The phosphatidylinositol 3-kinase alpha is required for DNA synthesis induced by some, but not all, growth factors.

Abstract

The phosphatidylinositol 3-kinase (PI 3-K) becomes activated when quiescent cells are stimulated with a variety of growth factors. We have micronijected antibodies specific for the pulla subunit of the PI 3-K into quiescent fibroblasts and tested their effect on the ability of growth factors to stimulate exit from quiescence and entry into S phase. The antibodies inhibited platelet-derived growth factorinduced DNA synthesis, a result in keeping with previous studies using mutant platelet-derived growth factor receptors. Interestingly, functional PI 3-K was required for the first 6 hr of G1-i.e., until -4 hr before the point at which the cells were committed to make DNA. A second tyrosine kinase receptor, the epidermal growth factor receptor, also required the PI 3-K for efficient signaling. However, colony-stimulating factor 1 (whose receptor is highly related to the platelet-derived growth factor receptor) could induce DNA synthesis in the absence of active PI 3-K, as could two growth factors (bombesin and lysophosphatidic acid) whose receptors are functionally coupled to G proteins. These data, therefore, demonstrate that some, but not all, growth factors require functional PI 3-K.The phosphatidylinositol 3-kinase (PI 3-K) associates with several proteins, including growth factor receptors, nonreceptor tyrosine kinases of the Src family, and crk and abl oncogene products (for review, see ref. 1). PI 3-K phosphorylates inositolphospholipids at the D3 position ofthe inositol ring, leading to the formation of phosphatidylinositol 3-phosphate, phosphatidylinositol 3,4-bisphosphate, and phosphatidylinositol 3,4,5-trisphosphate (2), which are poor substrates for any known phospholipase C (3, 4) and may form a new class of second messenger (1). PI 3-K activation and 3-phosphorylated inositolphospholipid formation occur rapidly after growth factor activation (5, 6); thus PI 3-K activation and the formation of its products could be important steps in mitogenic signal transduction. PI 3-K usually consists of a heterodimer of an 85-kDa and a 110-kDa subunit.There are at least two 85-kDa proteins (p85a and p85(), which contain one Src homology region 3 (SH3) domain and two Src homology domain 2 (SH2) domains (for review, see ref. 7). The latter bind with high specificity and affinity to tyrosine phosphorylated sequences (8); indeed, p85a associates with growth factor receptors and the mT-cSrc complex via its SH2 domains (9-12). The catalytic activity resides in the 110-kDa subunit, which has similarity to a Saccharomyces cerevisiae protein, VPS34 (13), that has been implicated in targeting of proteins to the yeast vacuole and vacuole morphogenesis during budding (14), leading to speculation on a possible role of PI 3-K in protein trafficking and vesicular morphogenesis (7). A second catalytic subunit, called p110,8, has recently been described (15); little is known about its biological activity.The binding sites for the SH2 domains of PI 3-K are in the kinase insert region of the platelet-derived growth factor (PD...

PTEN tumour suppressor is linked to the cell cycle control through the retinoblastoma protein

“…Finally, given that PI-3K signalling appears to mediate the functional interrelation between PTEN and Akt, we also analysed the expression of the catalitic subunit of PI-3K (p110) during the cell cycle progression. The results demonstrate no major alterations of p110 expression ( Figure 4a), suggesting that, as expected, the increase in PI-3K activity during cell cycle is regulated by signalling mechanisms independent of PI-3K gene expression (Roche et al, 1994).…”

Signal transduction pathways activated and required for mammary carcinogenesis in response to specific oncogenes

“…Three classes of mammary tumor cell lines, initiated by TGFa, neu, and heregulin/NDF, showed high levels of active PI 3-kinase compared to v-Ha-ras and c-myc induced mammary tumor cell lines. Our results showing that a TGFa initiated cell line is especially sensitive to low concentrations of LY294002 are also noteworthy in light of studies with neutralizing antibodies demonstrating that PI 3-kinase is required for signaling by the EGF receptor (Auger et al, 1989;Jackson et al, 1992;Roche et al, 1994). Furthermore, studies with a dominant negative p85 construct (encoding a mutant version of the regulatory subunit of PI 3-kinase) and chemical inhibitors such as we use here, have showed that PI 3-kinase is required for EGF receptor mediated transformation of mouse epidermal cells in vitro (Huang et al, 1996).…”

Gas6-mediated survival in NIH3T3 cells activates stress signalling cascade and is independent of Ras

“…On the contrary within the same experimental conditions PDGF-mediated survival activity yet required Ras, thus indicating a clear di erence between Gas6 and PDGF-survival inducing pathways. Both have the requirement of (Roche et al, 1994(Roche et al, , 1995 PI3K activity to induce cell survival (Datta et al, 1997;Goruppi et al, 1997) however several reports clearly identi®ed Ras as an upstream e ector required for PDGF-dependent PI3K activation (Rodriguez-Viciana et al, 1994;Datta et al, 1997), while it is likely that Gas6-dependent PI3K activation does not require Ras in this biological system. In fact both wortmannin or p85 dominant negative e ciently block Gas6 survival activities, together with its mitogenic e ects (Goruppi et al, 1997) and Ras was indeed shown to be exclusively required for Gas6-dependent mitogenesis (this manuscript).…”