Spacing of palindromic half sites as a determinant of selective STAT (signal transducers and activators of transcription) DNA binding and transcriptional activity.

Abstract

Signal transducers and activators of transcription (STAT proteins) bind to palindromic sequence elements related to interferon y (IFN-y) activation sites, which were first identified in the promoters of IFN-'yinducible genes. Although the sequences of the natural palindromic STAT-binding elements vary considerably, they conform to the general structure TT(N)5AA. We have systematically examined the effects of the spacing between the TT and AA core half sites on the binding of the STAT complexes activated by IFN-y, interleukin (IL) 6, granulocytemacrophage colony-stimulating factor, and IL-4. We show that (i) as suggested earlier, a core palindromic TT-AA motifwith a 5-bp spacing displays general STAT binding, (ii) a palindromic motif with a spacing of 4 bp selectively binds to complexes containing Stat3, and (iii) a motif with a 6-bp spacing selectively binds the STAT complexes activated by IL-4. We have examined natural elements in the promoters of cytokine-responsive genes that differ in half-site spacing and found that they display binding properties predicted from the synthetic binding sites. Furthermore, the observed differential selective binding characteristics for the most part correlate with the ability to mediate transcriptional activation of transfected test genes in response to the cytokines tested. Our results thus demonstrate that the specificity of STAT-directed transcription in response to particular cytokines or cytokine families depends in part on the spacing of half sites within the conserved response element sequence.

Unmasking by soluble IL-6 receptor of IL-6 effect on metastatic melanoma: growth inhibition and differentiation of B16-F10.9 tumor cells

“…Myeloleukemic M1 cells which stop growing and di erentiate with IL-6, had similar long-lasting Stat3 activation and biphasic induction of complex C or III (Nakajima et al, 1996). While most studies focused on the early Stat response around 15 min (Seidel et al, 1995;Wegenka et al, 1994;Zhong et al, 1994), it is clear that Stat complexes can show sustained and elaborate activation such as described here with the Figure 4 Transcriptional activities induced by IL-6 and sIL-6R in F10.9 melanoma cells. The luciferase reporter genes (see Materials and methods) were transfected into F10.9 cells and 24 h later the cells were treated as indicated (IL-6, 40 ng/ml; sIL-6R, 2 mg/ml).…”

Anti-sense oligonucleotide of p21(waf1/cip1) prevents Interleukin 4-mediated elevation of p27(kip1) in low grade astrocytoma cells

“…Binding sites for STAT1, another member of the STAT family, have been detected within the p21(waf1/cip1) promoter region, and activation of STAT1 and p21(waf1/cip1) are linked in some cells growth-arrested by EGF or ®broblast growth factor (Chin et al, 1996;Johnson et al, 1998). Interestingly, a DNA sequence recognized by STAT1 (TTC{N 3 }GAA) can also be bound by STAT6 (Seidel et al, 1995). That IL-4 signals STAT6 phosphorylation in both IL-4 responsive RTLGA and unresponsive WITG3 cells in our studies, however, suggests that activation of STAT6 alone is not su cient for growth arrest.…”

The role of STATs in myeloid differentiation and leukemia

“…Firstly, upon cytokine stimulation the activated STAT transcription factors are present both as homo-dimers and hetero-dimers (Leonard and O'Shea, 1998). Secondly, di erent STAT binding sequences have di erent a nities for the di erent homo-and hetero-dimers Seidel et al, 1995). Thirdly, besides tyrosine phosphorylation, serine phosphorylation of the STAT proteins seems to be essential for STATs to trans-activate target genes (Wen et al, 1995;Yamashita et al, 1998).…”