Aberrant folding of a mutant Stat5b causes growth hormone insensitivity and proteasomal dysfunction

Dennis J. Chia, Ezhilkani Subbian, Teresa M. Buck, Vivian Hwa, Ron G. Rosenfeld, William R. Skach, Ujwal Shinde, Peter Rotwein

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26 Scopus citations

Abstract

A predicted alanine to proline substitution in Stat5b that results in profound short stature, growth hormone insensitivity, and immunodeficiency represents the first natural mutation of this transcription factor in a human. To understand the mechanisms responsible for these pathophysiological abnormalities, we have studied the biochemical and biophysical properties of the mutant Stat5b molecule. In a cellular reconstitution model growth hormone robustly stimulated tyrosine phosphorylation and transcriptional activity of wild-type Stat5b while Stat5bA630P was minimally modified and did not promote reporter gene expression. Steady state levels of Stat5bWT were ∼3-fold higher than Stat5bA630P in cell extracts prepared with nonionic detergents. Although initial rates of biosynthesis of both proteins were similar, pulse-chase experiments established that the apparent half-life of newly synthesized soluble Stat5bA630P was <15% of Stat5bWT (3.5 h versus >24 h). Stat5bA630P accumulated in cells primarily in cytoplasmic inclusion bodies. Structural analysis of the isolated SH2 domain containing the A630P mutation showed that it resembled the wild-type SH2 segment but that it exhibited reduced thermodynamic stability and slower folding kinetics, displayed an increased hydrophobic surface, and was prone to aggregation in solution. Our results are compatible with a model in which Stat5bA630P is an inactive transcription factor by virtue of its aberrant folding and diminished solubility triggered by amisfolded SH2 domain. The potential for aggregation and formation of cytoplasmic inclusions raises the possibility that Stat5bA630P could produce additional defects through inhibition of proteasome function.

Original languageEnglish (US)
Pages (from-to)6552-6558
Number of pages7
JournalJournal of Biological Chemistry
Volume281
Issue number10
DOIs
StatePublished - Mar 10 2006

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Chia, D. J., Subbian, E., Buck, T. M., Hwa, V., Rosenfeld, R. G., Skach, W. R., Shinde, U., & Rotwein, P. (2006). Aberrant folding of a mutant Stat5b causes growth hormone insensitivity and proteasomal dysfunction. Journal of Biological Chemistry, 281(10), 6552-6558. https://doi.org/10.1074/jbc.M510903200