The janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway plays a critical role in the signaling of a wide array of cytokines and growth factors leading to various cellular functions, including proliferation, growth, hematopoiesis, and immune response.1-2

The binding of cytokines and growth factors to their corresponding receptors activates JAK, which then phosphorylates the receptor and STAT proteins on specific tyrosine residues. STATs then dimerize, translocate to the nucleus, bind to the consensus DNA sequence of 5’-TT(N4–6)AA-3’ and initiate the transcription of target genes.1-2

Life Technologies offers antibodies, ELISAs, Luminex® multiplex assays and growth factors for key targets in the JAK signaling cascade.

 

 

     JAK-STAT Pathway

Find JAK-STAT signaling pathway primary antibodies, antibody pairs, ELISAs and Luminex® assays by protein

Intracellular Proteins   Membrane Receptors   Cytokine Proteins

Key JAK-STAT Pathway Targets

Four JAK family kinases, including JAK1, JAK2, JAK3, and TYK2, and seven STAT family members, including STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b, and STAT6, have been identified. JAK1, JAK2, and TYK2 appear to be ubiquitously expressed, while JAK3 expression is normally limited to lymphoid cells. The JAKs are structurally unique in having a C-terminal kinase domain (JH1) preceded by a pseudokinase domain (JH2), which lacks the catalytic activity but has a critical regulatory function. JAKs also have a Src homology 2 (SH2) domain and an N-terminal band four-point-one, ezrin, radixin, moesin (FERM) domain that is critical for mediating the association with cytokine receptors. STAT proteins contain a SH2 domain for dimerization and a DNA-binding domain. The amino acid sequence diversity and their tissue-specific distributions account for the diverse roles of STATs in response to extracellular cytokines.1-2 The JAK-STAT pathways are up-regulated by a vast array of cytokines/growth factors. One mechanism for negative regulation of JAK-STAT pathways is through suppresser of cytokine signaling (SOCS) proteins, which directly bind to and inactivate JAKs3, and protein inhibitors of activated STATs (PIAS) that bind to phosphorylated STAT dimers, preventing DNA binding.4

Abnormal constitutive activation of JAK-STAT pathways has been implicated in various cancers and immune disorders. For example, STAT3 is persistently activated in many tumors, including major carcinomas and some hematologic tumors.5 Activating mutations in JAK2 have been linked to leukemia. TEL-JAK2 fusion due to chromosomal translocation was identified in a small set of human T cell acute lymphoblast leukemia patients.6 The V617F mutation in the JH2 pseudo-kinase domain of JAK2 was found in a high percentage of patients with myeloproliferative disorders, including polycythaemia vera.7 Inhibitors of JAK-STAT pathways are currently being sought in the areas of oncology and immune disorders.

Recombinant Antibody

Recombinant Rabbit Monoclonal Antibody
(click to enlarge)

Flow cytometry of Jurkat cells labeled with rabbit anti-STAT4.
Jurkat cells were fixed and permeabilized using FIX & PERM® reagents.

Cells were stained with (black trace) or without (blue trace) 0.5 μg anti-STAT4 followed by Alexa Fluor® 488 goat anti-rabbit Ig. Pre-incubation with immunogenic peptide decreased the signal (red trace).  Data were generated using a Novex® STAT4 ABfinity™ Recombinant Rabbit Monoclonal Antibody     (700185).

Recombinant Protein

Recombinant Protein

Expression of STAT5b (pY699) in TF-1 cells.   
TF-1 cells were treated with IFN-α or IL-3, or left unstimulated.

Cell lysates were analyzed using the STAT5b [pY699] ELISA kit. Treatment with IFN-α or IL-3 results in activation of the JAK/STAT pathway, as seen by the upregulation of phosphorylated STAT5b. Data were generated using a Novex® IFN-α Pure Recombinant Protein (PHC4014).

ELISA Quantification

ELISA Quantification

Total ELISA kits make effective controls.
Cell extracts were prepared and analyzed with the STAT5a [pY694] ELISA and STAT5a (Total) ELISA kits. 

Phosphorylation of STAT5a is increased in sodium vanadate-treated HEL cells, whereas the total level of STAT5a remains relatively constant in treated vs. untreated control, demonstrating the utility of the Total ELISA kits as controls.  Data were generated using a Novex®
STAT5a [pY694] ELISA Kit (KHO0761) and    
STAT5a (Total) ELISA Kit (KHO0751).

Events leading to STAT activation.

Events leading to STAT activation
(click to enlarge)
     Events leading to STAT activation. (A) Extracellular IFN (blue triangle) prior to binding to its receptor. (B) Upon binding, JAK kinases constitutively associated with the receptor subunits interact. The interacting JAK proteins activate one another by reciprocal tyrosine phosphorylation, and phosphorylate a tyrosine on two subunits contained in the receptor complex. These phosphorylated tyrosine residues provide paired docking sites for STAT via its SH2 domain (C). STAT, recruited to the receptor complex, is then phosphorylated at a tyrosine residue by the JAKs. This tyrosine phosphorylation promotes STAT homo and heterodimerization mediated by reciprocal phosphotyrosine SH2 domain interaction and the dissociation of the dimers from the receptor complex (D). The STAT dimer is then translocated to the cell nucleus, where they complex with other nuclear proteins and regulate gene expression by binding to promotors or other response elements on DNA.

Membrane Receptors

Gene Symbol
ALKF2R (Par1)KDR (CD309)
AXL (UFO)F2RL2KIT (CD117)
BTKFGFR1 (CD331)MERTK
CD2FGFR2 (CD332)MET
CD4FGFR3 (CD333)MST1R (CD136)
CD80FGFR4 (CD334)MUSK
CSF1R (CD115)FLT1NGFR (CD27)1
CXCR4 (CD184)FLT3 (CD135)NTRK1
DDR2FLT4NTRK2
EGFRIFNAR1NTRK3
EPHA1IFNAR2 (CD118)PDGFRA (CD140a)
EPHA2IFNGR1 (CD119)PDGFRB (CD140b)
EPHA3IGF1R (CD221)PRLR
EPHA4IL1R1 (CD121a)ROR2
EPHA5IL2RA (CD25)TEK
EPHA7IL2RG (CD132)TNFRSF10A (CD120a)
EPHA8IL3RA (CD123)TNFRSF10B (CD262)
EPHB1IL4R (CD124)TNFRSF1A (CD120a)
EPHB2IL6R (CD126)TNFRSF1B (CD120b)
EPHB3IL12BRTNFRSF25 (DR3)
EPHB4IL13RA (CD213A1)TYRO3
ERBB2 (Her2)INSR (CD220) 
ERBB4 (Her4)INSRR 

Cytokine Proteins

Gene Symbol
ANGPT1FGF1IL5
ANGPT2FGF2IL6
ANGPT4FGF3IL7
BMP2FGF4IL8
BMP7FGF5IL9
CCL2 (MCP1)FGF6IL10
CCL3 (MIP-1α)FGF7IL11
CCL4 (MIP-1β)FGF8IL12A
CCL5 (RANTES)FGF9IL12B
CCL7 (MCP-3)FGF10IL13
CCL8 (MCP-2)FGF16IL15
CCL11 (Eotaxin)FGF17IL16
CCL13 (MCP4)FGF19IL17A
CCL15 (MIP-5)GH1IL18
CCL17 (TARC)IFNA1IL19
CCL18 (MIP-4)IFNA2IL20
CCL19 (MIP-3β)IFNB1IL23A
CCL20 (MIP-3α)IFNGPRL
CCL21 (6Ckine)IL1ATNFSF10 (TRAIL)
CCL25 (TECK)IL1BTNFSF13B (BAFF)
CXCL12 (SDF1)IL2VEGFA
EGFIL3VEGFC
EPOIL4 

References

  1. Aaronson, D.S. et al. (2002) Science 296: 1653-1655. 
  2. O’Shea, J.J. et al. (2004) Nat Rev Drug Discovery 3: 555-564.
  3. Kishimoto, T. et al. (2001) Nat Genetics 28: 4-5.
  4. Shuai, K. et al. (2000) Oncogene 19 : 2638-2645.
  5. Darnell, J.E. et al. (2005) Nat Medicine 11: 595-596.
  6. Lacronique, V. et al. (1997) Science 278: 1309-1312.
  7. Ferrajoli, A. et al. (2006) Curr Cancer Drug Targets 6: 671-679.

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