Multiple protein-protein interactions converging on the Prp38 protein during activation of the human spliceosome

作者：Schuetze Tonio; Ulrich Alexander K C; Apelt Luise; Will Cindy L; Bartlick Natascha; Seeger Martin; Weber Gert; Luehrmann Reinhard; Stelzl Ulrich^*; Wahl Markus C^*

来源：RNA-A Publication of the RNA Society, 2016, 22(2): 265-277.

DOI：10.1261/rna.054296.115

• pre-mRNA processing factor 38
• pre-mRNA splicing
• protein-protein interactions
• spliceosomal B complex
• spliceosome
• yeast two-hybrid analysis

Spliceosomal Prp38 proteins contain a conserved amino-terminal domain, but only higher eukaryotic orthologs also harbor a carboxy-terminal RS domain, a hallmark of splicing regulatory SR proteins. We show by crystal structure analysis that the amino-terminal domain of human Prp38 is organized around three pairs of antiparallel alpha-helices and lacks similarities to RNA binding domains found in canonical SR proteins. Instead, yeast two-hybrid analyses suggest that the amino-terminal domain is a versatile protein protein interaction hub that possibly binds 12 other spliceosomal proteins, most of which are recruited at the same stage. as Prp38. By quantitative, alanine surface-scanning two-hybrid screens and biochemical analyses we delineated four distinct interfaces on the Prp38 amino-terminal domain. In vitro interaction assays using recombinant proteins showed that Prp38 can bind at least two proteins simultaneously via two different interfaces. Addition of excess Prp38 amino-terminal domain to in vitro splicing assays, but not of an interaction-deficient mutant, stalled splicing at a precatalytic stage. Our results show that human Prp38 is an unusual SR protein, whose amino-terminal domain is a multi-interface protein protein interaction platform that might organize the relative positioning of other proteins during splicing.

• 出版日期2016-2