The KH domains of Xenopus Vg1RBP mediate RNA binding and self-association.
Xenopus Vg1 mRNA is localized to the vegetal cortex during oogenesis in a process involving microtubules and microfilaments and proteins that specifically recognize the vegetal localization element (VLE) within the 3' untranslated region. One of the best characterized VLE-binding proteins is Vg1RBP or Vera. Primary sequence analysis of Vg1RBP and its homologs suggests that most of its open reading frame is occupied by RNA-binding modules, including two RRMs and four KH domains, arranged as three pairs of didomains. In the first detailed domain analysis of Vg1RBP, we show that the interaction of Vg1RBP with the VLE requires both KH didomains, but not the RRM didomain, and moreover that the KH didomains contribute cooperatively to RNA binding. In the full-length protein, individual KH domains display significant redundancy, and their relative importance appears to vary with the RNA target. We also demonstrate that the KH34 didomain mediates Vg1RBP self-association, which is stabilized by RNA, and occurs in vivo as well as in vitro. Altogether, our findings highlight the importance of multiple KH domains in mediating RNA-protein and protein-protein interactions in the formation of a stable complex of Vg1RBP and Vg1 mRNA.
Git
Anna
A
Department of Biochemistry, University of Cambridge, United Kingdom.
Standart
Nancy
N
D020413:3' Untranslated Regions; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D005260:Female; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009865:Oocytes / Q000502:physiology; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D016212:Transforming Growth Factor beta; D014981:Xenopus / Q000235:genetics / Q000378:metabolism*; D029867:Xenopus Proteins
D020413:3' Untranslated Regions; C513103:GDF1 protein, Xenopus; D006023:Glycoproteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D029867:Xenopus Proteins; D012313:RNA