Transient Loss of αB-Crystallin: An Early Cellular Response to Mechanical Stretch

Abstract

Human trabecular meshwork (HTM) is distended and stretched with increases in intraocular pressure. During this stretching, there is a rearrangement of actin filaments. The HTM cells express αB-crystallin, a small heat shock protein that may have a role in the stabilization and regulation of the cytoskeleton in mammalian cells. The levels of αB-crystallin were examined in trabecular meshwork cells after mechanical stretch. Human TM primary cell cultures, plated onto silicone sheets, were subjected to a single 10% linear stretch and samples were prepared at various times after stretch for immunoblotting or Northern blotting. Immunoblots of total protein extracts with antibody specific for αB-crystallin detected a 26% decrease of cellular αB-crystallin levels within 2 minutes. After 1 hour αB-crystallin levels had decreased 90% compared to control cells. The levels of αB-crystallin began to recover in cells stretched for 2 hours and returned to initial levels by 24 hours. Northern blots probed with αB-crystallin exon III cDNA detected a transcript of 0.65 kb in human TM cells and the levels of the αB mRNA remained constant during αB-crystallin protein decrease. Later, levels of the 0.65 kb transcript of αB-crystallin increased during the cellular recovery. These results suggest that decreased levels of αB-crystallin after mechanical stretch were probably not due to transcriptional changes but rather to increased degradation of αB-crystallin protein. An increase in mRNA levels may play a role in the recovery of αB-crystallin during reorganization of the cytoskeleton and attachment to the substratum. These data raise the possibility of a specific proteolysis of αB-crystallin protein in cells after a physiological challenge.