Split-APEX implicates splicing factor SRSF1 and splicing helicases in ribosomal biogenesis

Vasileios Paschalis^1*Max F.K. Wills¹Philippe De Gusmao Araujo¹Christian Lucas¹Sumera Tubasum¹Shijie Cui¹Hesna Kara¹Carlos Bueno-Alejo¹Marina Santana Vega²Andrea Taladriz Sender³Zhengyun Zhao³Alexander Axer³Cyril Dominguez¹Alasdair W. Clark²Glenn Burley³Andrew J. Hudson¹Ian Eperon^1*

• ¹University of Leicester, Leicester, United Kingdom
• ²University of Glasgow, Glasgow, United Kingdom
• ³University of Strathclyde, Glasgow, United Kingdom

SR proteins are RNA-binding proteins with one or two RNA recognition motif (RRM)-type RNA-binding domains and a C-terminal region rich in arginine-serine dipeptides. They function in cellular processes ranging from transcription to translation. The best-known SR protein, SRSF1, modulates RNA splicing by stabilizing the binding of constitutive splicing factors, but there is also evidence that it participates in constitutive splicing reactions and is present in spliceosomal complexes. It has been shown recently that it interacts with DDX23, an RNA helicase that triggers the transition from complex pre-B to complex B during activation of the spliceosome. To identify in which other steps of spliceosome assembly and reaction it might be present, we have used split-APEX with SRSF1 and a number of helicases, each of the latter being involved in a particular step. Peroxidase activity should only be reconstituted if SRSF1 and the helicase were in contact, and the consequent biotinylation should reveal proteins in the vicinity. Our results show that all the helicases tested can complement SRSF1, but that the proximal proteins are very similar in all cases. Moreover, the proteins identified fall into two major classes: splicing-related proteins and ribosomal proteins. The results raise the possibility that SRSF1 and the canonical helicases have hitherto unsuspected collaborative roles in ribosomal assembly or translation.