Mitogen-induced tyrosine phosphorylation of 41 kDa and 43 kDa proteins. Potential role in integrating multiple mitogenic signalling pathways

Abstract

We have examined the possible involvement of pertussis toxin (PT)-sensitive GTP-binding protein and protein kinase C (PKC) in mitogen-induced tyrosine phosphorylation of the 41 kDa and 43 kDa cytosol proteins using PT-pretreated (inactivation of PT-sensitive GTP-binding protein) or phorbol 12-myristate 13-acetate (PMA)-pretreated (depletion of PKC) mouse fibroblasts. The effects of the inactivation of PT-sensitive GTP-binding protein and the depletion of PKC on mitogen-stimulated tyrosine phosphorylation of the proteins were similar and varied significantly and systematically in response to growth factors. The important finding was that such inhibitory effects of PT-sensitive GTP-binding protein inactivation and PKC depletion on protein tyrosine phosphorylation induced by each mitogen always correlated well with their inhibitory effects on each mitogen-stimulated DNA synthesis. Although the extent of platelet-derived-growth-factor-induced phosphorylation of the proteins was decreased to approx. 50% in PT- and PMA-pretreated cells compared with native cells, protein phosphorylation itself was not affected and occurred at identical sites on each protein in native, PT- and PMA-pretreated cells. These results suggest that: (1) 41 kDa and 43 kDa proteins are located downstream of PT-sensitive GTP-binding protein and PKC in the mitogenic signalling pathways of growth factors, (2) protein phosphorylation occurs via a cascade of events which includes the activation of the receptor tyrosine kinases, PKC and other unidentified kinase(s) which directly participate(s) in the phosphorylation of the 41 kDa and 43 kDa proteins, and (3) their phosphorylation may play an important role in integrating multiple mitogenic signalling pathways.

Pertussis toxin induces angiogenesis in brain microvascular endothelial cells

“…We followed up by immunoblotting analysis with 23 phospho‐site‐specific antibodies that revealed the most marked changes in phosphorylation as a consequence of PTX intoxication. In previous studies, PTX has been demonstrated to elicit reductions of ERK1 and ERK2 phosphorylation in response to mitogens in mouse fibroblasts (Kohno et al, 1992), in response to cannabinoids in rabbit vascular smooth muscle cells (Su and Vo, 2007), by sphingosine‐1‐phosphatase receptor agonists such as Fingolimod in rat brain cortical astrocytes (Osinde et al, 2007), and by angiotensin II in rat pulmonary artery EC (which appears to be mediated via PTX inhibition of Gα i3 stimulation of Src and Raf1; Li et al, 2007). In the present study, in MBEC treated with PTX alone, there was clear evidence for inhibition of ERK1/2 MAP kinase signaling at multiple levels (Table V, Fig.…”

Hypoxia Enhances Cellular Proliferation and Inositol 1,4,5-Triphosphate Generation in Fibroblasts from Bovine Pulmonary Artery But Not from Mesenteric Artery

“…Activation of cellsurface receptors and their associated tyrosine kinase and phospholipases (C-␥, A 2 , or D) leads to the generation of lipid second messengers (IP 3 and diacyl glycerol [DAG]) (13,14). IP 3 promotes mobilization of intracellular Ca 2ϩ stores (15), whereas DAG is an endogenous activator of protein kinase C (PKC) (14).…”

Cell Type-specific Modulation of Cell Growth by Transforming Growth Factor β1 Does Not Correlate with Mitogen-activated Protein Kinase Activation

“…The cytoplasmic signaling pathway of TGF-␤1 appears to be totally independent of the MAP kinase cascade. Thus, TGF-␤1 is clearly distinct from the majority, if not all, of other peptide growth factors such as EGF, PDGF, and fibroblast growth factor whose ability to modulate cell proliferation is closely associated with the activation of MAP kinases (54). In this respect, interleukin-4 (55), activin A (another member of the TGF-␤ superfamily) (56), and thyrotropin (57, 58) have been reported to induce cellular proliferation without activation of MAP kinases, although thyrotropin has also been shown to induce the activation of MAP kinases in human thyroid cells (59).…”