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Rpb7 subunit of RNA polymerase II interacts with an RNA-binding protein involved in processing of transcripts.

ABSTRACT: Rpb4-Rpb7, a dissociable subcomplex of RNA polymerase II (pol II), is required for transcription initiation. To understand the role of Rpb7 in transcription initiation or other processes in transcription, we carried out a two-hybrid screen for proteins that interact with Rpb7 of the fission yeast Schizosaccharomyces pombe. The screen identified the S.pombe homolog of the Saccharomyces cerevisiae Nrd1, an RNA-binding protein implicated in 3' end formation of small nucleolar and small nuclear RNAs transcribed by pol II. The S.pombe protein, named Seb1 for seven binding, was essential for cell viability, and bound directly to Rpb7 in vitro. Saccharomyces cerevisiae Rpb7 also interacted with Nrd1, indicating that the interaction is conserved in evolution. Glu166 and/or Asp167 of S.pombe Rpb7, residues near the C-terminus of the 172 amino acid protein, were found to be important for its interaction with Seb1. Our results suggest that Rpb7 may function to anchor a processing factor to the pol II apparatus, thereby coupling RNA processing to transcription. The role for Rpb7 is consistent with its location in the pol II complex determined by recent structural studies.

SUBMITTER: Mitsuzawa H

PROVIDER: S-EPMC169969 | biostudies-literature | 2003 Aug

REPOSITORIES: biostudies-literature

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Rpb7 subunit of RNA polymerase II interacts with an RNA-binding protein involved in processing of transcripts.

Mitsuzawa Hiroshi H Kanda Emi E Ishihama Akira A

Nucleic acids research 20030801 16

Rpb4-Rpb7, a dissociable subcomplex of RNA polymerase II (pol II), is required for transcription initiation. To understand the role of Rpb7 in transcription initiation or other processes in transcription, we carried out a two-hybrid screen for proteins that interact with Rpb7 of the fission yeast Schizosaccharomyces pombe. The screen identified the S.pombe homolog of the Saccharomyces cerevisiae Nrd1, an RNA-binding protein implicated in 3' end formation of small nucleolar and small nuclear RNAs ...[more]

PMID: 12907709

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Project description:Rpb11 subunit of RNA polymerase II of Eukaryotes is related to N-terminal domain of eubacterial ? subunit and forms a complex with Rpb3 subunit analogous to prokaryotic ?2 homodimer, which is involved in RNA polymerase assembly and promoter recognition. In humans, a POLR2J gene family has been identified that potentially encodes several hRPB11 proteins differing mainly in their short C-terminal regions. The functions of the different human specific isoforms are still mainly unknown. To further characterize the minor human specific isoform of RNA polymerase II subunit hRPB11b?, the only one from hRPB11 (POLR2J) homologues that can replace its yeast counterpart in vivo, we used it as bait in a yeast two-hybrid screening of a human fetal brain cDNA library. By this analysis and subsequent co-purification assay in vitro, we identified transcription factor ATF4 as a prominent partner of the minor RNA polymerase II (RNAP II) subunit hRPB11b?. We demonstrated that the hRPB11b? interacts with leucine b-Zip domain located on the C-terminal part of ATF4. Overexpression of ATF4 activated the reporter more than 10-fold whereas co-transfection of hRPB11b? resulted in a 2.5-fold enhancement of ATF4 activation. Our data indicate that the mode of interaction of human RNAP II main (containing major for of hRPB11 subunit) and minor (containing hRPB11b? isoform of POLR2J subunit) transcription enzymes with ATF4 is certainly different in the two complexes involving hRPB3-ATF4 (not hRPB11a-ATF4) and hRpb11b?-ATF4 platforms in the first and the second case, respectively. The interaction of hRPB11b? and ATF4 appears to be necessary for the activation of RNA polymerase II containing the minor isoform of the hRPB11 subunit (POLR2J) on gene promoters regulated by this transcription factor. ATF4 activates transcription by directly contacting RNA polymerase II in the region of the heterodimer of ?-like subunits (Rpb3-Rpb11) without involving a Mediator, which provides fast and highly effective activation of transcription of the desired genes. In RNA polymerase II of Homo sapiens that contains plural isoforms of the subunit hRPB11 (POLR2J), the strength of the hRPB11-ATF4 interaction appeared to be isoform-specific, providing the first functional distinction between the previously discovered human forms of the Rpb11 subunit.

Dataset Information

Rpb7 subunit of RNA polymerase II interacts with an RNA-binding protein involved in processing of transcripts.

Publications

Rpb7 subunit of RNA polymerase II interacts with an RNA-binding protein involved in processing of transcripts.

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