The heat shock proteins A4 (HSPA4) ameliorates pathological myocardial remodeling through improving protein homeostasis

蛋白质质量控制（PQC）依赖于主要由分子伴侣进行的蛋白质适当折叠、泛素-蛋白酶体（UPS）系统介导的靶向蛋白水解和自噬之间的协同作用。蛋白质病是由蛋白质错误折叠引起的以蛋白质异常聚集为特征的疾病。有丝分裂后的细胞如心肌细胞不能简单地通过细胞分裂或再生来避免蛋白质错误折叠。因此，心脏中错误折叠蛋白负荷的增加可能导致心肌功能障碍的进展。通过药物干预改善PQC的措施代表了治疗人类蛋白质病变和心力衰竭的一种新的治疗潜力。作为分子伴侣，热休克蛋白（HSPs）有助于维持细胞中适当的PQC。HSPA4是HSP70家族的一员，在应激条件下的细胞存活中起作用。我们已经证明HSPA4蛋白水平在病理重构的心脏中增加。有趣的是，经典Hspa4基因敲除（Hspa4-/-）小鼠显示与多泛素化蛋白聚集相关的病理性重塑明显增加。我们迄今为止的数据表明，Hspa4-/-心脏中泛素聚集体的积累不是由蛋白酶体缺陷引起的。在Hspa4-/-心脏中，通过自噬优先降解的洗涤剂不溶性部分中检测到更多泛素化蛋白聚集。2)。蛋白酶体活性和蛋白酶体的典型底物p53蛋白水平在Hspa4+/+和Hspa4-/-心脏中均无差异影响。3)。在Hspa4-/-心脏中观察到自噬体的积累。因此，我们希望深入研究自噬作为蛋白质聚集体积累的可能机制，这可能是导致病理性重塑大大增加的原因。为此，我们利用腺病毒建立了HSPA4的体外过表达，并利用siRNA建立了HSPA4的体外低表达。此外，我们目前正在使用腺相关病毒（AAV9）在体内产生心肌细胞特异性HSPA4过表达。利用功能获得和功能丧失的方法，我们希望探索HSPA4蛋白抑制病理性心肌重塑并延缓心力衰竭进展的假设，可能是通过改善自噬介导的有毒错误折叠蛋白的降解。此外，我们希望确定控制Hspa4表达的转录因子。

Protein quality control (PQC) depends on the collaboration between proper protein folding, which is mainly performed by molecular chaperones, and targeted proteolysis mediated by the ubiquitin-proteasome (UPS) system and autophagy. Proteinopathies are diseases caused by protein misfolding and characterized by abnormal protein aggregation. Post-mitotic cells such as cardiomyocytes cannot simply avoid protein misfolding by cell division or regeneration. An increased load of misfolded proteins in the heart may therefore contribute to a progression of myocardial dysfunction. Measures to improve PQC by pharmacological intervention represent a novel therapeutic potential to treat human proteopathies and thereby heart failure. As molecular chaperones, the heat shock proteins (HSPs) help to maintain the proper PQC in the cells. HSPA4 is a member of the HSP70 family that plays a role in cell survival under stress conditions. We have already shown that HSPA4 protein levels increase in pathologically-remodeled hearts. Interestingly, classical Hspa4 knockout (Hspa4-/-) mice show markedly increased pathological remodeling associated with the aggregation of polyubiquitinated proteins. Our data to date indicate that the accumulation of ubiquitin aggregates in Hspa4-/- hearts was not caused by a defect in the proteasome: 1.) More aggregation of ubiquitinated proteins were detected in In Hspa4-/- hearts in the detergent-insoluble fraction that is preferentially degraded by autophagy. 2.) Neither proteasome activity nor p53 protein level, a typical substrate of the proteasome, was differentially affected in Hspa4+/+ and Hspa4-/- hearts. 3.) An accumulation of autophagosomes was observed in Hspa4-/- hearts. Therefore, we want to deeply investigate the autophagy as a possible mechanism for the accumulation of protein aggregates, which may be responsible for the greatly increased pathological remodeling. For this purpose, we established in vitro HSPA4 overexpression using adenovirus, as well as HSPA4 knockdown using siRNA. Moreover, we are currently generating in vivo cardiomyocyte-specific HSPA4 overexpression using adeno-associated virus (AAV9). Using gain and loss-of-function approaches, we would like to explore the hypothesis that HSPA4 protein suppresses pathologic myocardial remodeling and delays the progression to heart failure, possibly by improving autophagy-mediated degradation of toxic, misfolded proteins. Moreover, we want to identify the transcription factors controlling Hspa4 expression.