Phosphorylation Regulates Tau Interactions with Src Homology 3 Domains of Phosphatidylinositol 3-Kinase, Phospholipase Cγ1, Grb2, and Src Family Kinases

Phosphatidylinositol 3-kinase (PI 3-kinase) has been implicated as a participant in signaling pathways regulating cell growth by virtue of its activation in response to various mitogenic stimuli. Here we describe the cloning of a novel and ubiquitously expressed human PI 3-kinase. The 4.8-kb cDNA encodes a putative translation product of 1,070 amino acids which is 42% identical to bovine PI 3-kinase and 28% identical to Vps34, a Saccharomyces cerevisiae PI 3-kinase involved in vacuolar protein sorting. Human PI 3-kinase is also similar to Tor2, a yeast protein required for cell cycle progression. Northern (RNA) analysis demonstrated expression of human PI 3-kinase in all tissues and cell lines tested. Protein synthesized from an epitope-tagged cDNA had intrinsic PI 3-kinase activity and associated with the adaptor 85-kDa subunit of PI 3-kinase (p85) in intact cells, as did endogenous human PI 3-kinase. Coprecipitation assays showed that a 187-amino-acid domain between the two src homology 2 domains of p85 mediates interaction with PI 3-kinase in vitro and in intact cells. These results demonstrate the existence of different PI 3-kinase isoforms and define a family of genes encoding distinct PI 3-kinase catalytic subunits that can associate with p85.

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Using a Phage Display Library to Identify Basic Residues in A-Raf Required to Mediate Binding to the Src Homology 2 Domains of the p85 Subunit of Phosphatidylinositol 3′-Kinase

Journal of Biological Chemistry ◽

10.1074/jbc.m004720200 ◽

2000 ◽

Vol 275 (46) ◽

pp. 36450-36456 ◽

Cited By ~ 22

Author(s):

Tamela R. King ◽

Yun Fang ◽

Elizabeth S. Mahon ◽

Deborah H. Anderson

Keyword(s):

Phage Display ◽

Phage Display Library ◽

Phosphatidylinositol 3 Kinase ◽

Src Homology 2 ◽

Src Homology ◽

Phosphatidylinositol 3

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Genetic analysis of a phosphatidylinositol 3-kinase SH2 domain reveals determinants of specificity.

Molecular and Cellular Biology ◽

10.1128/mcb.14.9.5929 ◽

1994 ◽

Vol 14 (9) ◽

pp. 5929-5938 ◽

Cited By ~ 16

Author(s):

M Yoakim ◽

W Hou ◽

Z Songyang ◽

Y Liu ◽

L Cantley ◽

...

Keyword(s):

Phosphorylation Site ◽

Growth Factor Receptor ◽

Sh2 Domain ◽

T Antigen ◽

Site Directed Mutagenesis ◽

Phosphatidylinositol 3 Kinase ◽

Sh2 Domains ◽

Src Homology ◽

Polyomavirus Middle T Antigen ◽

Phosphatidylinositol 3

Phosphatidylinositol 3-kinase is an important element in both normal and oncogenic signal transduction. Polyomavirus middle T antigen transforms cells in a manner depending on association of its tyrosine 315 phosphorylation site with Src homology 2 (SH2) domains on the p85 subunit of the phosphatidylinositol 3-kinase. Both nonselective and site-directed mutagenesis have been used to probe the interaction of middle T with the N-terminal SH2 domain of p85. Most of the 24 mutants obtained showed reduced middle T binding. However, mutations that showed increased binding were also found. Comparison of middle T binding to that of the platelet-derived growth factor receptor showed that some mutations altered the specificity of recognition by the SH2 domain. Mutations altering S-393, D-394, and P-395 were shown to affect the ability of the SH2 domain to select peptides from a degenerate phosphopeptide library. These results focus attention on the role of the EF loop in the SH2 domain in determining binding selectivity at the third position after the phosphotyrosine.

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Effect of nutritional state on the formation of a complex involving insulin receptor IRS-1, the 52 kDa Src homology/collagen protein (Shc) isoform and phosphatidylinositol 3′-kinase activity

Biochemical Journal ◽

10.1042/bj3350293 ◽

1998 ◽

Vol 335 (2) ◽

pp. 293-300 ◽

Cited By ~ 23

Author(s):

Joëlle DUPONT ◽

Michel DEROUET ◽

Jean SIMON ◽

Mohammed TAOUIS

Keyword(s):

Insulin Receptor ◽

Kinase Activity ◽

Residual Activity ◽

Nutritional State ◽

Phosphatidylinositol 3 Kinase ◽

Regulatory Subunits ◽

Normal Tissues ◽

Collagen Protein ◽

Src Homology ◽

Phosphatidylinositol 3

The Src homology and collagen protein (Shc) is tyrosine phosphorylated in response to insulin; however, evidence for its interaction with insulin receptor (IR) in normal tissues is missing. Interactions between IR, Shc and regulatory subunits of the phosphatidylinositol 3´-kinase (PI 3´-kinase) were characterized in the present study in liver and muscles of chickens submitted to various nutritional states. A chicken liver Shc cDNA fragment encoding a 198 amino acid long fragment, including the phosphotyrosine binding domain was sequenced. It shows 89% homology with the corresponding human homologue. The amounts of the three Shc isoforms (66, 52 and 46 kDa) and Shc messenger were not altered by the nutritional state. Shc tyrosine phosphorylation was decreased by fasting in both liver and muscle. Importantly, Shc was immunoprecipitated by IR antibody (mostly the 52 kDa isoform) or by αIRS-1(mostly the 46 kDa isoform). IR–Shc association was decreased by fasting and restored by refeeding. In liver, αShc immunoprecipitated the three forms of regulatory subunits of PI 3´-kinase and a PI 3´-kinase activity which was decreased by fasting. In muscle, αShc immunoprecipitated only the p85 isoform; the associated PI 3´-kinase activity was not altered by the nutritional state. Conversely, in both tissues anti-p85 antibody precipitated only the 52 kDa Shc isoform. In liver, antibodies to insulin receptor substrate-1 (αIRS-1), Shc or IR immunoprecipitated the three regulatory subunits of PI 3´-kinase and an equal PI 3´-kinase activity, without any residual activity left in the supernatants, suggesting the presence of a large complex involving IR, IRS-1, Shc (mainly the 52 kDa isoform) and PI 3´-kinase activity. The presence of another complex containing IRS-1 and the 46 kDa Shc isoform, but no PI 3´-kinase activity, is suggested.

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