Phospho-NTRK1 (Y496) Antibody
Description
Introduction to Phospho-NTRK1 (Y496) Antibody
Phospho-NTRK1 (Y496) Antibody is a specialized immunological reagent designed to specifically detect the neurotrophic tyrosine kinase receptor type 1 (NTRK1), commonly known as TrkA, only when phosphorylated at tyrosine residue 496. This site-specific antibody serves as a critical tool for researchers investigating neuronal development, oncogenic signaling, and various neurological disorders. The antibody's high specificity allows for precise monitoring of TrkA activation status, providing valuable insights into nerve growth factor (NGF) signaling pathways and their dysregulation in disease states .
These antibodies are available in various formats, including polyclonal and recombinant monoclonal versions, each optimized for specific applications such as Western blotting, immunofluorescence, and enzyme-linked immunosorbent assays (ELISA). The development of these specialized reagents has significantly advanced our understanding of neurotrophin signaling mechanisms and their implications in both normal physiological conditions and pathological states .
Scientific Background of NTRK1 and Y496 Phosphorylation
NTRK1 encodes the high-affinity nerve growth factor receptor TrkA, a transmembrane receptor tyrosine kinase primarily expressed in neural tissues. This receptor plays crucial roles in the development and maintenance of the central and peripheral nervous systems through regulation of neuronal proliferation, differentiation, and survival .
The TrkA receptor becomes activated through a specific sequence of molecular events. Upon binding its primary ligand, nerve growth factor (NGF), or alternatively neurotrophin-3 (NTF3), TrkA undergoes homodimerization followed by autophosphorylation at multiple tyrosine residues, including Y496 . This phosphorylation event represents a critical step in the initiation of downstream signaling cascades that ultimately regulate neuronal function and survival.
Tyrosine 496 phosphorylation specifically serves as a docking site for adaptor proteins that mediate signal transduction. The phosphorylated Y496 residue in TrkA corresponds functionally to phosphorylated Y516 in TrkB and TrkC, highlighting the evolutionary conservation of this signaling mechanism across the Trk family of receptors .
Research Applications
These antibodies find application in diverse experimental techniques:
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Western Blotting: Used at dilutions ranging from 1:500 to 1:10,000 to detect phosphorylated TrkA in cell or tissue lysates, typically visualized as a 140 kDa band .
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Immunofluorescence: Applied at dilutions of 1:200 to 1:1,000 to visualize the subcellular localization of activated TrkA in fixed cells or tissue sections .
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ELISA: Employed at higher dilutions (approximately 1:5,000) in sandwich ELISA formats to quantify phosphorylated TrkA levels .
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Proximity Ligation Assay: Used in modified form to detect functional signaling complexes formed by phosphorylated TrkA with adaptor proteins such as SHC1, providing spatial information about active signaling in situ .
Molecular Mechanisms and Signaling Pathways
The phosphorylation of Y496 in TrkA represents a critical event in neurotrophin signaling, serving as a direct binding site for key adaptor proteins. This molecular interaction initiates multiple signaling cascades essential for neuronal function.
Y496 Phosphorylation and Downstream Signaling
When phosphorylated, Y496 directly binds and activates SHC-transforming protein (SHC) and fibroblast growth factor receptor substrate 2 (FRS2) . These interactions initiate signal transduction through several major pathways:
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Ras-MAPK Pathway: Through SHC1 and FRS2, phosphorylated TrkA activates a GRB2-Ras-MAPK cascade regulating cell differentiation and survival .
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PI3K-AKT Pathway: Phosphorylated Y496, via SHC1 and SH2B1, controls a Ras-PI3 kinase-AKT1 signaling cascade critical for neuronal survival .
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PLCγ Pathway: Although PLCγ primarily binds to phosphorylated Y791 of TrkA, signaling initiated at Y496 can influence this pathway, which controls NF-κB activation and transcription of genes involved in cell survival .
These pathways ultimately regulate the transcription of genes involved in neuronal differentiation and survival, including those encoding cyclic AMP-responsive element-binding (CREB) transcriptional co-activator proteins .
Research Applications and Clinical Significance
Phospho-NTRK1 (Y496) antibodies have proven instrumental in advancing our understanding of both physiological processes and pathological conditions involving neurotrophin signaling.
Neurological Research
In neurological research, these antibodies enable scientists to monitor TrkA activation in response to neurotrophic factors, providing insights into mechanisms of neuronal development, survival, and plasticity. This has facilitated studies on neurodegenerative diseases, pain disorders, and neurodevelopmental conditions associated with aberrant neurotrophin signaling .
Cancer Research
TrkA signaling has emerged as a significant focus in oncology, particularly with the identification of oncogenic NTRK fusions in various cancer types. Phospho-NTRK1 (Y496) antibodies have been utilized to characterize signaling in these contexts, contributing to the development and assessment of TRK inhibitors as targeted therapies .
In a notable application, researchers employed a proximity ligation assay based on TrkA-Y496 phosphorylation to detect active TrkA signaling in a patient with soft-tissue sarcoma harboring an oncogenic LMNA-NTRK1 gene fusion. This assay demonstrated robust signaling associated with tumor nuclei, consistent with oncogenic activity of the fusion protein .
Functional Analysis of NTRK1 Mutations
Phospho-NTRK1 (Y496) antibodies have been instrumental in the functional characterization of NTRK1 mutations associated with hereditary sensory and autonomic neuropathy type IV (HSAN IV). By measuring Y496 phosphorylation levels, researchers have demonstrated that complete abolition of TrkA kinase activity is not the only pathogenic mechanism underlying this condition, revealing a diverse range of functional effects across different mutations .
Comparative Analysis of Available Antibodies
Several manufacturers offer Phospho-NTRK1 (Y496) antibodies with varying specifications and optimized applications. The following table compares key products:
| Manufacturer | Catalog Number | Type | Key Features | Applications | Cross-Reactivity |
|---|---|---|---|---|---|
| St John's Labs | STJ91325 | Polyclonal | Detects endogenous levels of TrkA phosphorylated at Y496 | WB, IF, ELISA | Human, Mouse, Rat |
| Proteintech | 84976-1-PBS | Recombinant | PBS-only formulation | WB, Indirect ELISA | Human |
| Abcam | ab197071 | Recombinant Monoclonal | Cross-reacts with TrkB (Y516) and TrkC (Y516) | Dot Blot, WB | Human, Rat |
These products offer researchers options tailored to specific experimental needs, with variations in specificity, cross-reactivity, and optimized applications .
Product Specs
Target Background
- Two novel compound heterozygous variants of NTRK1 (c.632T > A and c.1253_1254delTC) were identified in a pair of Chinese identical twins with Congenital Insensitivity to Pain and Anhidrosis. PMID: 30461622
- The above results suggest that rutin preconditioning ameliorates cerebral I/R injury in OVX rats through ER-mediated BDNF-TrkB and NGF-TrkA signaling. PMID: 29420916
- The TrkA peptide is competitive for metal binding with analogous peptides due to the N-terminal domain of NGF. These data provide cues for future exploration of the effect of metal ions on the activity of the NGF and its specific cellular receptor. PMID: 30103559
- The LMNA-NTRK1 fusion was likely the molecular driver of tumorigenesis and metastasis in this patient, and the observed effectiveness of crizotinib treatment provides clinical validation of this molecular target. PMID: 30134855
- that lipofibromatosis-like tumour represents a novel entity of NTRK1-associated neoplasms PMID: 29958731
- System xC(-)-mediated TrkA activation therefore presents a promising target for therapeutic intervention in cancer pain treatment. PMID: 29761734
- Results identified two known splice-site mutations, one known nonsense mutation and one novel missense mutation in three congenital insensitivity to pain with anhidrosis (CIPA) pedigrees. These findings expanded the spectrum of the NTRK1 mutations associated with CIPA patients, providing additional clues for the phenotype-genotype relationship beneath CIPA. PMID: 30201336
- 27 mutations in NTRK1 from Congenital insensitivity to pain with anhidrosis cohort, including 15 novel mutations, are reported. PMID: 29770739
- NTRK1 was upregulated in 80% of head and neck squamous carcinoma tissue. PMID: 29904026
- TRKA expression can be found in 1.6% of solid tumours and can be paralleled by NTRK1 gene rearrangements or mostly copy number gain PMID: 29802225
- These results suggest that polymorphisms in NTRK1 play an important role in pain sensitivity in young Han Chinese women PMID: 29054434
- We developed a comprehensive model of acquired resistance to NTRK inhibitors in cancer with NTRK1 rearrangement and identified cabozantinib as a therapeutic strategy to overcome the resistance PMID: 28751539
- TrkA plays an important role in the pathogenesis of NPM-ALK(+) T-cell lymphoma. PMID: 28557340
- Results show frequent BRCA2, EGFR, and NTRK1/2/3 mutations in mismatch repair-deficient colorectal cancers , sugggesting personalized medicine strategies to treat the patients with advanced disease who may have no remaining treatment options PMID: 28591715
- novel deletional mutation has enriched the spectrum of NTRK1 mutations PMID: 28981924
- This study identify four novel NTRK1 mutations (IVS14+3A>T, p.Ser235*, p.Asp596Asn, and p.Leu784Serfs*79) and demonstrate that they are pathologic mutations using an mRNA splicing assay and an NTRK autophosphorylation assay. PMID: 28177573
- Report a novel mechanism for the TRAIL-induced apoptosis of TrkAIII expressing NB cells that depends upon SHP/Src-mediated crosstalk between the TRAIL-receptor signaling pathway and TrkAIII. PMID: 27821809
- This show evidence of variation in plasmatic monocytic TrkA expression during the progression of dementia. PMID: 27802234
- TrkA was detected in 20% of thyroid cancers, compared with none of the benign samples. TrkA expression was independent of histologic subtypes but associated with lymph node metastasis, suggesting the involvement of TrkA in tumor invasiveness. Nerves in the tumor microenvironment were positive for TrkA. PMID: 29037860
- phenotypes, as well as both recurrent and novel mutations in NTRK1 in 2 Chinese patients with CIPA PMID: 28192073
- we conclude that complete abolition of TRKA kinase activity is not the only pathogenic mechanism underlying HSAN IV. PMID: 27676246
- Nine patients have been reported from nine unrelated families with hereditary sensory and autonomic neuropathy IV due to various mutations in NTRK1, five of which are novel. PMID: 28328124
- Data suggest that kinase domains of neurotrophin receptor isoforms, TRKA, TRKB, and TRKC, exhibit a bulky phenylalanine gatekeeper, leading to a small and unattractive back pocket/binding site for antineoplastic kinase inhibitors. [REVIEW] PMID: 28215291
- Pan-Trk immunohistochemistry is a time-efficient and tissue-efficient screen for NTRK fusions, particularly in driver-negative advanced malignancies and potential cases of secretory carcinoma and congenital fibrosarcoma. PMID: 28719467
- analysis of NTRK1 transcripts in peripheral blood cells of the patient revealed an influence of the variant on mRNA splicing. The C>A transversion generated a novel splice-site, which led to the incorporation of 10 intronic bases into the NTRK1 mRNA and consequently to a non-functional gene product. PMID: 27184211
- NTRK fusions occur in a subset of young patients with mesenchymal or sarcoma-like tumors at a low frequency PMID: 28097808
- A novel nonsense mutation and a known splice-site mutation were detected in NTRK1 in two siblings and were shown to be associated with congenital insensitivity to pain with anhidrosis. PMID: 28345382
- NTRK1 gene fusion in spitzoid neoplasms results in tumors with Kamino bodies and were typically arranged in smaller nests with smaller predominantly spindle-shaped cells, occasionally forming rosettes. PMID: 27776007
- Results suggest that NTRK1 oncogenic activation through gene fusion defines a novel and distinct subset of soft tissue tumors resembling lipofibromatosis (LPF), but displaying cytologic atypia and a neural immunophenotype, provisionally named LPF-like neural tumors. PMID: 27259011
- This review highlights treatment options, including clinical trials for ROS1 rearrangement, RET fusions, NTRK1 fusions, MET exon skipping, BRAF mutations, and KRAS mutations. PMID: 27912827
- ShcD binds to active Ret, TrkA, and TrkB neurotrophic factor receptors predominantly via its phosphotyrosine-binding (PTB) domain. PMID: 28213521
- TrkA misfolding and aggregation induced by some Insensitivity to Pain with Anhidrosis mutations disrupt the autophagy homeostasis causing neurodegeneration. PMID: 27551041
- USP36 actions extend beyond TrkA because the presence of USP36 interferes with Nedd4-2-dependent Kv7.2/3 channel regulation. PMID: 27445338
- Our results demonstrated that TrkA expression was associated with tumor progression and poor survival, and was an independent predictor of poor outcomes in gastric cancer patients PMID: 26459250
- High NTRK1 expression is associated with colon cancer. PMID: 26716414
- TrkA immunohistochemistry is an effective, initial screening method for NTRK1 rearrangement detection in the clinic. PMID: 26472021
- This work identifies GGA3 as a key player in a novel DXXLL-mediated endosomal sorting machinery that targets TrkA to the plasma membrane, where it prolongs the activation of Akt signaling and survival responses. PMID: 26446845
- Data show that p.G595R and p.G667C TRKA mutations drive acquired resistance to entrectinib in colorectal cancers carrying NTRK1 rearrangements. PMID: 26546295
- Two key biological processes for progressive hearing loss, TrkA signaling pathway and EGF receptor signaling pathway were significantly and differentially enriched by the two sets of allele-specific target genes of miR-96. PMID: 26564979
- Report novel variant of myo/haemangiopericytic sarcoma with recurrent NTRK1 gene fusions. PMID: 26863915
- TrkA as a candidate oncogene in malignant melanoma and support a model in which the NGF-TrkA-MAPK pathway may mediate a trade-off between neoplastic transformation and adaptive anti-proliferative response. PMID: 26496938
- IL-13 confers epithelial cell responsiveness to NGF by regulating NTRK1 levels by a transcriptional and epigenetic mechanism and that this process likely contributes to allergic inflammation. PMID: 25389033
- findings suggest that Cbl-b limits NGF-TrkA signaling to control the length of neurites. PMID: 25921289
- mRNA expression of NTRK1 genes was higher in low-grade gliomas vs. high-grade and control samples. Poor survival was associated with NTRK1 mRNA. Promoter methylation does not regulate NTRK1 genes in glioma. PMID: 24840578
- Translocations in the NTRK1 gene are recurring events in colorectal cancer, although occurring at a low frequency (around 0.5%). PMID: 26001971
- Findings have implications for understanding the mature and less malignant neuroblastoma phenotype associated with NTRK1 expression, and could assist the development of new therapeutic strategies for neuroblastoma differentiation PMID: 25361003
- TrkA expression in neurons was found to be regulated at the gene promoter level by Bex3 protein. PMID: 25948268
- Causative role for M379I and R577G NTRK1 mutations in melanoma development is highly unlikely. PMID: 24965840
- Increased NTRK1 expression is associated with spontaneous abortions. PMID: 24825909
- Data indicate how the neurotrophins function through tyrosine kinase receptors TrkC and TrkA. PMID: 24603864
