RBM4 Regulates Neuronal Differentiation of Mesenchymal Stem Cells by Modulating Alternative Splicing of Pyruvate Kinase M

ABSTRACT

RBM4 promotes differentiation of neuronal progenitor cells and neurite outgrowth of cultured neurons via its role in splicing regulation. In this study, we further explored the role of RBM4 in neuronal differentiation. During neuronal differentiation, energy production shifts from glycolysis to oxidative phosphorylation. We found that the splice isoform change of the metabolic enzyme pyruvate kinase M (PKM) from PKM2 to PKM1 occurs during brain development and is impaired in RBM4-deficient brains. The PKM isoform change could be recapitulated in human mesenchymal stem cells (MSCs) during neuronal induction. Using a PKM minigene, we demonstrated that RBM4 plays a direct role in regulating alternative splicing of PKM. Moreover, RBM4 antagonized the function of the splicing factor PTB and induced the expression of a PTB isoform with attenuated splicing activity in MSCs. Overexpression of RBM4 or PKM1 induced the expression of neuronal genes, increased the mitochondrial respiration capacity in MSCs, and, accordingly, promoted neuronal differentiation. Finally, we demonstrated that RBM4 is induced and is involved in the PKM splicing switch and neuronal gene expression during hypoxia-induced neuronal differentiation. Hence, RBM4 plays an important role in the PKM isoform switch and the change in mitochondrial energy production during neuronal differentiation.

KEYWORDS:

• alternative splicing
• hypoxia
• mesenchymal stem cells
• neuronal differentiation
• pyruvate kinase M

Supplemental material for this article may be found at https://doi.org/10.1128/MCB.00466-16.

ACKNOWLEDGMENTS

We thank D. Dhananjaya for recombinant protein and for discussion, Yi-Shuian Huang for an expression vector, and the Core Facility of the Institute of Biomedical Sciences, Academia Sinica, for technical assistance.

C.-H.S. designed and performed the majority of experiments and contributed to manuscript preparation. K.-Y.H. provided mouse embryos and performed RT-qPCR. S.-C.H. provided MSCs and cell lines. W.-Y.T. oversaw the project, designed the experiments, and wrote the manuscript.

We declare that we have no conflicts of interest.

This work was supported by Ministry of Science and Technology grant 104-2321-B-001-066.