Phosphorylation of pICln by the autophagy activating kinase ULK1 regulates snRNP biogenesis and splice activity of the cell

The spliceosome, responsible for all mature protein-coding transcripts of eukaryotic intron-containing genes, consists of small uridine-rich nuclear ribonucleoproteins (UsnRNPs). The assembly of UsnRNPs de-pends, on one hand, on the arginine methylation of Sm proteins catalyzed by the PRMT5 complex. On the other hand, it depends on the phosphorylation of the PRMT5 subunit pICln by the Uncoordinated Like Kinase 1 (ULK1). In consequence, phosphorylation of pICln affects the stability of the UsnRNP assembly intermediate, the so-called 6 S complex. The detailed mechanisms of phosphorylation-dependent integrity and subsequent UsnRNP assembly of the 6 S complex in vivo have not yet been analyzed.By using a phospho-specific antibody against ULK1-dependent phosphorylation sites of pICln, we vi-sualize the intracellular distribution of phosphorylated pICln. Furthermore, we detect the colocaliphosphor-pICln1 with phospho-pICln by size-exclusion chromatography and immunofluorescence techniques. We also show that phosphorylated pICln is predominantly present in the 6 S complex. The addition of ULK1 to in vitro produced 6 S complex, as well as the reconstitution of ULK1 in ULK1-deficient cells, increases the efficiency of snRNP biogenesis. Accordingly, inhibition of ULK1 and the associated decreased pICln phos-phorylation lead to accumulation of the 6 S complex and reduction in the spliceosomal activity of the cell.(c) 2023 Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/ licenses/by-nc-nd/4.0/).

Automated summary

The spliceosome, consisting of UsnRNPs, plays a crucial role in the production of mature protein-coding transcripts of eukaryotic intron-containing genes. The assembly of UsnRNPs depends on the arginine methylation of Sm proteins by the PRMT5 complex and the phosphorylation of the PRMT5 subunit pICln by ULK1. However, the detailed mechanisms of phosphorylation-dependent integrity and UsnRNP assembly of the 6 S complex have not been fully analyzed yet.