Domain analysis of the Saccharomyces cerevisiae heterogeneous nuclear ribonucleoprotein, Nab2p. Dissecting the requirements for Nab2p-facilitated poly(A) RNA export.
Mature poly(A) RNA transcripts are exported from the nucleus in complex with heterogeneous nuclear ribonucleoproteins (hnRNPs). Nab2p is an essential Saccharomyces cerevisiae hnRNP protein that interacts with poly(A) RNA and shuttles between the nucleus and cytoplasm. Functional Nab2p is required for export of poly(A) RNA from the nucleus. The Nab2 protein consists of the following four domains: a unique N-terminal domain, a glutamine-rich domain, an arginine-glycine (RGG) domain, and a zinc finger domain. We generated Nab2p deletion mutants to analyze the contribution of each domain to the in vivo function of Nab2p. We first tested whether the deletion mutants could replace the essential NAB2 gene. We then examined the impact of these mutations on Nab2p localization, poly(A) RNA localization, and association of Nab2p with poly(A) RNA. Our analyses revealed that the N-terminal domain is required for nuclear export of both poly(A) RNA and Nab2p. We confirm that the RGG domain is important for Nab2p import in vivo. Finally, the zinc finger domain is critical for the interaction between Nab2p and poly(A) RNA in vivo. Our data support a model where Nab2p associates with poly(A) RNA in the nucleus through the zinc finger domain and facilitates the export of the poly(A) RNA through protein interactions mediated by the N-terminal domain.
Marfatia
Kavita A
KA
Department of Biochemistry, Graduate Program in Genetics and Molecular Biology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Crafton
Emily B
EB
Green
Deanna M
DM
Corbett
Anita H
AH
D000483:Alleles; D001120:Arginine / Q000737:chemistry; D002467:Cell Nucleus / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D017353:Gene Deletion; D005973:Glutamine / Q000737:chemistry; D015151:Immunoblotting; D017404:In Situ Hybridization, Fluorescence; D008853:Microscopy; D008957:Models, Genetic; D009154:Mutation; D029543:Nucleocytoplasmic Transport Proteins / Q000737:chemistry* / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011061:Poly A / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism; D013696:Temperature; D016335:Zinc Fingers
D003432:Cross-Linking Reagents; C080773:NAB2 protein, S cerevisiae; D029543:Nucleocytoplasmic Transport Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D005973:Glutamine; D011061:Poly A; D012313:RNA; D001120:Arginine