Ras GTPase‐activating protein‐associated p62 is a major v‐Src‐SH3‐binding protein

Williger, Ben-Tsion; Liscovitch, Mordechai

doi:10.1016/s0014-5793(97)00027-6

Cited by 3 publications

(2 citation statements)

References 45 publications

(9 reference statements)

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“…Sam68 has been found to bind SH3 domains of different molecules in vitro: Src, p85, PLCγ, and Grb2 [Williger and Liscovitch, 1997; Shen et al, 1999]. Nevertheless, although the SH3‐mediated interactions with p85 and PLC‐γ has been confirmed [Richard et al, 1995; Taylor et al, 1995], no evidence of in vivo interaction of Sam68 with Grb2 in cells has been provided.…”

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confidence: 99%

Sam68 associates with the SH3 domains of Grb2 recruiting GAP to the Grb2‐SOS complex in insulin receptor signaling

Najib

Sánchez-Margalet

2002

J of Cellular Biochemistry

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The 68 kDa Src substrate associated during mitosis (Sam68) is an RNA binding protein with Src homology (SH) 2 and 3 domain binding sites. We have recently found that Sam68 is a substrate of the insulin receptor (IR) that translocates from the nucleus to the cytoplasm and that Tyr-phosphorylated Sam68 associates with the SH2 domains of p85 PI3K and GAP, in vivo and in vitro. In the present work, we have further demonstrated the cytoplasmic localization of Sam68, which is increased in cells overexpressing IR. Besides, we sought to further study the association of Sam68 with the Ras-GAP pathway by assessing the interactions with SH3 domains of Grb2. We employed GST-fusion proteins containing the SH3 domains of Grb2 (N or C), and recombinant Sam68 for in vitro studies. In vivo studies of protein-protein interaction were assessed by co-immunoprecipitation experiments with specific antibodies against Sam68, GAP, Grb2, SOS, and phosphotyrosine; and by affinity precipitation with the fusion proteins (SH3-Grb2). Insulin stimulation of HTC-IR cells promotes phosphorylation of Sam68 and its association with the SH2 domains of GAP. Sam68 is constitutively associated with the SH3 domains of Grb2 and it does not change upon insulin stimulation, but Sam68 is Tyr-phosphorylated and promotes the association of GAP with the Grb2-SOS complex. In vitro studies with fusion proteins showed that Sam68 association with Grb2 is preferentially mediated by the C-terminal SH3 domains of Grb2. In conclusion, Sam68 is a substrate of the IR and may have a role as a docking protein in IR signaling, recruiting GAP to the Grb2-SOS complex, and in this way it may modulate Ras activity.

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confidence: 99%

Sam68 associates with the SH3 domains of Grb2 recruiting GAP to the Grb2‐SOS complex in insulin receptor signaling

Najib

Sánchez-Margalet

2002

J of Cellular Biochemistry

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“…To date, seven members of the DOK family of proteins, DOK‐1–7, have been identified and shown to serve as substrates of various protein tyrosine kinases, including ABL, SRC, BCR, TIE2, KIT, platelet‐derived growth factor receptor (PDGFR), epidermal growth factor receptor (EGFR), muscle‐specific tyrosine kinase (MuSK), and RET. ( 18,19,22–24,27–30,35–40 ) DOK‐1–3 are preferentially expressed in hematopoietic tissues and act as negative regulators of RAS/ERK signaling. On the other hand, DOK‐4–6 are mainly expressed in non‐hematopoietic tissues such as the nervous system and embryonic kidneys, and act as positive or negative regulators of RAS/ERK signaling.…”

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confidence: 99%

Analysis of DOK‐6 function in downstream signaling of RET in human neuroblastoma cells

Kurotsuchi

Murakumo²,

Jijiwa³

et al. 2010

Cancer Science

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Point mutations and structural alterations of the RET tyrosine kinase gene cause multiple endocrine neoplasia type 2 (MEN 2) and papillary thyroid carcinoma, respectively. RET activation by glial cell line-derived neurotrophic factor (GDNF) is essential for the development of the enteric nervous system and the kidney. The signal through RET tyrosine kinase requires several adaptor proteins including the DOK (downstream of kinase) family of proteins. Of the seven members of the DOK protein family, DOK-1, -4, -5, and -6 have been reported to play roles in the GDNF-RET signaling pathway. Although DOK-6 has been shown to bind to RET and promote GDNF-induced neurite outgrowth in mouse Neuro2A cells, DOK-6 function in human cells remains unclear. In the present study, we investigated the role of DOK-6 in GDNF-RET signaling in human cells including neuroblastoma cells. DOK-6 was constitutively localized to the plasma membrane via its pleckstrin homology (PH) domain, and was phosphorylated following RET activation via a MEN2A mutation or GDNF stimulation. However, DOK-6 could not significantly affect downstream signaling and neurite outgrowth in human neuroblastoma cells. The binding affinity of the DOK-6 phosphotyrosine-binding (PTB) domain to RET was much lower than that of the DOK-1, DOK-4, and SHC PTB domains to RET. These findings indicate that DOK-6 is involved in RET signaling with less influence when compared with DOK-1, DOK-4, and SHC. (Cancer Sci 2010; 101: 1147-1155 T he RET proto-oncogene encodes receptor tyrosine kinase (RTK) expressed in a wide variety of neurons and the developing kidney.(1-4) RET tyrosine kinase is activated by stimulation of the glial cell line-derived neurotrophic factor (GDNF) family of ligands including GDNF, neurturin, artemin, and persephin in the presence of glycosylphosphatidylinositolanchored co-receptors called GDNF family receptor alpha 1-4.(5) Consistent with the expression pattern of RET during embryogenesis, signaling through RET plays crucial roles in the development of the enteric nervous system and the kidney. RET mutations are responsible for the development of several human diseases including multiple endocrine neoplasia (MEN) type 2A and 2B, familial medullary thyroid carcinoma (FMTC), papillary thyroid carcinoma (PTC), and Hirschsprung's disease.(8-10) RET mutations in MEN2A, MEN2B, FMTC, and PTC are gain-of-function mutations, whereas Hirschsprung mutations are loss-of-function mutations.(11-17) Specific tyrosine residues in the RET intracellular domain are phosphorylated by GDNF stimulation or RET mutations, which results in the activation of several signaling pathways, including the RAS ⁄ ERK (extracellular signal-regulated kinase), PI3K (phosphatidylinositol-3 kinase) ⁄ AKT, p38MAPK (mitogen activated protein kinase), phospholipase Cc, and RAC ⁄ JNK (c-Jun N-terminal kinase) pathways.(8-10) These signaling pathways are important for cell proliferation, survival, migration, and invasion.Signal transduction through RET requires various adaptor proteins such as SHC,...

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A Chemical Genetic Screen for Direct v-Src Substrates Reveals Ordered Assembly of a Retrograde Signaling Pathway

Shah

Shokat

2002

Chemistry & Biology

130

118

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Using an ATP analog that is a specific substrate for an analog-specific allele of v-Src, we identified several novel cytoskeletal substrates that control actin assembly processes. A screen for less abundant v-Src substrates revealed the scaffolding protein Dok-1 as a direct substrate of v-Src. Further studies suggest that v-Src phosphorylation sites on Dok-1 are critical for its binding to RasGAP and Csk, negative regulators of Src signaling. This results in the downregulation of growth-promoting signals of the Src family kinases and the Ras pathway. Identification of the direct substrates of v-Src leads to a model for the precise order of assembly of a retrograde signaling pathway in v-Src-transformed cells and has provided new insight into the balance between those signals that promote cell transformation mediated by v-Src catalyzed tyrosine phosphorylation and those that inhibit it.

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Ras GTPase‐activating protein‐associated p62 is a major v‐Src‐SH3‐binding protein

Cited by 3 publications

References 45 publications

Sam68 associates with the SH3 domains of Grb2 recruiting GAP to the Grb2‐SOS complex in insulin receptor signaling

Sam68 associates with the SH3 domains of Grb2 recruiting GAP to the Grb2‐SOS complex in insulin receptor signaling

Analysis of DOK‐6 function in downstream signaling of RET in human neuroblastoma cells

A Chemical Genetic Screen for Direct v-Src Substrates Reveals Ordered Assembly of a Retrograde Signaling Pathway

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