Regulation of Actin Interacting Protein Drebrin by Mutations in HSPB1

Abstract

Heat shock protein B1 (HspB1) is an ATP‐independent molecular chaperone with a conserved α‐crystalline domain in the C‐termini region. A number of mutations in HspB1 have been identified in families with Charcot Marie Tooth type IIF (CMT‐IIF) disease or with distal hereditary motor neuropathy II (dHMN II). Mutations outside the crystalline domain (P39L, G84L, P128S, P182L) associate with dHMN. Mutations S135F, R136W and R127W associate with a CMT‐IIF phenotype.Wild type (wt) HspB1 reduces RhoA activation by translational inhibition of PDZ‐RhoGEF, a RhoA specific GEF. Decreased expression of PDZ‐RhoGEF by HspB1 was accomplished by enhancing expression of specific microRNAs (miR20a and miR128) that inhibit the translation of PDZ‐RhoGEF and resulted in an increased neurite growth response. The regulatory effect of HspB1 on RhoA activity is blocked by mutation in S135F of HspB1 but not by R136W and R127W. This suggested either that these HspB1 mutations alter independent cellular mechanisms or that their effects may converge downstream of RhoA GTPase signaling to cause similar disease phenotype.We constructed lentiviral vectors carrying full length human HspB1 cDNA of wt or mutant S135F, R136W or R127W. Cortical neurons derived from SD rat pups E15 were transfected to expressed wt or mutant HspB1transgene to determine their effect on actin cytoskeleton.The studies showed S135F, R16W and R127W mutations cause HspB1 to partition more readily with the cytoskeletal fraction and enhance the binding of HspB1 to F‐actin leading to the loss of Drebrin, an F‐actin side‐binding protein, from the F‐actin domain. These findings highlight the diverse mechanisms by which HspB1 functions as an important regulatory protein of the actin cytoskeleton.