Isolation and characterization of cDNA encoding mouse RNA polymerase II subunit RPB14

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Y Nishi1, K Yamamoto, Y Yao

  • 1Department of Biochemistry, Saitama Medical School, Iruma-gun, Japan.

Gene
|March 18, 1997
PubMed

Insights

Researchers identified a new mouse RNA polymerase II subunit, mRPB14, using yeast two-hybrid screening. This finding suggests conserved enzyme organization between yeast and mammals.

Area of Science:

  • Molecular Biology
  • Biochemistry

Background:

  • RNA polymerase I (Pol I) and RNA polymerase II (Pol II) are crucial enzymes for transcription.
  • Understanding the subunit composition of these polymerases is key to elucidating their function.

Purpose of the Study:

  • To identify novel subunits of mouse RNA polymerase II.
  • To investigate the conservation of RNA polymerase subunit organization between yeast and mammals.

Main Methods:

  • Yeast two-hybrid system was employed using the 40-kDa subunit of mouse RNA polymerase I (mRPA40) as bait.
  • A specific antibody was generated against the isolated cDNA product (mRPB14).
  • Immunoprecipitation assays were performed using the mRPB14 antibody.

Main Results:

  • A mouse cDNA, mRPB14, was isolated, encoding a protein homologous to yeast RNA polymerase II subunit B12.5.
  • The anti-mRPB14 antibody specifically recognized a 14-kDa polypeptide in purified mammalian RNA polymerase II.
  • The antibody co-immunoprecipitated the largest subunit of mouse RNA polymerase II, confirming mRPB14's association.

Conclusions:

  • Biochemical evidence confirms mRPB14 as a specific subunit of mouse RNA polymerase II.
  • The study indicates conserved subunit organization of RNA polymerase between yeast and mouse.
  • The potential role of the alpha-motif in protein-protein interactions within RNA polymerase is suggested.

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