TheJAK-STAT signaling pathway is a chain of interactions between proteins in a cell, and is involved in processes such asimmunity,cell division,cell death, andtumor formation. The pathway communicates information from chemical signals outside of a cell to thecell nucleus, resulting in the activation of genes through the process oftranscription. There are three key parts of JAK-STAT signalling:Janus kinases (JAKs),signal transducer and activator of transcription proteins (STATs), and receptors (which bind the chemical signals).^[1] Disrupted JAK-STAT signalling may lead to a variety of diseases, such as skin conditions,cancers, and disorders affecting the immune system.^[1]

Main articles:JAKs andSTATs

There are four JAK proteins:JAK1,JAK2,JAK3 andTYK2.^[1] JAKs contains aFERM domain (approximately 400 residues), an SH2-related domain (approximately 100 residues), akinase domain (approximately 250 residues) and apseudokinase domain (approximately 300 residues).^[2] The kinase domain is vital for JAK activity, since it allows JAKs tophosphorylate (add phosphate groups to) proteins.

There are seven STAT proteins:STAT1,STAT2,STAT3,STAT4,STAT5A,STAT5B andSTAT6.^[1] STAT proteins contain many different domains, each with a different function, of which the most conserved region is theSH2 domain.^[2] The SH2 domain is formed of 2α-helices and aβ-sheet and is formed approximately from residues 575–680.^[2][3] STATs also havetranscriptional activation domains (TAD), which are less conserved and are located at theC-terminus.^[4] In addition, STATs also contain: tyrosine activation, amino-terminal, linker,coiled-coil andDNA-binding domains.^[4]

The binding of variousligands, usually cytokines, such asinterferons andinterleukins, to cell-surface receptors, causes the receptors to dimerize, which brings the receptor-associated JAKs into close proximity.^[6] The JAKs then phosphorylate each other ontyrosine residues located in regions calledactivation loops, through a process calledtransphosphorylation, which increases the activity of their kinase domains.^[6] The activated JAKs then phosphorylate tyrosine residues on the receptor, creating binding sites for proteins possessingSH2 domains.^[6] STATs then bind to the phosphorylated tyrosines on the receptor using their SH2 domains, and then they are tyrosine-phosphorylated by JAKs, causing the STATs to dissociate from the receptor.^[2] At least STAT5 requiresglycosylation atthreonine 92 for strong STAT5 tyrosine phosphorylation.^[7] These activated STATs formhetero- orhomodimers, where the SH2 domain of each STAT binds the phosphorylated tyrosine of the opposite STAT, and the dimer then translocates to thecell nucleus to induce transcription of target genes.^[2] STATs may also be tyrosine-phosphorylated directly byreceptor tyrosine kinases - but since most receptors lack built-in kinase activity, JAKs are usually required for signalling.^[1]

To move from thecytosol to thenucleus, STAT dimers have to pass throughnuclear pore complexes (NPCs), which are protein complexes present along thenuclear envelope that control the flow of substances in and out of the nucleus. To enable STATs to move into the nucleus, an amino acid sequence on STATs, called thenuclear localization signal (NLS), is bound by proteins calledimportins.^[4] Once the STAT dimer (bound to importins) enters the nucleus, a protein calledRan (associated with GTP) binds to the importins, releasing them from the STAT dimer.^[8] The STAT dimer is then free in the nucleus.

Specific STATs appear to bind to specific importin proteins. For example,STAT3 proteins can enter the nucleus by binding to importin α3 and importin α6.^[9] On the other hand,STAT1 andSTAT2 bind to importin α5.^[4] Studies indicate that STAT2 requires a protein calledinterferon regulatory factor 9 (IRF9) to enter the nucleus.^[8] Not as much is known about nuclear entrance of other STATs, but it has been suggested that a sequence of amino acids in the DNA-binding domain ofSTAT4 might allow nuclear import; also,STAT5 andSTAT6 can both bind to importin α3.^[8] In addition, STAT3, STAT5 and STAT6 can enter the nucleus even if they are not phosphorylated at tyrosine residues.^[8]

After STATs are made byprotein biosynthesis, they have non-protein molecules attached to them, calledpost-translational modifications. One example of this is tyrosine phosphorylation (which is fundamental for JAK-STAT signalling), but STATs experience other modifications, which may affect STAT behaviour in JAK-STAT signalling. These modifications include:methylation,acetylation andserine phosphorylation.

• Methylation. STAT3 can be dimethylated (have two methyl groups) on alysine residue, at position 140, and it is suggested that this could reduce STAT3 activity.^[10] There is debate as to whether STAT1 is methylated on anarginine residue (at position 31), and what the function of this methylation could be.^[11]
• Acetylation. STAT1, STAT2, STAT3, STAT5 and STAT6 have been shown to be acetylated.^[12] STAT1 may have an acetyl group attached to lysines at positions 410 and 413, and as a result, STAT1 can promote the transcription of apoptotic genes - triggering cell death.^[12] STAT2 acetylation is important for interactions with other STATs, and for the transcription of anti-viral genes.^[4]

Acetylation of STAT3 has been suggested to be important for its dimerization, DNA-binding and gene-transcribing ability, andIL-6 JAK-STAT pathways that use STAT3 require acetylation for transcription of IL-6 response genes.^[12] STAT5 acetylation on lysines at positions 694 and 701 is important for effective STAT dimerization inprolactin signalling.^[13] Adding acetyl groups to STAT6 is suggested to be essential for gene transcription in some forms ofIL-4 signalling, but not all the amino acids which are acetylated on STAT6 are known.^[12]

• Serine phosphorylation. Most of the seven STATs (except STAT2) undergo serine phosphorylation.^[2] Serine phosphorylation of STATs has been shown to reduce gene transcription.^[14] It is also required for the transcription of some target genes of the cytokines IL-6 andIFN- γ.^[11] It has been proposed that phosphorylation of serine can regulate STAT1 dimerization,^[11] and that continuous serine phosphorylation on STAT3 influences cell division.^[15]

Like many other transcription factors, STATs are capable of recruitingco-activators such asCBP andp300, and these co-activators increase the rate of transcription of target genes.^[2] The coactivators are able to do this by making genes on DNA more accessible to STATs and by recruiting proteins needed for transcription of genes. The interaction between STATs and coactivators occurs through the transactivation domains (TADs) of STATs.^[2] The TADs on STATs can also interact withhistone acetyltransferases (HATs);^[16] these HATs add acetyl groups to lysine residues on proteins associated with DNA calledhistones. Adding acetyl groups removes the positive charge on lysine residues, and as a result there are weaker interactions between histones and DNA, making DNA more accessible to STATs and enabling an increase in the transcription of target genes.

JAK-STAT signalling is able to interconnect with other cell-signalling pathways, such as thePI3K/AKT/mTOR pathway.^[17] When JAKs are activated and phosphorylate tyrosine residues on receptors, proteins with SH2 domains (such as STATs) are able bind to the phosphotyrosines, and the proteins can carry out their function. Like STATs, thePI3K protein also has an SH2 domain, and therefore it is also able to bind to these phosphorylated receptors.^[17] As a result, activating the JAK-STAT pathway can also activate PI3K/AKT/mTOR signalling.

JAK-STAT signalling can also integrate with theMAPK/ERK pathway. Firstly, a protein important for MAPK/ERK signalling, calledGrb2, has an SH2 domain, and therefore it can bind to receptors phosphorylated by JAKs (in a similar way to PI3K).^[17] Grb2 then functions to allow the MAPK/ERK pathway to progress. Secondly, a protein activated by the MAPK/ERK pathway, calledMAPK (mitogen-activated protein kinase), can phosphorylate STATs, which can increase gene transcription by STATs.^[17] However, although MAPK can increase transcription induced by STATs, one study indicates that phosphorylation of STAT3 by MAPK can reduce STAT3 activity.^[18]

One example of JAK-STAT signalling integrating with other pathways isInterleukin-2 (IL-2) receptor signaling inT cells. IL-2 receptors have γ (gamma) chains, which are associated withJAK3, which then phosphorylates key tyrosines on the tail of the receptor.^[19] Phosphorylation then recruits an adaptor protein calledShc, which activates the MAPK/ERK pathway, and this facilitates gene regulation bySTAT5.^[19]

An alternative mechanism for JAK-STAT signalling has also been suggested. In this model,SH2 domain-containingkinases, can bind to phosphorylated tyrosines on receptors and directly phosphorylate STATs, resulting in STAT dimerization.^[6] Therefore, unlike the traditional mechanism, STATs can be phosphorylated not just by JAKs, but by other receptor-bound kinases. So, if one of the kinases (either JAK or the alternative SH2-containing kinase) cannot function, signalling may still occur through activity of the other kinase.^[6] This has been shown experimentally.^[20]

Given that many JAKs are associated withcytokine receptors, the JAK-STAT signalling pathway plays a major role in cytokine receptor signalling. Sincecytokines are substances produced by immune cells that can alter the activity of neighbouring cells, the effects of JAK-STAT signalling are often more highly seen in cells of the immune system. For example,JAK3 activation in response toIL-2 is vital forlymphocyte development and function.^[21] Also, one study indicates thatJAK1 is needed to carry out signalling for receptors of the cytokines IFNγ, IL-2, IL-4 andIL-10.^[22]

The JAK-STAT pathway in cytokine receptor signalling can activate STATs, which can bind to DNA and allow the transcription of genes involved in immune cell division, survival, activation and recruitment. For example,STAT1 can enable the transcription of genes which inhibit cell division and stimulateinflammation.^[2] Also,STAT4 is able to activateNK cells (natural killer cells), andSTAT5 can drive theformation of white blood cells.^[2][23] In response to cytokines, such as IL-4, JAK-STAT signalling is also able to stimulateSTAT6, which can promoteB-cell proliferation, immune cell survival, and the production of an antibody calledIgE.^[2]

JAK-STAT signalling plays an important role in animal development. The pathway can promote blood cell division, as well asdifferentiation (the process of a cell becoming more specialised).^[24] In some flies with faulty JAK genes, too much blood cell division can occur, potentially resulting inleukaemia.^[25] JAK-STAT signalling has also been associated with excessivewhite blood cell division in humans and mice.^[24]

The signalling pathway is also crucial for eye development in the fruit fly (Drosophila melanogaster). When mutations occur in genes coding for JAKs, some cells in the eye may be unable to divide, and other cells, such asphotoreceptor cells, have been shown not to develop correctly.^[24]

The entire removal of a JAK and a STAT inDrosophila causes death ofDrosophila embryos, whilst mutations in the genes coding for JAKs and STATs can cause deformities in the body patterns of flies, particularly defects in forming body segments.^[24] One theory as to how interfering with JAK-STAT signalling might cause these defects is that STATs may directly bind to DNA and promote the transcription of genes involved in forming body segments, and therefore by mutating JAKs or STATs, flies experience segmentation defects.^[26] STAT binding sites have been identified on one of these genes, calledeven-skipped (eve), to support this theory.^[27] Of all the segment stripes affected by JAK or STAT mutations, the fifth stripe is affected the most, the exact molecular reasons behind this are still unknown.^[24]

Given the importance of the JAK-STAT signalling pathway, particularly in cytokine signalling, there are a variety of mechanisms that cells possess to regulate the amount of signalling that occurs. Three major groups of proteins that cells use to regulate this signalling pathway areprotein inhibitors of activated STAT (PIAS),^[28]protein tyrosine phosphatases (PTPs)^[29] andsuppressors of cytokine signalling (SOCS).^[30] Computational models of JAK-STAT signaling based on the laws ofchemical kinetics have elucidated the importance of these different regulatory mechanisms on JAK-STAT signaling dynamics.^[31][32][33]

PIAS are a four-member protein family made of:PIAS1,PIAS3,PIASx, andPIASγ.^[34] The proteins add a marker, calledSUMO (small ubiquitin-like modifier), onto other proteins – such as JAKs and STATs, modifying their function.^[34] The addition of a SUMO group ontoSTAT1 by PIAS1 has been shown to prevent activation of genes by STAT1.^[35] Other studies have demonstrated that adding a SUMO group to STATs may block phosphorylation of tyrosines on STATs, preventing their dimerization and inhibiting JAK-STAT signalling.^[36] PIASγ has also been shown to prevent STAT1 from functioning.^[37] PIAS proteins may also function by preventing STATs from binding to DNA (and therefore preventing gene activation), and by recruiting proteins calledhistone deacetylases (HDACs), which lower the level of gene expression.^[34]

Since adding phosphate groups on tyrosines is such an important part of how the JAK-STAT signalling pathway functions, removing these phosphate groups can inhibit signalling. PTPs are tyrosine phosphatases, so are able to remove these phosphates and prevent signalling. Three major PTPs areSHP-1,SHP-2 andCD45.^[38]

• SHP-1. SHP-1 is mainly expressed inblood cells.^[39] It contains two SH2 domains and a catalytic domain (the region of a protein that carries out the main function of the protein) - the catalytic domain contains the amino acid sequence VHCSAGIGRTG (a sequence typical of PTPs).^[40] As with all PTPs, a number of amino acid structures are essential for their function: conservedcysteine,arginine andglutamine amino acids, and a loop made oftryptophan,proline andaspartate amino acids (WPD loop).^[40] When SHP-1 is inactive, the SH2 domains interact with the catalytic domain, and so the phosphatase is unable to function.^[40] When SHP-1 is activated however, the SH2 domains move away from the catalytic domain, exposing the catalytic site and therefore allowing phosphatase activity.^[40] SHP-1 is then able to bind and remove phosphate groups from the JAKs associated with receptors, preventing the transphosphorylation needed for the signalling pathway to progress.

One example of this is seen in the JAK-STAT signalling pathway mediated by theerythropoietin receptor (EpoR). Here, SHP-1 binds directly to a tyrosine residue (at position 429) on EpoR and removes phosphate groups from the receptor-associated JAK2.^[41] The ability of SHP-1 to negatively regulate the JAK-STAT pathway has also been seen in experiments using mice lacking SHP-1.^[42] These mice experience characteristics ofautoimmune diseases and show high levels of cell proliferation, which are typical characteristics of an abnormally high level of JAK-STAT signalling.^[42] Additionally, addingmethyl groups to the SHP-1 gene (which reduces the amount of SHP-1 produced) has been linked tolymphoma (a type of blood cancer) .^[43]

However, SHP-1 may also promote JAK-STAT signalling. A study in 1997 found that SHP-1 potentially allows higher amounts of STAT activation, as opposed to reducing STAT activity.^[44] A detailed molecular understanding for how SHP-1 can both activate and inhibit the signalling pathway is still unknown.^[38]

• SHP-2. SHP-2 has a very similar structure to SHP-1, but unlike SHP-1, SHP-2 is produced in many different cell types - not just blood cells.^[45] Humans have two SHP-2 proteins, each made up of 593 and 597 amino acids.^[40] The SH2 domains of SHP-2 appear to play an important role in controlling the activity of SHP-2. One of the SH2 domains binds to the catalytic domain of SHP-2, to prevent SHP-2 functioning.^[38] Then, when a protein with a phosphorylated tyrosine binds, the SH2 domain changes orientation and SHP-2 is activated.^[38] SHP-2 is then able to remove phosphate groups from JAKs, STATs and the receptors themselves - so, like SHP-1, can prevent the phosphorylation needed for the pathway to continue, and therefore inhibit JAK-STAT signalling. Like SHP-1, SHP-2 is able to remove these phosphate groups through the action of the conserved cysteine, arginine, glutamine and WPD loop.^[40]

Negative regulation by SHP-2 has been reported in a number of experiments - one example has been when exploringJAK1/STAT1 signalling, where SHP-2 is able to remove phosphate groups from proteins in the pathway, such as STAT1.^[46] In a similar manner, SHP-2 has also been shown to reduce signalling involvingSTAT3 andSTAT5 proteins, by removing phosphate groups.^[47][48]

Like SHP-1, SHP-2 is also believed to promote JAK-STAT signalling in some instances, as well as inhibit signalling. For example, one study indicates that SHP-2 may promote STAT5 activity instead of reducing it.^[49] Also, other studies propose that SHP-2 may increaseJAK2 activity, and promote JAK2/STAT5 signalling.^[50] It is still unknown how SHP2 can both inhibit and promote JAK-STAT signalling in the JAK2/STAT5 pathway; one theory is that SHP-2 may promote activation of JAK2, but inhibit STAT5 by removing phosphate groups from it.^[38]

• CD45. CD45 is mainly produced in blood cells.^[4] In humans it has been shown to be able to act on JAK1 and JAK3,^[51] whereas in mice, CD45 is capable of acting on all JAKs.^[52] One study indicates that CD45 can reduce the amount of time that JAK-STAT signalling is active.^[52] The exact details of how CD45 functions is still unknown.^[38]

There are eight protein members of theSOCS family:cytokine-inducible SH2 domain-containing protein (CISH),SOCS1,SOCS2,SOCS3,SOCS4,SOCS5,SOCS6, andSOCS7, each protein has anSH2 domain and a 40-amino-acid region called the SOCS box.^[53] The SOCS box can interact with a number of proteins to form a protein complex, and this complex can then cause the breakdown of JAKs and the receptors themselves, therefore inhibiting JAK-STAT signalling.^[4] The protein complex does this by allowing a marker called ubiquitin to be added to proteins, in a process calledubiquitination, which signals for a protein to be broken down.^[54] The proteins, such as JAKs and the receptors, are then transported to a compartment in the cell called theproteasome, which carries out protein breakdown.^[54]

SOCS can also function by binding to proteins involved in JAK-STAT signalling and blocking their activity. For example, the SH2 domain of SOCS1 binds to a tyrosine in the activation loop of JAKs, which prevents JAKs from phosphorylating each other.^[4] The SH2 domains of SOCS2, SOCS3 and CIS bind directly to receptors themselves.^[54] Also, SOCS1 and SOCS3 can prevent JAK-STAT signalling by binding to JAKs, using segments called kinase inhibitory regions (KIRs) and stopping JAKs binding to other proteins.^[55] The exact details of how other SOCS function is less understood.^[4]

Since the JAK-STAT pathway plays a major role in many fundamental processes, such asapoptosis andinflammation, dysfunctional proteins in the pathway may lead to a number of diseases. For example, alterations in JAK-STAT signalling can result incancer and diseases affecting the immune system, such assevere combined immunodeficiency disorder (SCID).^[56]

JAK3 can be used for the signalling ofIL-2,IL-4,IL-15 andIL-21 (as well as other cytokines); therefore patients with mutations in the JAK3 gene often experience issues affecting many aspects of the immune system.^[57][58] For example, non-functional JAK3 causes SCID, which results in patients having noNK cells,B cells orT cells, and this would make SCID individuals susceptible to infection.^[58] Mutations of theSTAT5 protein, which can signal with JAK3, has been shown to result inautoimmune disorders.^[59]

It has been suggested that patients with mutations inSTAT1 andSTAT2 are often more likely to develop infections from bacteria and viruses.^[60] Also,STAT4 mutations have been associated withrheumatoid arthritis, andSTAT6 mutations are linked toasthma.^[61][62]

Patients with a faulty JAK-STAT signalling pathway may also experience skin disorders. For example, non-functional cytokine receptors, and overexpression ofSTAT3 have both been associated withpsoriasis (an autoimmune disease associated with red, flaky skin).^[58] STAT3 plays an important role in psoriasis, as STAT3 can control the production ofIL-23 receptors, and IL-23 can help the development ofTh17 cells, and Th17 cells can induce psoriasis.^[63] Also, since many cytokines function through the STAT3 transcription factor, STAT3 plays a significant role in maintainingskin immunity.^[58] In addition, because patients with JAK3 gene mutations have no functional T cells, B cells or NK cells, they would more likely to develop skin infections.

Cancer involves abnormal and uncontrollable cell growth in a part of the body. Therefore, since JAK-STAT signalling can allow the transcription of genes involved in cell division, one potential effect of excessive JAK-STAT signalling is cancer formation. High levels of STAT activation have been associated with cancer; in particular, high amounts of STAT3 and STAT5 activation is mostly linked to more dangerous tumours.^[64] For example, too much STAT3 activity has been associated with increasing the likelihood ofmelanoma (skin cancer) returning after treatment and abnormally high levels of STAT5 activity have been linked to a greater probability of patient death fromprostate cancer.^[65][64] Altered JAK-STAT signalling can also be involved in developingbreast cancer. JAK-STAT signalling inmammary glands (located within breasts) can promote cell division and reduce cell apoptosis during pregnancy and puberty, and therefore if excessively activated, cancer can form.^[66] High STAT3 activity plays a major role in this process, as it can allow the transcription of genes such asBCL2 andc-Myc, which are involved in cell division.^[66]

Mutations inJAK2 can lead toleukaemia andlymphoma.^[6] Specifically, mutations inexons 12, 13, 14 and 15 of the JAK2 gene are proposed to be a risk factor in developing lymphoma or leukemia.^[6] Additionally, mutated STAT3 and STAT5 can increase JAK-STAT signalling in NK and T cells, which promotes very high proliferation of these cells, and increases the likelihood of developing leukaemia.^[66] Also, a JAK-STAT signalling pathway mediated byerythropoietin (EPO), which usually allows the development of red blood cells, may be altered in patients with leukemia.^[67]

The Janus kinase (JAK)/signal transducer and the activator of the transcription (STAT) pathway were at the centre of attention for driving hyperinflammation inCOVID-19, i.e., theSARS-CoV-2infection triggers hyperinflammation through the JAK/STAT pathway, resulting in the recruitment ofdendritic cells,macrophages, andnatural killer (NK) cells, as well asdifferentiation ofB cells andT cells progressing towardscytokine storm.^[68]

Since excessive JAK-STAT signalling is responsible for some cancers and immune disorders,JAK inhibitors have been proposed as drugs for therapy. For instance, to treat some forms of leukaemia, targeting and inhibiting JAKs could eliminate the effects of EPO signalling and perhaps prevent the development of leukaemia.^[67] One example of a JAK inhibitor drug isruxolitinib, which is used as a JAK2 inhibitor.^[64] STAT inhibitors are also being developed, and many of the inhibitors target STAT3.^[66] It has been reported that therapies which target STAT3 can improve the survival of patients with cancer.^[66] Another drug, calledTofacitinib, has been used for psoriasis and rheumatoid arthritis treatment, and has been approved for treatment ofCrohn's disease andulcerative colitis.^[56]

• Janus kinase inhibitor, a type of Janus kinases-blocking drugs used for cancer therapy.
• Signal transducing adaptor protein, a helper protein used by major proteins in signalling pathways.

• JAK-STAT, peer-reviewed journal published by Landes Bioscience
• Jak/Stat pathway (human) on wikipathwaysArchived 6 February 2018 at theWayback Machine
• Web Site of Austrian Special Research Program (SFB) on Jak STAT signaling

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• Gene expression
• Transcription factors

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