme. The molecular composition and proteins responsible for the regulation of changes in mitochondria during spermatogenesis are not fully understood
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