Abstract
A variety of cytokines and growth factors act through an induction of gene expression mediated by a family of latent transcription factors called STAT (signal transducers and activators of transcription) proteins. Ligand-induced tyrosine phosphorylation of the STATs promotes their homodimer and heterodimer formation and subsequent nuclear translocation. We demonstrate here that STAT protein heterocomplexes exist prior to cytokine treatment. When unstimulated HeLa cells are ruptured in hypotonic buffer without salt or detergent, immunoadsorption of either STAT1 or STAT2 from the resulting cytosol yields coimmunoadsorption of the other STAT protein. Similarly, STAT1-STAT3 heterocomplexes are coimmunoadsorbed from hypotonic cytosol. STAT1 and STAT2 or STAT1 and STAT3 translated in reticulocyte lysate spontaneously form heterocomplexes when the translation lysates are mixed at 0 degrees C. Our data suggest that interferon-alpha /beta-induced tyrosine phosphorylation increases the stability of a preexisting, latent, STAT1-STAT2 signaling complex. Newly translated STAT1 binds in equilibrium fashion to STAT2 and STAT3, but we show that STAT2 and STAT3 exist in separate heterocomplexes with STAT1, consistent with a model in which STAT1 contains a common binding site for other STAT proteins.
Highlights
Ʈ To whom correspondence should be addressed: Dept. of Pharmacology, 1301 Medical Science Research Bldg
We found that rupture of HeLa cells in a hypotonic buffer followed by immunoadsorption of STAT1 yields coimmunoadsorption of STAT2 and vice versa
STAT1 and STAT2 Are Preassociated—In the experiment of Fig. 1, HeLa cells were ruptured under two conditions: 1) in a hypotonic buffer that has been used to isolate heat shock protein heterocomplexes in which some of the proteins are rather weakly bound [15] or 2) in the same buffer but with the addition of 1% Triton X-100 and 150 mM NaCl
Summary
Ʈ To whom correspondence should be addressed: Dept. of Pharmacology, 1301 Medical Science Research Bldg. It is known that interaction of IFN-␣ or IFN- with the IFN-␣/ receptor activates JAK1 and Tyk with resulting tyrosine phosphorylation of the 91-kDa STAT1 and the 113-kDa STAT2 proteins, which are thought to form a STAT heterodimer that associates with a 48-kDa DNA-binding protein. This multiprotein unit is called the interferon-stimulated gene factor 3, and it appears to be the primary positive regulator of interferon-stimulated response element-controlled genes [3,4,5]. This may imply that STAT1 contains a common binding site for other STAT proteins
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