HSPB1, HSPB6, HSPB7 and HSPB8 Protect against RhoA GTPase-Induced Remodeling in Tachypaced Atrial Myocytes

BACKGROUND: We previously demonstrated the small heat shock protein, HSPB1, to prevent tachycardia remodeling in in vitro and in vivo models for Atrial Fibrillation (AF). To gain insight into its mechanism of action, we examined the protective effect of all 10 members of the HSPB family on tachycardia remodeling. Furthermore, modulating effects of HSPB on RhoA GTPase activity and F-actin stress fiber formation were examined, as this pathway was found of prime importance in tachycardia remodeling events and the initiation of AF. METHODS AND RESULTS: Tachypacing (4 Hz) of HL-1 atrial myocytes significantly and progressively reduced the amplitude of Ca²⁺ transients (CaT). In addition to HSPB1, also overexpression of HSPB6, HSPB7 and HSPB8 protected against tachypacing-induced CaT reduction. The protective effect was independent of HSPB1. Moreover, tachypacing induced RhoA GTPase activity and caused F-actin stress fiber formation. The ROCK inhibitor Y27632 significantly prevented tachypacing-induced F-actin formation and CaT reductions, showing that RhoA activation is required for remodeling. Although all protective HSPB members prevented the formation of F-actin stress fibers, their mode of action differs. Whilst HSPB1, HSPB6 and HSPB7 acted via direct prevention of F-actin formation, HSPB8-protection was mediated via inhibition of RhoA GTPase activity. CONCLUSION: Overexpression of HSPB1, as well as HSPB6, HSPB7 and HSPB8 independently protect against tachycardia remodeling by attenuation of the RhoA GTPase pathway at different levels. The cardioprotective role for multiple HSPB members indicate a possible therapeutic benefit of compounds able to boost the expression of single or multiple members of the HSPB family

The human genome encodes ten alpha-crystallin-related small heat shock proteins: HspB1-10

To obtain an inventory of all human genes that code for alpha-crystallin-related small heat shock proteins (sHsps), the databases available from the public International Human Genome Sequencing Consortium (IHGSC) and the private Celera human genome project were exhaustively searched. Using the human Hsp27 protein sequence as a query in the protein databases, which are derived from the predicted genes in the human genome, 10 sHsp-like proteins were retrieved, including Hsp27 itself. Repeating the search procedure with all 10 proteins and with a variety of more distantly related animal sHsps, no further human sHsps were detected, as was the case when searches were performed at deoxyribonucleic acid level. The 10 retrieved proteins comprised the 9 earlier recognized human sHsps (Hsp27/HspB1, HspB2, HspB3, alphaA-crystallin/HspB4, alphaB-crystallin/HspB5, Hsp20/HspB6, cvHsp/HspB7, H11/HspB8, and HspB9) and a sperm tail protein known since 1993 as outer dense fiber protein 1 (ODF1). Although this latter protein probably serves a structural role and has a high cysteine content (14%), it clearly contains an alpha-crystallin domain that is characteristic for sHsps. ODF1 can as such be designated as HspB10. The expression of all 10 human sHsp genes was confirmed by expressed sequence tag (EST) searches. For Hsp27/HspB1, 2 retropseudogenes were detected. The HspB1-10 genes are dispersed over 9 chromosomes, reflecting their ancient origin. Two of the genes (HspB3 and HspB9) are intronless, and the others have 1 or 2 introns at various positions. The transcripts of several sHsp genes, notably HspB7, display low levels of alternative splicing, as supported by EST evidence, which may result in minor amounts of isoforms at the protein level