Stimulation of mRNA export by an F-box protein, Mdm30p, in vivo.
Mdm30p, a nucleus-encoded F-box protein, which binds to the substrate for ubiquitin-mediated proteolysis, is involved in maintenance of fusion-competent mitochondria for various cellular functions. Recently, Mdm30p has been implicated in regulation of gene expression. However, its mode of action in gene regulation is not clearly known in vivo. With this view, we have systematically analyzed here the role of Mdm30p in regulation of transcriptional initiation, elongation, mRNA processing, and export in Saccharomyces cerevisiae, using a formaldehyde-based in vivo cross-linking and chromatin immunoprecipitation assay in conjunction with RT-PCR and fluorescence in situ hybridization. We show that Mdm30p is dispensable for formation of the preinitiation complex assembly, association of elongating RNA polymerase II, and recruitment of mRNA capping enzyme, cap-binding complex, and 3' end formation machinery at the transcriptionally active genes such as ADH1, PHO84, and RPS5. Intriguingly, we find that Mdm30p facilitates the recruitment of the transcription-export complex at these genes. Consistently, the export of mRNAs of these genes is significantly impaired in the absence of Mdm30p as revealed by fluorescence in situ hybridization and RT-PCR analysis of cytoplasmic mRNA. However, such an impaired mRNA export is not dependent on mitochondrial fusion, as the deletion of FZO1, an essential gene for mitochondrial fusion, does not alter the export of ADH1, PHO84, and RPS5 mRNAs. Together, our data demonstrate that Mdm30p selectively controls mRNA export independently of mitochondrial fusion, revealing a novel function of an F-box protein in mRNA export.
Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, 1245 Lincoln Drive, Carbondale, IL 62901, USA.
Abhijit
A
Shukla
Geetha
G
Durairaj
Jessica
J
Schneider
Zhen
Z
Duan
Thomas
T
Shadle
Sukesh R
SR
Bhaumik
D044783:F-Box Proteins / Q000235:genetics / Q000502:physiology*; D005786:Gene Expression Regulation; D008928:Mitochondria / Q000378:metabolism / Q000648:ultrastructure; D034443:RNA Transport; D012333:RNA, Messenger / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic
D044783:F-Box Proteins; C481653:Mdm30 protein, S cerevisiae; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins