Upon activation by tyrosine kinases, members of the STAT family of transcription factors form stable dimers that are able to rapidly translocate to the nucleus and bind DNA. Although crystal structures of activated, near full-length, Stat1 and Stat3 illustrate how STATs bind to DNA, they provide little insight into the dynamic regulation of STAT activity. To explore the unique structural changes Stat1 and Stat3 undergo when they become activated, full-length inactive recombinant proteins were prepared. To our surprise, even though these proteins are unable to bind DNA, our studies demonstrate that they exist as stable homodimers. Similarly, the Stat1 and Stat3 found in the cytoplasm of unstimulated cells also exhibit a dimeric structure. These observations indicate that Stat1 and Stat3 exist as stable homodimers prior to activation.Cytokines are important regulators of intercellular communication. They mediate pleiotropic cellular responses by binding to specific transmembrane spanning receptors (reviewed in Refs. 1 and 2). These receptors in turn activate intracellular signaling pathways, resulting in the induction of gene expression. The STAT 1 (signal transducers and activators of transcription) family of transcription factors transmits signals in response to cytokines. This family consists of seven members, Stat1, Stat2, Stat3, Stat4, Stat5a, Stat5b, and Stat6. Structural and functional studies have led to the identification of six conserved STAT domains (3-5): 1) the amino-terminal domain (ϳamino acids 1-125), which has been implicated in cooperativity of DNA binding to tandem gamma activation site (GAS) elements, nuclear translocation, and in receptor association (2); 2) the coiled-coil domain (ϳamino acids 125-325), which has been shown to mediate interactions with several other proteins (2); 3) the DNA binding domain (ϳamino acids 325-475); 4) the linker domain (ϳamino acids 475-575); 5) the SH2 domain ϩ tyrosine activation motif (ϳamino acids 575-710), which is essential for STAT recruitment to the receptor, STAT activation, and dimerization (2); and 6) the carboxyl-terminal transcriptional activation domain.The general outline for the STAT signaling paradigm elucidated a decade ago is now widely accepted (1, 2, 6). In this model, a specific interaction between a cytokine and its cognate receptor brings the cytoplasmic domains of this receptor into apposition, thereby promoting the transphosphorylation of receptor associated tyrosine kinases from the Janus kinase family. These activated Janus kinases in turn phosphorylate specific tyrosine motifs found in receptor endodomains, which then serve to mediate recruitment of the specific monomeric STATs to the receptor complex. Once at the receptor, the STATs become activated by a single tyrosine phosphorylation event. The activated STATs are then released, where upon they dimerize through the interaction between the SH2 domain of one STAT and the tyrosine-phosphorylated tail segment of the other STAT. This renders them competent for both rapid translocation ...

No abstract

Immature myeloid cells have been shown to transduce signals through a carboxyl-terminally truncated isoform of Stat5. This functionally distinct signal transducer and activator of transcription isoform is generated through a unique protein-processing event. Evaluation of numerous cell lines has determined that there is a direct correlation between the expression of truncated Stat5 and protease activity. Moreover, protease activity is found only in the myeloid and not in lymphoid progenitors. To further characterize the protease small quantities have been purified to near homogeneity. Studies on this purified material indicate that the protease has an apparent molecular mass of ϳ25 kDa and is active over a wide range of pH values. The protease will also cleave both activated (i.e. tyrosine-phosphorylated) and inactivate Stat5. Although this activity is sensitive to phenylmethylsulfonyl fluoride, it is notably not sensitive to several other serine protease inhibitors. Additional studies have led to the identification of the unique site where the protease cleaves Stat5. Mutagenesis of this site renders Stat5 resistant to cleavage. Consistent with the model that Stat5 cleavage is important for early myeloid development, introduction of a "non-cleavable" isoform of Stat5 into FDC-P1 cells (a myeloid progenitor line) leads to significant phenotypic changes.

Characterization of the ability of interferons to induce immediate early genes led to the identification of the signal transducer and activators of transcription (STAT) signaling paradigm, where a single protein transduces signals directly from the receptor to the nucleus. Subsequent studies have determined that all cytokines transduce pivotal signals through at least one of the seven members of this STAT family. Notably, cytokines can be placed in functional subgroups based on the STATs they activate.