Ecd promotes U5 snRNP maturation and Prp8 stability

Abstract

Pre-mRNA splicing catalyzed by the spliceosome represents a critical step in the regulation of gene expression contributing to transcriptome and proteome diversity. The spliceosome consists of five small nuclear ribonucleoprotein particles (snRNPs), the biogenesis of which remains only partially understood. Here we define the evolutionarily conserved protein Ecdysoneless (Ecd) as a critical regulator of U5 snRNP assembly and Prp8 stability. Combining Drosophila genetics with proteomic approaches, we demonstrate the Ecd requirement for the maintenance of adult healthspan and lifespan and identify the Sm ring protein SmD3 as a novel interaction partner of Ecd. We show that the predominant task of Ecd is to deliver Prp8 to the emerging U5 snRNPs in the cytoplasm. Ecd deficiency, on the other hand, leads to reduced Prp8 protein levels and compromised U5 snRNP biogenesis, causing loss of splicing fidelity and transcriptome integrity. Based on our findings, we propose that Ecd chaperones Prp8 to the forming U5 snRNP allowing completion of the cytoplasmic part of the U5 snRNP biogenesis pathway necessary to meet the cellular demand for functional spliceosomes.

Ecdysoneless Protein Regulates Viral and Cellular mRNA Splicing to Promote Cervical Oncogenesis

“…Our recent publication showed multiple components of the mRNA splicing machinery are associated with ECD (12), suggesting a role for ECD in cellular and potential viral mRNA splicing. Such a role was consistent with our' and others' findings that ECD associates with the core spliceosome scaffold protein PRPF8 (11)(12)(13).…”

“…It is notable that a major binding partner of ECD is R2TP complex (9), suggesting ECD may link the spliceosome to R2TP/PAQosome, with latter involved in remodeling necessary during mRNA splicing. Consistent with this idea, Drosophila Ecd promotes the U5 snRNP maturation (13). The R2TP/PAQosome complex is also known to regulate the assembly of TERT (38).…”

“…Author Manuscript Published OnlineFirst on October 20, 2021; DOI: 10.1158/1541-7786.MCR-21-0567 PRPF8, an essential scaffolding component of the spliceosome machinery (9,(11)(12)(13), suggesting a role of ECD in splicing. We recently showed that ECD is in a complex with proteins involved in mRNA export and splicing (12).…”

CryoEM of RUVBL1–RUVBL2–ZNHIT2, a complex that interacts with pre-mRNA-processing-splicing factor 8

“…Two factors, Zinc finger HIT domain-containing protein 2 (ZNHIT2) ( 4 ) and ECD (Protein ecdysoneless homolog) ( 20 , 21 ), have recently been found to interact with the R2TP chaperone system to regulate the biogenesis of PRPF8 and U5 snRNP. ZNHIT2 interacts with RUVBL2 and regulates the composition of U5 snRNP and splicing ( 4 ).…”

“…PRPF8-based purifications detect RUVBL1–RUVBL2 and ZNHIT2 but no other components of the R2TP complex, which has been interpreted as indication that ZNHIT2 competes for RPAP3 binding to RUVBL1–RUVBL2 ( 4 , 22 ). ECD also interacts with several components of R2TP and regulates the stability of PRPF8 and the assembly of U5 snRNP ( 20 , 21 ). ECD contains CK2 phosphorylation sites at consensus motifs recognized by PIH1D1, but the functional relevance of the interaction between ECD and PIH1D1 is unclear as it seems dispensable for at least some of the functions assigned to ECD.…”