Conserved functional antagonism of CELF and MBNL proteins controls stem cell-specific alternative splicing in planarians

Abstract

In contrast to transcriptional regulation, the function of alternative splicing (AS) in stem cells is poorly understood. In mammals, MBNL proteins negatively regulate an exon program specific of embryonic stem cells; however, little is known about the in vivo significance of this regulation. We studied AS in a powerful in vivo model for stem cell biology, the planarian Schmidtea mediterranea. We discover a conserved AS program comprising hundreds of alternative exons, microexons and introns that is differentially regulated in planarian stem cells, and comprehensively identify its regulators. We show that functional antagonism between CELF and MBNL factors directly controls stem cell-specific AS in planarians, placing the origin of this regulatory mechanism at the base of Bilaterians. Knockdown of CELF or MBNL factors lead to abnormal regenerative capacities by affecting self-renewal and differentiation sets of genes, respectively. These results highlight the importance of AS interactions in stem cell regulation across metazoans.Stem cells are specialized cells found in all animals that can develop into several different types of mature cells. Stem cells are therefore well suited for maintaining organs that are in heavy use, such as the intestine, and for regenerating tissues that are prone to injury, like the skin.One reason why stem cells differ from mature cell types is because they activate, or “express”, different sets of genes. In addition, many genes can be expressed as one of several versions. These variants, also known as isoforms, are generated by a process called alternative splicing. In mature cells in mammals, a group of proteins called the MBNL proteins help to prevent the expression of gene isoforms that are characteristic to stem cells.The adult flatworm Schmidtea mediterranea contains stem cells that can regenerate any part of the body. Solana, Irimia et al. have now investigated whether alternative splicing is important for controlling how the worm’s stem cells behave. After establishing which gene isoforms are expressed in the stem cells and the mature cells, the levels of different sets of proteins that control alternative splicing were experimentally reduced.The results indicate that just as seen in mammals, the MBNL proteins reduce the expression of stem cell-related gene isoforms in the flatworms. Furthermore, Solana, Irimia et al. found that another protein called CELF counteracts MBNL proteins by helping to express gene isoforms that are active in stem cells.The interplay between the MBNL and CELF proteins has also been observed in human cells. Thus, it appears that this way of controlling alternative splicing is common to flatworms and mammals and is therefore evolutionarily ancient. This suggests that other similar ways of controlling stem cells by interactions between regulatory proteins might be working in all animal stem cells. Further studies are now needed to investigate these control proteins.