A long non-coding RNA interacts with Gfra1 and maintains survival of mouse spermatogonial stem cells

Abstract

Spermatogonial stem cells (SSCs) are unique male germline stem cells that support spermatogenesis and male fertility. Long non-coding RNAs (lncRNA) have been identified as key regulators of stem cell fate; however, their role in SSCs has not been explored. Here, we report that a novel spermatogonia-specific lncRNA (lncRNA033862) is essential for the survival of murine SSCs. LncRNA033862 is expressed in early spermatogonia including SSC and was among 805 lncRNAs identified by global expression profiling as responsive to glial cell-derived neurotrophic factor (GDNF), a growth factor required for SSC self-renewal and survival. LncRNA033862 is an antisense transcript of the GDNF receptor alpha1 (Gfra1) that lacks protein coding potential and regulates Gfra1 expression levels by interacting with Gfra1 chromatin. Importantly, lncRNA033862 knockdown severely impairs SSC survival and their capacity to repopulate recipient testes in a transplantation assay. Collectively, our data provide the first evidence that long non-coding RNAs (lncRNAs) regulate SSC fate.

Systematic identification and comparison of expressed profiles of lncRNAs and circRNAs with associated co-expression and ceRNA networks in mouse germline stem cells

“…Interestingly, the strong enrichment PI3K-Akt pathway, which was observed in the highly co-expressed mRNAs and lncRNAs of SSCs and FGSCs (Supplementary Figure 4), suggests that FGSCs had a similar GDNF signaling mechanism as SSCs. To determine whether FGSCs had the same GDNF signaling mechanism as SSCs, we removed GDNF from the culture medium for a period of 7 days and found that the results were similar to those of a previous study on SSCs [64]. Withdrawal of GDNF for a week resulted in a significant reduction in cell numbers (of 1.0 × 10 5 plated cells, an average of 0.4×10 5 remained after GDNF depletion) (Figure 4C–4E).…”

Mrhl Long Noncoding RNA Mediates Meiotic Commitment of Mouse Spermatogonial Cells by Regulating Sox8 Expression

“…It is clear from such studies that the transcriptome levels of lncRNAs are greater than those of mRNAs during spermatogenesis and that the expression of lncRNAs is also dynamically regulated. Mechanistic details of lncRNA function during spermatogenesis have been provided by only a few studies, including our report of the role of mrhl RNA in regulation of Wnt signaling through p68 RNA helicase (27,(44)(45)(46). The present report describes a new dimension in the context of spermatogenesis, with lncRNA regulating the expression of an important transcription factor and such regulation being crucial for meiotic entry or spermatogonial differentiation.…”

Noncoding RNAs: Potential players in the self‐renewal of mammalian spermatogonial stem cells

“…In recent years, it has become clear that serval types of ncRNAs, involving miRNA, lncRNA, piRNA, small nucleolar RNA, and circRNAs, play important roles in stem cell self‐renewal by forming complicated epigenetic regulatory networks together with other epigenetic factors related to DNA methylation, histone methylation, and acetylation (Dupuis‐Sandoval, Poirier, & Scott, ; S. Hu & Shan, ; Rojas‐Rios, Chartier, Pierson, & Simonelig, ; C. Y. Yu et al, ; Z. Yu, Li, Fan, Liu, & Pestell, ). As for SSCs, although a large number of ncRNAs have been screened as potential regulatory molecules for SSC self‐renewal using high‐throughput RNA sequencing and bioinformatics approaches, only a small portion of them have been validated and functionally characterized using biological experiment to uncover roles in SSCs self‐renewal, particularly lncRNAs, piRNA, and circRNAs (K. Hu et al, ; Li et al, ; X. Li, Ao, et al, ), thus there is an urgent need to reveal more ncRNAs associated with SSC self‐renewal to elucidate the molecular mechanisms underlying SSC self‐renewal. Encouragingly, great improvements of technologies have been recently made on isolation, characterization, in vitro culture, and in vivo transplantation of SSCs will provide us a robust research platform to explore the function of ncRNAs in SSC self‐renewal (X. Li, Ao, et al, ; Mulder et al, ; Takashima & Shinohara, ; Zhang, Sun, & Zou, ).…”

“…Accumulating evidence has shown that lncRNAs are important players in diverse cellular processes including proliferation, differentiation, apoptosis, senescence, stress response through complicated mechanisms involving lncRNA–mRNA, lncRNA–miRNA, lncRNA–DNA, and lncRNA–protein interaction (Fatica & Bozzoni, ), although in vivo functional characterization of lncRNA are still facing many challenges, for instance, some of lncRNA mutant mice do not exhibit a severe or expected phenotype similar to that of in vitro experiments (Bassett et al, ; Kohtz, ; S. Nakagawa, ). To date, some lncRNAs have been demonstrated to play critical regulatory roles in posttranscriptional and transcriptional regulation in self‐renewal of stem cell, including SSCs (Lee et al, ; Li et al, ).…”

“…Liang et al () found 241 lncRNAs exhibited SSC‐specific expression pattern by comparing the lncRNA expression profiles of mouse SSC, type A spermatogonia, pachytene spermatocytes, and round spermatids using microarray and subsequently characterized a lncRNA AK015322, 1 of 241 lncRNAs, was highly expressed in mouse SSCs and could promote the proliferation of mouse SSC line C18‐4 in vitro via regulating ETV5 (a pivotal transcriptional factor for SSC self‐renewal) as a decoy of miR‐19b‐3p. To identify lncRNAs with potential regulatory roles in SSCs, L. Li et al () identified 805 lncRNAs transcripts in mouse SSCs were subjected to expression level changes in response to GDNF (an essential growth factor required for SSC self‐renewal) through applying high‐throughput RNA sequencing. They further found lncRNA‐033862, an lncRNA subjected to GDNF signaling, was specifically expressed in mouse SSCs and could regulate the expression level of Gfra1 which encodes the GDNF coreceptor by interacting with Gfra1 chromatin, as well as its knockdown could impair self‐renewal and survival of mouse SSCs both in vitro and in vivo, demonstrating that lncRNA‐033862 is involved in the maintenance of SSC (K. Hu, Zhang, & Liang, ).…”