C6ORF32, AN HDAC6-BINDING PROTEIN THAT REGULATES MYOBLAST DIFFERENTIATION

描述(申请人提供)：骨骼肌由成肌细胞融合形成的多核合胞体组成。通过微阵列研究发现，当肌管含有2-5个肌核时，C6ORF32的表达在人成肌细胞融合的早期上调(Cerletti等人，2006年)。C6ORF32表达的下调导致成肌细胞融合受阻，这是由于肌肉生成素的表达显著减少(Yoon等人，2007年)。通过计算分析，预测的C6ORF32蛋白只包含一个已知功能的结构域，即推测的HDAC结合域。研究发现，C6ORF32与HDAC6结合，HDAC6是一种IIb类HDAC，具有促进细胞运动的功能。假设HDAC6和C6ORF32在功能上是相连的，并且HDAC6和C6ORF32之间的相互作用通过降低HDAC6的去乙酰化活性来正向调节成肌分化。我们将通过下列特定目的在体内外研究骨骼肌中HDAC6和C6ORF32之间的调控联系：1)确定C6ORF32与HDAC6的关键结合区域，并证明这两种蛋白质在体内存在结合；2)确定HDAC6是否通过调节C6ORF32的乙酰化/去乙酰化来调节成肌细胞的分化；3)评估C6ORF32是否通过调节HDAC6的活性来调节成肌细胞的分化；4)确定在体内阻断C6ORF32的表达是否会通过影响HDAC6的功能而导致肌肉发育、功能和再生方面的缺陷。这些研究将为了解未来针对C6ORF32和HDAC6之间调节相互作用的药物治疗是否能够增强肌源性修复奠定基础。

DESCRIPTION (provided by applicant): Skeletal muscle is comprised of multinucleated syncytia formed by the fusion of myoblasts. Through microarray studies, it was found that expression of C6ORF32 is upregulated early during human myoblast fusion, when myotubes contain between 2-5 myonuclei (Cerletti et al., 2006). Downregulation of C6ORF32 expression causes a block in myoblast fusion due to a significant decrease in myogenin expression (Yoon et al., 2007). By computational analyses, the predicted C6ORF32 protein contains only one domain of known function, a putative HDAC-binding domain. It was found that C6ORF32 binds to HDAC6, a class IIb HDAC known for its function in promoting cell motility. It is hypothesized that HDAC6 and C6ORF32 are functionally connected, and the interaction between HDAC6 and C6ORF32 positively regulates myogenic differentiation by reducing the de-acetylation activity of HDAC6. The regulatory connection between HDAC6 and C6ORF32 in skeletal muscle will be studied both in vitro and in vivo via the following specific aims: 1) Define the critical binding region of C6ORF32 to HDAC6, and demonstrate that binding of these two proteins occurs in vivo; 2) Determine if HDAC6 regulates myoblast differentiation by modulating C6ORF32 acetylation/de-acetylation; 3) Evaluate if C6ORF32 regulates myoblast differentiation by modulating HDAC6 activity; 4) Determine if ablation of C6ORF32 expression in vivo yields to defects in muscle development, function and regeneration by affecting HDAC6 function. These studies will constitute the groundwork for understanding whether future pharmacological therapies targeting the regulatory interaction between C6ORF32 and HDAC6 are amenable to enhance myogenic repair.