ZFP207 interacts with U1 snRNP to promote spliceosome assembly via phase separation [CRISPR screen]

U1 snRNP plays an essential role in initiating spliceosome assembly, yet the mechanism underlying its synergy with other splicing regulators for efficient spliceosome assembly remains elusive. Here we identify ZFP207 as a key regulator of U1 snRNP function that substantially promotes spliceosome assembly. Acute depletion of ZFP207 recapitulates the molecular phenotypes observed with the depletion of SNRNP70, a core component of U1 snRNP. Mechanistically, the N-terminal zinc finger domains of ZFP207 directly bind to U1 snRNA, while its C-terminus undergoes phase separation via intrinsically disordered regions (IDRs). The coordination between the N-terminus and C-terminus of ZFP207 drives the formation of biomolecular condensate with U1 snRNP, which creates a molecular environment to promote spliceosome assembly by facilitating the interactions between U1 snRNP and other splicing regulators. Collectively, our study demonstrates the critical role of ZFP207-mediated phase separation in ensuring proper U1 snRNP function and spliceosome assembly. To identify other regulators of U1 snRNP and U1-PAS-mediated gene repression, we sought to conduct a genome-wide CRISPR interference (CRISPRi) screen on U1-PAS reporter cells, selecting cells in which reporter genes become upregulated. We performed two independent screens in parallel and amplified the genomic sequences encoding the sgRNA from surviving cells after 5 and 8 days of blasticidin selection, as well as non-selected control cells for deep-sequencing analysis. Gene enrichment analysis for CRISPRi screen data (D5 vs D0, D8 vs D0) using MAGeCK (model-based analysis of genome-wide CRISPR–Cas9 knockout).