Molecular Mechanisms of Steroid Hormone Action: An Examination of Relationships Between Rates of Synthesis of Individual Proteins and the Generation of Metabolic and Lethal Actions of Glucocorticoid Hormones in Lymphoid Cells

摘要

An improved high-resolution two-dimensional gel system for separating complex protein is described that allows at least a three fold increase in the number of proteins detected. Proteins are separated in the first dimension by isoelectric point and in the second dimension by size. Although best resolution is obtained by application of < 100 mu g of protein containing > 5 x 10 exp 6 cpm, as much as 150 mu g of protein may be applied without appreciable loss of resolution. The possibility that hormone-induced changes in the synthesis of individual proteins may serve as initiating events for the rapidly-evolving glucocorticoid-induced metabolic suppressions in rat thymus cells has been examined. Synthetic rates of about 2500 individual proteins have been screened using giant two-dimensional electrophoresis. Dexamethasone induces a rapid increase in synthesis of at least six proteins. The time course suggests that changes in the synthesis of some or all of these four proteins may serve to generate the rapidly evolving hormone-induced inhibition of glucose transport. Proteins associated with the emergence of resistance to glucocorticoids have been examined in the mouse P1798 corticoid-sensitive and-resistant lymphosarcoma cell line as well as in sensitive and resistant (those cells that survive glucorticoid challenge) rat thymus cells. Two-dimensional gel analysis reveals 7 consistent protein differences between highly resistant versus sensitive P1798 lymphosarcoma cells. Analysis of resistant and sensitive rat thymus cells reveals a total of 18 consistent protein differences. These results suggest that resistance of lymphoid cells to glucocorticoid killing in vivo may be related to the relative rates of synthesis of these few proteins, and that common mechanisms of resistance may be employed by both normal and neoplastic tissue. (ERA citation 09:044714)