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Interactions of Cdk7 and Kin28 with Hint/PKCI-1 and Hnt1 histidine triad proteins

N Korsisaari1, T P Mäkelä

  • 1Haartman Institute & Biocentrum Helsinki, P. O. Box 21, University of Helsinki, 00014 Helsinki, Finland.

The Journal of Biological Chemistry
|August 26, 2000
PubMed

Insights

Researchers discovered a new interaction between Cyclin-dependent kinase 7 (Cdk7) and histidine triad (HIT) proteins, Hint and Hnt1. This finding suggests a role for HIT proteins in regulating Cdk7 and Kin28 kinase functions.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Cyclin-dependent kinase 7 (Cdk7) is crucial for transcription as part of TFIIH and as a Cdk-activating kinase (CAK).
  • Cdk7 phosphorylates the RNA polymerase II C-terminal domain (CTD), regulating transcription.
  • The function of histidine triad (HIT) proteins in relation to Cdk7 is not well understood.

Purpose of the Study:

  • To investigate potential novel interactions with Cdk7.
  • To characterize the functional relationship between Cdk7 and HIT family proteins.

Main Methods:

  • Yeast two-hybrid screening to identify interacting proteins.
  • Co-immunoprecipitation assays to confirm physical interactions.
  • Subcellular localization studies.
  • Genetic interaction studies in Saccharomyces cerevisiae using gene disruption and temperature-sensitive alleles.

Main Results:

  • A novel physical interaction was identified between Cdk7 and the HIT protein Hint/PKCI-1.
  • The interaction was validated through co-immunoprecipitation and observed to cause partial nuclear relocalization of Hint upon Cdk7 overexpression.
  • This physical association is independent of cyclin H binding or Cdk7 kinase activity.
  • The interaction is conserved in yeast, with Cdk7's homolog Kin28 interacting with the HIT protein Hnt1.
  • Genetic disruption of HNT1 and a temperature-sensitive KIN28 allele in yeast resulted in cell elongation and reduced colony formation, indicating a genetic interaction.

Conclusions:

  • Histidine triad proteins Hint and Hnt1 physically and genetically interact with Cdk7 and Kin28, respectively.
  • These findings suggest a regulatory role for HIT proteins in the functions of Cdk7 and Kin28.
  • This study uncovers a new layer of regulation for key transcription and cell cycle kinases.

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