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.
Background
Dysregulation of microRNAs (miRNAs) play critical roles in cancerous processes. Although miR-3064 was reported to be an important tumor suppressor in ovarian cancer, the cellular impact of miR-3064 on pancreatic cancer (PC) progression, its downstream target genes and upstream mechanisms that control the expression of miR-3064 remain to be fully clarified.
Methods
We compared miRNA expression profiles between PC tissues compared with normal tissues using a miRNA microarray analysis of clinical samples, and screened the identified miRNAs for their influence on cell proliferation. We measured the expression of miR-3064 in PC tissues and PC cell lines using quantitative real-time PCR assays. Gain- and loss-of-function experiments were conducted to explore the biologic significance of miR-3064 in PC progression both in vitro and in vivo. The interactions between miR-3064 and long noncoding RNA (lncRNA) PXN-AS1 was verified using the luciferase reporter assay and RNA immunoprecipitation assay.
Results
We showed that miR-3064 was significantly overexpressed in PC tissues compared to normal tissues. High miR-3064 was associated with worse prognosis in patients with PC. Functionally, ectopic expression of miR-3064 promoted the proliferation, invasion, clone formation and sphere formation of PC cells in vitro and stimulated PC growth in vivo, while specific knockdown of miR-3064 or CRISPR/Cas9-mediated knockout of miR-3064 resulted in opposite phenotypes. Further investigation revealed that miR-3064 directly targeted PIP4K2B, which was reduced in PC tissues and attenuated PC cell proliferation, invasion and sphere formation induced by miR-3064. Importantly, lncRNA PXN-AS1 expression was downregulated in PC samples, and it directly interacted with miR-3064 and suppressed its levels in PC cells. Enforced expression of PXN-AS1 remarkably decreased cell proliferation, invasion and sphere formation, while re-expression of miR-3064 abrogated these effects of PXN-AS1.
Conclusions
MiR-3064, a key oncogenic miRNA, could promote PC cell growth, invasion and sphere formation via downregulating the levels of tumor suppressor PIP4K2B. PXN-AS1 functioned as a sponge to suppress the expression of miR-3064. These observations offer fresh insight into the mechanisms through which miR-3064 modulates the development of PC.
Electronic supplementary material
The online version of this article (10.1186/s13046-019-1379-5) contains supplementary material, which is available to authorized users.
In the original publication of this manuscript [1], there are errors in Fig. 3. The authors declare that the identified errors do not change the results or conclusions of this paper.
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