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Multiple protein–protein interactions converging on the Prp38 protein during activation of the human spliceosome
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| Zeitschriftentitel: | RNA |
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| Personen und Körperschaften: | , , , , , , , , , |
| In: | RNA, 22, 2016, 2, S. 265-277 |
| Format: | E-Article |
| Sprache: | Englisch |
| veröffentlicht: |
Cold Spring Harbor Laboratory
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| Schlagwörter: |
| Zusammenfassung: | <jats:p>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 α-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.</jats:p> |
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| Umfang: | 265-277 |
| ISSN: |
1355-8382
1469-9001 |
| DOI: | 10.1261/rna.054296.115 |