Met pS


Fig.10.2 The domain structure of STAT proteins. STATs contain several conserved domains. The activity of STATs may be regulated by several posttranslational modifications, including tyrosine phosphorylation (pY), serine phosphorylation (pS), acetylation (Ace), sumoylation (Sumo), and methylation (Met). Contradictory reports on the roles of sumoylation and methylation in the regulation of STAT activity have been reported. See text for details.

of STATs in unstimulated cells also existed as dimers (Braunstein et al, 2003; Ota et al, 2004; Yuan et al, 2005; Zhong et al, 2005), although the exact amount of non-phosphorylated STATs existing as dimers under physiological conditions is not known.

The crystal structures of tyrosine phosphorylated STAT1 or STAT3 dimers binding to DNA have been determined (Becker et al., 1998; Chen et al., 1998). The reciprocal and highly specific SH2-phosphotyrosyl peptide interactions are solely responsible for holding two STAT monomers together and may be involved in stabilizing the STAT-DNA binding (Chen et al., 1998).

B: DNA Binding Domain

The first evidence that suggests the direct binding of a STAT protein to DNA comes from studies on IFN-y signaling. Using two-dimensional mobility gel shift and SDS-PAGE analysis and UV cross-linking assays, STAT1 was found to directly bind to DNA (Shuai et al., 1992).

Sequence analysis of STAT proteins failed to reveal significant homology to any known DNA binding domains. The DNA binding domain of STATs was identified by domain-swapping studies (Horvath et al., 1995). It was shown that the region between residues 400 to 500 of STAT might contact DNA. The DNA binding domain has been further defined from the crystal structures of STATs binding to DNA (Becker et al., 1998; Chen et al., 1998). The region between residues 317 to 488 of STAT1 is in contact with DNA and this region contains an immunoglobulin-like fold, which resembles the DNA binding domains of the p50 subunit of NF-kB and the tumor suppressor p53.

All STAT proteins, except STAT2, can bind to DNA alone. The consensus DNA binding sequence for STAT1, 3, 4, 5 is TTN5AA (where N represents any nucleotide). The optimal DNA binding sequence for STAT6 is TTN6AA (Darnell, 1997). However, it should be noted that the natural DNA binding sequences present in the promoters of endogenous genes have different affinity toward STAT binding, which may provide additional specificity in transcriptional regulation.

C: Transcriptional Activation Domain

STAT 1(3, a differentially spliced product of Statl gene lacking the COOH-terminal 38 amino acids present in STAT la, failed to activate IFN-y-induced genes, although it was tyrosine phosphorylated, translocated to the nucleus, and bound to DNA (Muller et al., 1993; Shuai et al., 1993a). These findings suggest that the COOH-terminal 38 amino acid residues may act as the transcriptional activation domain of STATl. Indeed, when fused to the GAL4 DNA-binding domain, the COOH-terminal regions of STATs can act as transcriptional activation domains (Bhattacharya et al., 1996).

The COOH-terminal regions of STATs have been shown to interact with several transcriptional coactivators, including the histone acetyltransferases p300/CREB-binding protein (CBP) (Bhattacharya et al., 1996; Zhang et al., 1996) and the acetyltransferase general control non-repressed 5 (GCN5) (Paulson et al., 2002), resulting in the enhanced transcriptional activity of STATl and STAT2.

D: The NH2-TerminalRegion

The NH2-terminal region of STATs contains a N-terminai domain and a coil-coil domain (Fig. 10.2). Tandem arrays of multiple copies of weak STAT-binding sites are present in some genes such as MIG (monokine induced by IFN-y) and IFN-y (Xu et al., 1996). The binding of STAT dimers to two adjacent DNA sites is cooperative and involves the formation of a tetrameric STAT-DNA binding complex. The NH2-terminal regions of STATl and STAT4, although not required for the binding of a dimer to a single high-affinity binding site, were required for a cooperative interaction between two STAT dimers (Vinkemeier et al, 1996; Xu et al, 1996).

Recently, a portion of STATs in untreated cells was found to exist as dimers (Braunstein et al, 2003; Ota et al, 2004; Yuan et al, 2005; Zhong et al., 2005). The crystal structure of unphosphorylated STATl has been resolved (Yuan et al, 2005). The NH2-terminal region of STATl was shown to mediate the dimerization of unphosphorylated STATl molecules. However, the NH2-region appears to play different roles in the activation of STATs. For example, mutations in the NH2-terminal region of STAT4 abolished the tyrosine phosphorylation of STAT4 (Ota et al, 2004). In contrast, the mutation of several conserved residues at the NH2-terminal region of STATl increased the tyrosine phosphorylation of STATl, which probably resulted from the defects in the tyrosine dephophorylation of STATl caused by these mutations (Zhong et al, 2005). Thus, the NH2-terminal regions of different STATs may play different roles in the regulation of STAT activity.

Regulation of STATs

The activity of STATs is tightly regulated by multiple mechanisms, including posttranslational modifications of STATs, inhibition of STAT activity by PIAS proteins, and the dephosphoiylation of STATs by protein tyrosine phosphatases (Shuai and Liu, 2003).

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