Unraveling transcriptome dynamics in human spermatogenesis

Abstract

Spermatogenesis is a dynamic developmental process that includes stem cell proliferation and differentiation, meiotic cell divisions and extreme chromatin condensation. Although studied in mice, the molecular control of human spermatogenesis is largely unknown. Here, we developed a protocol that enables next-generation sequencing of RNA obtained from pools of 500 individually laser-capture microdissected cells of specific germ cell subtypes from fixed human testis samples. Transcriptomic analyses of these successive germ cell subtypes reveals dynamic transcription of over 4000 genes during human spermatogenesis. At the same time, many of the genes encoding for well-established meiotic and post-meiotic proteins are already present in the pre-meiotic phase. Furthermore, we found significant cell type-specific expression of post-transcriptional regulators, including expression of 110 RNA-binding proteins and 137 long non-coding RNAs, most of them previously not linked to spermatogenesis. Together, these data suggest that the transcriptome of precursor cells already contains the genes necessary for cellular differentiation and that timely translation controlled by post-transcriptional regulators is crucial for normal development. These established transcriptomes provide a reference catalog for further detailed studies on human spermatogenesis and spermatogenic failure.

Distinct prophase arrest mechanisms in human male meiosis

“…3B, ). We therefore looked more closely at the expression of genes that we previously found to be upregulated during the leptotene/zygotene to pachytene transition (Jan et al, 2017). This analysis first revealed that many genes from this gene set were not upregulated in type I or type II arrested spermatocytes but remained at leptotene/zygotene-like expression levels (Fig.…”

“…Importantly, we used the transcriptome of early pachytene spermatocytes, collected in a previous study describing gene expression throughout normal spermatogenesis (Jan et al, 2017), as controls. In this study, almost all genes characteristic for meiosis appeared to be already expressed in early spermatocytes, with early and late spermatocytes only displaying 24 DEGs.…”

“…For each patient, 500 histologically pachytene-like spermatocytes were individually laser dissected and pooled as described previously (Jan et al, 2017). Laser-dissected cells were captured in silicon-coated adhesive caps (Adhesive cap 500 opaque tube, Zeiss) and were lysed at 42°C in 10 µl of extraction buffer provided in the PicoPure RNA isolation kit (Arcturus).…”

Spermatogenesis and the Evolution of Mammalian Sex Chromosomes

“…Newly arisen male-biased genes are often X-linked, but over time the movement of these genes from the X to the autosomes has led to the de-enrichment of X-linked male meiotic genes [46]. Meiotic silencing may also select for X-linked genes that are critical in the later stages of meiosis to be highly expressed earlier in development (prior to meiosis or in early meiosis), so that transcripts persist through MSCI [48,49]. If true, then the constraints of MSCI may partially explain the enrichment for X-linked genes that are expressed earlier in spermatogenesis.…”

“…DNA becomes highly condensed and the morphological features of the mature spermatozoa take shape through the process of spermiogenesis (Figure 1A). Spermiogenesis is a highly specialized developmental process and many genes expressed during this phase are specific to postmeiotic cells [48,49,71,72]. Similar to MSCI, sex-linked genes are also partially silenced during postmeiotic development through a process known as postmeiotic sex chromosome repression (PSCR) .…”