Growth rate dependence of differential incorporation of a guanosine triphosphate photoaffinity probe into the alpha subunit of a guanine nucleotide binding protein, Gs, from metastatic variants of B16 melanoma cells.
The signal transducing regulatory protein (Gs alpha) was examined in B16 melanoma clones of low (F1C29) and high (F10C23) experimental metastatic potential. Incorporation of the photoaffinity analogue, [8-azido-gamma-32P]GTP, into Gs alpha was decreased in F10C23 extracts when compared to F1C29. This difference disappeared when the photolabeling reaction was carried out at an elevated temperature which enhanced the rate of GTP exchange, suggesting functional differences in the ability of Gs alpha to bind or release GTP rather than dissimilar intracellular Gs alpha concentrations. Differential Gs alpha photolabeling occurred only during the period of rapid growth when F10C23 cells proliferated faster than F1C29 cells. During the recovery phase of growth immediately following plating and at confluence, periods in which F1C29 and F10C23 growth rates are similar, Gs alpha photolabeling between the two clones was equal. CMT lung carcinoma clones of differential metastatic potential grew at a uniform rate at all stages of growth and also exhibited equal Gs alpha photolabeling. F10C23 cells were more responsive to alpha-melanocyte-stimulating hormone stimulation of adenylate cyclase activity than F1C29 cells at all growth stages. These results confirm previously observed functional differences in Gs alpha between B16 metastatic variants and show that photolabeling differences in Gs alpha are related to growth rate.
Lange-Carter
C A
CA
Molecular and Environmental Toxicology Program, School of Pharmacy, University of Colorado, Boulder 80309-0297.
Droms
K A
KA
Lester
B R
BR
Malkinson
A M
AM
D000345:Affinity Labels / Q000378:metabolism*; D000818:Animals; D001345:Autoradiography; D001386:Azides / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D014644:Genetic Variation; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism; D007700:Kinetics; D046911:Macromolecular Substances; D008546:Melanoma, Experimental / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D010761:Phosphorus Radioisotopes; D010777:Photochemistry
C021961:8-azidoguanosine triphosphate; D000345:Affinity Labels; D001386:Azides; D046911:Macromolecular Substances; D010761:Phosphorus Radioisotopes; D006160:Guanosine Triphosphate; D000242:Cyclic AMP; D019204:GTP-Binding Proteins