Application of percent labeled mitoses (PLM) analysis to the investigation of melanoma cell responsiveness to MSH stimulation throughout the cell cycle

Abstract

Using a modification of the percent labeled mitoses (PLM) method of cell cycle analysis, the approximate lengths of the cell cycle phases of Cloudman S91 mouse melanoma cells were determined to be: G1, 10.0 h; S, 8.0 h; G2, 2.0 h; and M, 4.0 h. Studies were then carried out to investigate melanoma cell responsiveness to MSH (melanocyte stimulating hormone) throughout the cell cycle. In contrast to previous reports of a G2-specific cell response to MSH, melanoma cells demonstrated increased tyrosinase activity when exposed to MSH for the time periods: 0–8 h, 8–16 h, or 16–24 h. These times correspond to late M-G1; G1-S; and late S-G2-M, respectively. Cells exposed to cycloheximide (10 −6 M) during any of these time periods were inhibited from responding to MSH, suggesting that continued protein synthesis is required throughout the cell cycle for MSH action. These results demonstrate that MSH responsiveness is not necessarily expressed exclusively during the G2 phase of the cell cycle in Cloudman S91 mouse melanoma cells in culture, as reported by others.