Srsf3 mediates alternative RNA splicing downstream of PDGFRα signaling in the facial mesenchyme

Abstract

Signaling through the platelet-derived growth factor receptor alpha (PDGFRα) is critical for mammalian craniofacial development, though the mechanisms by which the activity of downstream intracellular effectors is regulated to mediate gene expression changes have not been defined. We find that the RNA-binding protein Srsf3 is phosphorylated at Akt consensus sites downstream of PI3K-mediated PDGFRα signaling in mouse palatal mesenchyme cells, leading to its nuclear translocation. We further demonstrate that ablation of Srsf3 in the mouse neural crest lineage leads to facial clefting due to defective cranial neural crest cell proliferation and survival. Finally, we show that Srsf3 regulates the alternative RNA splicing of transcripts encoding protein kinases in the mouse facial process mesenchyme to regulate PDGFRα-dependent intracellular signaling. Collectively, our findings reveal that alternative RNA splicing is an important mechanism of gene expression regulation downstream of PI3K/Akt-mediated PDGFRα signaling in the facial mesenchyme and identify Srsf3 as a critical regulator of craniofacial development.

The Role of RNA-Binding Proteins in Vertebrate Neural Crest and Craniofacial Development

“…While the target transcripts of individual RBPs do not appear to overlap in the context of craniofacial morphogenesis, even within related tissues, common cellular mechanisms are regulated by these RBPs to contribute to proper development. For example, only six transcripts are commonly differentially alternatively spliced ( Exoc1 , Meg3 , Nfya , Smarca2 , Tpm1 and Uap1 ) and only two transcripts are commonly differentially expressed ( Lama5 and Thrb ) between E12.5 Rbfox2 fl/fl ;Pax3-Cre +/Tg craniofacial tissue and E11.5 Srsf3 fl/fl ;Wnt1-Cre +/Tg maxillary process mesenchyme tissue relative to their respective controls [ 48 , 58 ]. Notably, however, knockdown or ablation of Esrp1/2 , Rbfox2 , Srsf3 and Ddx3x each results in altered expression of transcripts encoding components of developmental signaling pathways.…”

“…Notably, however, knockdown or ablation of Esrp1/2 , Rbfox2 , Srsf3 and Ddx3x each results in altered expression of transcripts encoding components of developmental signaling pathways. These include the FGF, WNT, SHH, BMP and TGFβ signaling pathways in the case of ESRP1/2 [ 30 ], and the TGFβ, PDGF and Wnt signaling pathways for RBFOX2 [ 48 ], SRSF3 [ 58 ] and Ddx3x [ 70 , 72 ], respectively. These findings raise the intriguing possibility that RNA processing mediated by RBPs serves to regulate both intracellular and intercellular signaling during craniofacial development, often in feedback loops involving developmental signaling pathways and the phosphorylation of intracellular effectors.…”

“…These defects stem from reduced proliferation of the mesenchyme underlying the cranial neural folds at E8.0 and increased cell death in the mesenchyme of pharyngeal arch 1 at E9.5. Additionally, conditional ablation of Srsf3 in the murine NC lineage leads to a significant increase in expression of multiple genes involved in NC differentiation, including Col2a1 and Zic5 [ 58 ]. PI3K is the primary effector of PDGFRα signaling during skeletal development in the mouse, and disruption of this axis leads to palatal clefting [ 59 , 60 ].…”

“…SRSF3 is phosphorylated at AKT consensus sites in response to PI3K-mediated PDGFRα signaling in immortalized mouse embryonic palatal mesenchyme cells, driving translocation of phosphorylated SRSF3 into the nucleus where alternative RNA splicing occurs. Relatedly, SRSF3 coordinates the alternative RNA splicing of transcripts encoding protein serine/threonine kinases in the E11.5 maxillary process mesenchyme to regulate PDGFRα-dependent intracellular signaling, specifically, phosphorylation of ERK1/2 [ 58 ].…”

Biological function and molecular mechanism of SRSF3 in cancer and beyond (Review)

“…Overexpression of protein phosphatase, Mg 2+ /Mn 2+ dependent 1G (PPM1G) promotes the dephosphorylation of SRSF3 and changes the alternative splicing patterns of genes related to the cell cycle and transcriptional regulation in HCC cells ( 55 ). SRSF3 is phosphorylated at AKT consensus sites downstream of PI3K-mediated platelet-derived growth factor receptor α signaling in mouse palatal mesenchymal cells, resulting in its nuclear translocation ( 56 ). However, SRSF3 appears to be hypo-phosphorylated and stable in cells ( 57 ), which may be a way to maintain cell homeostasis.…”