In Vivo mutational analysis of the DNA binding domain of the tissue-specific transcription factor, Pit-1

Jie Liang, Scott Moye-Rowley, Richard A. Maurer

Research output: Contribution to journalArticle

24 Scopus citations

Abstract

Pit-1 is a member of the POU family of transcription factors, which contain a bipartite DNA binding domain. The DNA binding domain consists of a POU-specific domain and a POU homeodomain. Each of the subdomains can interact with DNA independently, but both subdomains are required for high affinity, sequence-specific DNA binding. To examine the contributions of individual amino acids to the function of the DNA binding domain of Pit-1, we developed an approach involving random, in vitro mutagenesis followed by functional screening in Saccharomyces cerevisiae. Using this strategy, we identified a number of point mutations that altered the function of the Pit-1 DNA binding domain. Mutations that altered Pit-1 function were found in both the POU-specific and the POU homeodomain. Most of the mutations involve amino acid residues that are conserved in POU factors. One of the more frequent kinds of mutation affected residues located in the hydrophobic core of the protein. Another common mutation involved amino acids that are thought to make specific contacts with DNA. These mutations define a number of amino acid residues that are important for the function of the DNA binding domain of Pit-1.

Original languageEnglish (US)
Pages (from-to)25520-25525
Number of pages6
JournalJournal of Biological Chemistry
Volume270
Issue number43
DOIs
StatePublished - Oct 27 1995

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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