The signal transducer and activator of transcription (STAT) proteins were first identified as signaling proteins that function as second messengers and transcription factors in response to cytokines and growth factors (Santos and Costa-Pereira 2011; Li 2008). Mammals have seven STAT genes, namely, STAT1, STAT2, STAT3, STAT4, STAT5A, STAT5B, and STAT6 (Santos and Costa-Pereira 2011). All the STAT proteins are highly conserved and contain six domains including the SH2, linker, coiled -coil, and DNA- binding domain (Bromberg and Darnell 2000). Traditionally, unphosphorylated STAT is believed to reside in the cytoplasm in an inactive form; upon phosphorylation by JAK or another tyrosine kinase, the phosphorylated (active) STAT translocates into the nucleus to induce transcription of target genes (Bromberg and Darnell 2000). However, recent studies have prompted revisions to this paradigm. In addition to the canonical signaling pathway, there is increasing evidence that unphosphorylated STAT shuttles back and forth between the nucleus and cytoplasm (Santos and Costa-Pereira 2011). In this noncanonical mode of signaling, unphospho-STAT seems to carry additional functions, such as chromatin remodeling and heterochromatin dynamics (Li 2008).
STAT’s Protein Structure and Isoforms
The N-terminal domain (NTD) is required for dimerization of unphosphorylated STATs and tetramerization between STAT1, STAT3, and STAT5. In addition, the NTD can also recruit transcriptional co-activators, such as CREB-binding protein (CBP)/p300, by promoting protein-protein interaction, and is also involved in tyrosine phosphorylation and dephosphorylation of STATs. The coiled-coil domain (CCD) is important for the interaction of STATs with other proteins to mediate transcription. For example, STAT3 interacts with c-Jun to induce transcription of target genes in response to IL-6 induction and binds to epidermal growth factor (EGF) receptors via its CCD domain. Besides, a leucine-rich segment is located within the CCD domain, which also regulates nuclear translocation of STAT proteins. Even though the DNA-binding domain (DBD) is highly conserved across the protein family, the small differences in the amino acid sequences of DBD across different STATs are responsible for their different DNA-binding specificities. Upon cytokine stimulation, the heterodimers formed by phosphorylated STAT1 and STAT3 require the DBD for nuclear translocation as well. Next to the DBD is the α-helical linker domain (LD). It links two crucial domains in STAT, namely, DBD and the Src-homology 2 (SH2) domain (Santos and Costa-Pereira 2011). Additionally, LD is also crucial for the transcriptional activation of STAT1 following IFN-γ stimulation and has been implicated in the constitutive nucleocytoplasmic shuttling of unphosphorylated STATs in resting cells (Yang et al. 1999; Marg et al. 2004). The 100-amino-acid-long SH2 domain, located at the C-terminal end of the peptide, is probably the most well-conserved domain in STAT; it plays a crucial role in protein-protein interactions between STATs and other proteins (Santos and Costa-Pereira 2011). SH2 domain recognizes phosphorylated tyrosine residue and binds to phosphorylated Janus kinase (JAK) or STAT (Huang et al. 2008; Watanabe and Arai 1996). Moreover, a number of residues within this domain are important for STAT to mediate cellular functions. In particular, the phosphorylation of STAT1 on tyrosine 701 and its subsequent homodimerization are dependent on the arginine 602 residue within the SH2 domain (Li 2008); when proline 630 is mutated, tyrosine phosphorylation of STAT2 is impaired, and it no longer dimerizes with STAT1 (Santos and Costa-Pereira 2011). The last domain on the C-terminal end is the transactivation domain (TAD). In addition to its function in activating transcription, TAD is used to interact with other proteins as well. For instance, TAD is required for the interaction of STAT1 with breast cancer 1 (BRCA1) and CBP/p300 (Santos and Costa-Pereira 2011).
STAT in the Canonical JAK-STAT Pathway
STAT is involved in regulating many cellular processes such as cell proliferation, apoptosis, growth, hematopoiesis, antiviral responses, and immune systems (O’Shea et al. 2004). Typically, STAT binds to well-defined DNA consensus sequences and initiates transcription of interferon (IFN)-stimulated genes (ISGs). It is also known to regulate the transcription of many genes including the B-cell lymphoma 2 (Bcl-2) family proteins, cyclin D1, Myc, and many genes implicated in angiogenesis or metastasis (Li 2008). In addition to the consensus sequence, STAT tetramer complexes can also recognize and bind weakly to non-consensus regions via the NTD domain for added specificity in its regulation. Besides inducing transcription, STAT can also act as a negative regulator of transcription in some instances (Boucheron et al. 1998). As an example, STAT3β, a naturally occurring splice variant of STAT3, inhibits the transcription of STAT3’s target genes (Caldenhoven et al. 1996).
Unphosphorylated STATs in Gene Regulation
The Role of STATs in Cancer
STAT1, STAT3, and STAT5 are often misregulated in many cancer cells, especially STAT3, which is a known oncogene (Silva 2004). STAT3 is believed to contribute to cancer cell transformation via its inflammatory response (Yu and Jove 2004). In most cases, the abnormal activities of the upstream kinases are the cause of STAT signaling misregulation in cancer cells. In fact, a well-established tumor model in Drosophila involves a hyperactive JAK kinase called hop Tumorous-L (hop Tum-L) that causes STAT over-activation and over-proliferation of blood cells, analogous to human leukemia (Li 2008; Shi et al. 2006). The role of STAT activation in these pathological conditions has so far been attributed mostly to over-activation of the canonical JAK-STAT pathway that upregulates STAT target genes. In addition to the canonical pathway, disruption of STAT’s function in stabilizing heterochromatin may also contribute to tumorigenesis since heterochromatin has been implicated in tumor suppression. Unphospho-STAT5A has been shown to be prominently present in the nucleus of cultured human cells and plays a role in heterochromatin stability and tumor suppression (Hu et al. 2013). Therefore, respective contributions of canonical and noncanonical JAK-STAT signaling in cancer development are currently under investigation.
In conclusion, STAT is a crucial signaling molecule in animal cells that translates extracellular signals into changes in gene transcription. In the canonical JAK-STAT pathway, only phosphorylated STATs translocate into the nucleus and act as transcription factors, whereas unphosphorylated STATs are assumed to remain latent in the cytoplasm. In the noncanonical mode of JAK-STAT signaling, unphosphorylated STATs are also found, at least in fruit flies, to localize in the nucleus and play important roles in regulating gene expression by modulating heterochromatin formation. It has been shown that mammalian JAK2 plays a role in regulating heterochromatin formation (Dawson et al. 2009). And STAT’s function in heterochromatin seems conserved in mammals (Hu et al. 2013). Proper function of STAT is essential for different stages of development, and maintaining the health of individual organisms and misregulation of STAT has been implicated in multiple human diseases. STAT thus poses as an attractive potential therapeutic target, because it mediates a rather straightforward signaling pathway, and the phospho-tyrosine residue can conceivably be targeted. However, since STAT has proven to be versatile and involved in multiple functions via distinct mechanisms, targeting STAT may cause unexpected cellular responses. Therefore, the biological functions and cellular effects of STAT proteins must be thoroughly investigated even though STAT remains a potential therapeutic target.
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