STAT5a Antibody (Rabbit mAb) [J3A20]

Application: Reactivity: Mouse, Rat, Human

Usage Information

Dilution
WB1:1000 IP1:20 IHC1:1000 IF1:100 - 1:500 FCM1:100

Application
WB, IP, IHC, IF, FCM

Reactivity
Mouse, Rat, Human

Source
Rabbit Monoclonal Antibody

Storage Buffer
PBS, pH 7.2+50% Glycerol+0.05% BSA+0.01% NaN3

Storage (from the date of receipt)
-20°C (avoid freeze-thaw cycles), 2 years

Predicted MW Observed MW
91 kDa 92 kDa
^*Why do the predicted and actual molecular weights differ? The following reasons may explain differences between the predicted and actual protein molecular weight. Post-translational modifications(e.g., phosphorylation, glycosylation); Splice variants and isoforms; Relative charge; Multimerization.

Positive Control	Mouse brain tissue; Rat brain tissue; Human breast tissue; Human squamous lung carcinoma tissue; Recombinant Human STAT5a protein (His tag N-Terminus); Jurkat cells; TF-1 cells; K-562 cells; A431 cells;
Negative Control

Datasheet & SDS

Biological Description

Specificity
STAT5a Antibody (Rabbit mAb) [J3A20] detects endogenous levels of total STAT5a protein.

Clone
J3A20

Synonym(s)
STAT5, STAT5A, Signal transducer and activator of transcription 5A

Background
STAT5A belongs to the STAT family of latent cytoplasmic transcription factors, sharing with its six paralogs, including the closely related STAT5B, a conserved modular architecture built from an N-terminal multimerization domain, a coiled-coil domain, a DNA-binding domain, a linker domain, a single SH2 domain, and a C-terminal transactivation domain. STAT5A activation begins when a cytokine such as growth hormone or prolactin engages its class I cytokine receptor, triggering a receptor-mediated dimerization of two JAK2 molecules through their FERM domains; this JAK2 dimerization requires a membrane-proximal receptor motif termed the switch region, and mutating this switch region disrupts downstream STAT phosphorylation without affecting JAK2 binding to the receptor itself, showing that receptor-driven JAK2 dimer formation, not merely JAK2 recruitment, is the structural step required for kinase activation. Once dimerized and transactivated, JAK2 phosphorylates tyrosine residues on the receptor's cytoplasmic tail, creating docking sites that recruit STAT5A through its SH2 domain; JAK2 then phosphorylates STAT5A itself at a single conserved tyrosine residue, and this phosphorylation drives STAT5A dissociation from the receptor, dimerization with another phosphorylated STAT5 monomer through reciprocal phosphotyrosine-SH2 domain interactions, and translocation to the nucleus, where the STAT5 dimer binds gamma-interferon activation sequence elements within target gene promoters. This dimerization-dependent activation mechanism is directly subverted in BCR-ABL1-positive leukemia: Src-family kinases undergo autophosphorylation at a conserved tyrosine within their own activation loop, generating a docking site that recruits the STAT5A SH2 domain, and this SH2-activation-loop interaction is required for Src-family-kinase-mediated phosphorylation of STAT5A at its activating tyrosine, yet the same interaction simultaneously traps phosphorylated STAT5A in the cytoplasm rather than allowing normal nuclear translocation, producing attenuated target gene expression and reduced colony formation in chronic myeloid leukemia cells despite STAT5A being fully tyrosine-phosphorylated. This finding demonstrates that STAT5A phosphorylation status alone does not predict transcriptional output, since the SH2 domain engaged by Src-family kinases competes directly with the reciprocal STAT5-STAT5 dimerization interaction normally required for nuclear entry. Activated STAT5 drives proliferation and survival of leukemic cells and contributes to both initial transformation and maintenance of BCR-ABL1-driven disease, and because STAT5A additionally mediates hormone-responsive gene programs relevant to prostate and breast cancer through interaction with androgen and estrogen receptor signaling, the JAK2 dimerization interface and the STAT5A SH2 domain represent two mechanistically distinct, independently characterized points for therapeutic intervention in STAT5-driven malignancy.

Background

STAT5A belongs to the STAT family of latent cytoplasmic transcription factors, sharing with its six paralogs, including the closely related STAT5B, a conserved modular architecture built from an N-terminal multimerization domain, a coiled-coil domain, a DNA-binding domain, a linker domain, a single SH2 domain, and a C-terminal transactivation domain. STAT5A activation begins when a cytokine such as growth hormone or prolactin engages its class I cytokine receptor, triggering a receptor-mediated dimerization of two JAK2 molecules through their FERM domains; this JAK2 dimerization requires a membrane-proximal receptor motif termed the switch region, and mutating this switch region disrupts downstream STAT phosphorylation without affecting JAK2 binding to the receptor itself, showing that receptor-driven JAK2 dimer formation, not merely JAK2 recruitment, is the structural step required for kinase activation. Once dimerized and transactivated, JAK2 phosphorylates tyrosine residues on the receptor's cytoplasmic tail, creating docking sites that recruit STAT5A through its SH2 domain; JAK2 then phosphorylates STAT5A itself at a single conserved tyrosine residue, and this phosphorylation drives STAT5A dissociation from the receptor, dimerization with another phosphorylated STAT5 monomer through reciprocal phosphotyrosine-SH2 domain interactions, and translocation to the nucleus, where the STAT5 dimer binds gamma-interferon activation sequence elements within target gene promoters. This dimerization-dependent activation mechanism is directly subverted in BCR-ABL1-positive leukemia: Src-family kinases undergo autophosphorylation at a conserved tyrosine within their own activation loop, generating a docking site that recruits the STAT5A SH2 domain, and this SH2-activation-loop interaction is required for Src-family-kinase-mediated phosphorylation of STAT5A at its activating tyrosine, yet the same interaction simultaneously traps phosphorylated STAT5A in the cytoplasm rather than allowing normal nuclear translocation, producing attenuated target gene expression and reduced colony formation in chronic myeloid leukemia cells despite STAT5A being fully tyrosine-phosphorylated. This finding demonstrates that STAT5A phosphorylation status alone does not predict transcriptional output, since the SH2 domain engaged by Src-family kinases competes directly with the reciprocal STAT5-STAT5 dimerization interaction normally required for nuclear entry. Activated STAT5 drives proliferation and survival of leukemic cells and contributes to both initial transformation and maintenance of BCR-ABL1-driven disease, and because STAT5A additionally mediates hormone-responsive gene programs relevant to prostate and breast cancer through interaction with androgen and estrogen receptor signaling, the JAK2 dimerization interface and the STAT5A SH2 domain represent two mechanistically distinct, independently characterized points for therapeutic intervention in STAT5-driven malignancy.

References
https://pubmed.ncbi.nlm.nih.gov/25885255/ https://pubmed.ncbi.nlm.nih.gov/30044226/

Protein Size (kDa)	PVDF Transfer Membrane Pore Size (μm)
< 10	0.22
10-20	0.22
20-30	0.45
30-45	0.45
45-70	0.45
80-100	0.45
100-140	0.45
> 140	0.45

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Protein Size (kDa)	Separating Gel Percentage
4-40	20%
12-45	15%
10-70	12.5%
15-100	10%
25-100	8%
60-210	5%