MicroRNAs-10a and -10b Contribute to Retinoic Acid-induced Differentiation of Neuroblastoma Cells and Target the Alternative Splicing Regulatory Factor SFRS1 (SF2/ASF)

Abstract

MicroRNAs (miRNAs) are an emerging class of non-coding endogenous RNAs involved in multiple cellular processes, including cell differentiation. Treatment with retinoic acid (RA) results in neural differentiation of neuroblastoma cells. We wanted to elucidate whether miRNAs contribute to the gene expression changes induced by RA in neuroblastoma cells and whether miRNA regulation is involved in the transduction of the RA signal. We show here that RA treatment of SH-SY5Y neuroblastoma cells results in profound changes in the expression pattern of miRNAs. Up to 42 different miRNA species significantly changed their expression (26 up-regulated and 16 down-regulated). Among them, the closely related miR-10a and -10b showed the most prominent expression changes. Induction of miR-10a and -10b by RA also could be detected in LA-N-1 neuroblastoma cells. Loss of function experiments demonstrated that miR-10a and -10b are essential mediators of RA-induced neuroblastoma differentiation and of the associated changes in migration, invasion, and in vivo metastasis. In addition, we found that the SR-family splicing factor SFRS1 (SF2/ASF) is a target for miR-10a -and -10b in HeLa and SH-SY5Y neuroblastoma cells. We show here that changes in miR10a and -10b expression levels may regulate SFRS1-dependent alternative splicing and translational functions. Taken together, our results give support to the idea that miRNA regulation plays a key role in RA-induced neuroblastoma cell differentiation. The discovery of SFRS1 as direct target of miR-10a and -10b supports the emerging functional interaction between two post-transcriptional mechanisms, microRNAs and splicing, in the neuronal differentiation context. MicroRNAs (miRNAs, miRs)5 are an emerging class of small non-coding endogenous RNAs that are involved in multiple biological processes. These tiny regulatory elements are transcribed as primary longer transcripts, which are then processed by Dicer and Drosha complexes into 21-23-nt mature miRNAs. One strand of the mature miRNA is then incorporated into the RNA-induced silencing complex to regulate gene expression by targeting the 3Ј-untranslated region (3Ј-UTR) of mRNAs with consequent translational repression and/or target mRNA degradation. This mode of action demonstrates the great regulatory potential of miRNAs, as a unique mRNA can be targeted by diverse miRNAs, and conversely, each miRNA may have 100s of different target mRNAs (for review, see Refs. 1 and 2).In vertebrates, the highest variety of miRNAs is expressed in the brain than in any other tissue, suggesting an important role in nervous system development (3, 4). Recent studies have revealed the importance of miRNA expression in neural development and differentiation. For instance, it has been reported that Dicer-deleted mice exhibit malformations of the midbrain and cerebellum and failure of neural crest and dopaminergic differentiation (5). In addition, three brain-specific miRNAs (miR-124a, miR-9, and miR-132) have been proposed to regulate the transcription factor R...

Early leptin blockade predisposes fat-fed rats to overweight and modifies hypothalamic microRNAs

“…This miRNA is a potent inducer of neuroblastoma and muscle cell differentiation (Huang et al 2010, Foley et al 2011, Meseguer et al 2011. As leptin is involved in neuronal network organization in the hypothalamus, we suggest that blocking leptin action early in life promotes neuronal differentiation instead of proliferation leading to the impairment of energy homeostasis later in life as our findings suggest.…”

Retinoic acid signaling and neuronal differentiation

“…A sharp change in Numb isoform expression was observed in P19 embryonal carcinoma cells when neuronal differentiation was stimulated by RA [201]. The molecular mechanism of Numb alternative splicing remains an open question, although RA-induced differentiation alters splicing machinery in P19 cells [202] and in SH-SY5Y neuroblastoma cells [203].…”

Regulating the regulators: Serine/arginine‐rich proteins under scrutiny

“…Regarding the regulation of SRSF1 expression levels by miRNA, recent work from the Barettino laboratory (19) has shown that miR-10a and 10b directly target SRSF1 3 0 UTR, leading to SRSF1 mRNA destabilization during retinoic-acid induced differentiation of neuroblastoma cell line SH-SY5Y. In addition, changes in miR-10a and 10b levels alter SRSF1 regulatory activity both at alternative splicing and mRNA translation level, as evaluated by the analysis of tau exon 10 inclusion and translation of a reporter mRNA harboring an SRSF1-binding sequence, respectively.…”