Src family kinase activity is required for Kit-mediated mitogen-activated protein (MAP) kinase activation, however loss of functional retinoblastoma protein makes MAP kinase activation unnecessary for growth of small cell lung cancer cells.
Small cell lung cancer (SCLC) is characterized by multiple genetic alterations that include inactivation of the retinoblastoma protein (Rb), the establishment of several autocrine loops including that induced by coexpression of stem cell factor (SCF) and Kit, and the ectopic expression and activation of Src family kinases. Previous studies have shown that Lck associates with, and becomes activated by, Kit after SCF stimulation of SCLC cells. In the present study, we have demonstrated that PP1, a pharmacological inhibitor of Src kinases, blocked SCF-mediated activation of mitogen-activated protein (MAP) kinase, but it also inhibited Kit activation. However, MAP kinase activation was more sensitive than Kit activation to the effects of PP1. Overexpression of Lck reduced the sensitivity of MAP kinase activation to PP1 without altering the sensitivity of Kit activation, which suggested a role for Lck in SCF-mediated MAP kinase activation. Inducible expression of a dominant negative Lck inhibited MAP kinase activation in a dose-dependent manner, which confirmed that Src family kinase activity is required for SCF-induced MAP kinase activation. The growth of cells that expressed dominant negative Lck was unaffected, however, despite the inhibition of MAP kinase. Growth was also unaffected by the inhibition of the MAP kinase pathway using PD 98059, but sensitivity to the MAP/extracellular signal-regulated kinase kinase inhibitor could be partially restored by expression of wild-type Rb. Therefore, MAP kinase activation seems to be dispensable for the growth of SCLC only in the absence of Rb expression. These data suggest that the SCF/Kit autocrine loop, through activation of Lck and subsequently MAP kinase, and the mutational inactivation of Rb contribute to the loss of G1-S phase checkpoint regulation during the pathogenesis of SCLC. Furthermore, the data demonstrate that, in established SCLC cell lines, proliferative signal transduction initiated by Kit is mediated by pathways other than the classic MAP kinase pathway.
Department of Microbiology/Immunology, Medical College of Virginia of Virginia Commonwealth University and McGuire Veterans Affairs Medical Center, Richmond 23249, USA.
C
C
Bondzi
J
J
Litz
P
P
Dent
G W
GW
Krystal
D015153:Blotting, Western; D018288:Carcinoma, Small Cell / Q000378:metabolism*; D002455:Cell Division; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D005799:Genes, Dominant; D006801:Humans; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck) / Q000235:genetics / Q000378:metabolism; D020935:MAP Kinase Signaling System; D010766:Phosphorylation; D011233:Precipitin Tests; D019009:Proto-Oncogene Proteins c-kit / Q000235:genetics / Q000378:metabolism; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D016160:Retinoblastoma Protein / Q000378:metabolism*; D019089:Stem Cell Factor / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured
C108123:4-amino-5-(4-methylphenyl)-7-(tert-butyl)pyrazolo(3,4-d)pyrimidine; D004791:Enzyme Inhibitors; D005419:Flavonoids; D011720:Pyrazoles; D011743:Pyrimidines; D016160:Retinoblastoma Protein; D019089:Stem Cell Factor; D019009:Proto-Oncogene Proteins c-kit; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D016392:Proto-Oncogene Proteins pp60(c-src); C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one