Department of Otolaryngology-Head and Neck Surgery, Beijing Tongren Hospital, Capital Medical University, Beijing, China.
Key Laboratory of Otolaryngology Head and Neck Surgery, Ministry of Education, Capital Medical University, Beijing, China.
Laryngoscope. 2024 Feb;134(2):855-864. doi: 10.1002/lary.31021. Epub 2023 Sep 2.
Muscle RING-finger protein-1 (MuRF-1), an E3 ubiquitin ligase, has been reported to aggravate skeletal muscle denervated atrophy by mediating the ubiquitination degradation of multiple proteins, whereas the molecular mechanism underlying MuRF-1-mediated internal laryngeal muscle denervated atrophy remains unknown.
A rat unilateral recurrent laryngeal nerve (RLN) transection model was established to evaluate denervated muscle atrophy of the larynx. The expression of MuRF-1, G- and F-actin in thyroarytenoid muscle (TA) myocytes before and after RLN injury was analyzed by immunofluorescence and Western blotting. Coimmunoprecipitation experiments detected molecular interactions between MuRF-1 and G-actin. Immunoprecipitation tested MuRF-1-mediated ubiquitination of G-actin in denervated and innervated TA muscle tissues. The shRNA-MuRF-1 AAV was used to suppress MuRF-1 expression in denervated TA muscles in vivo.
First, MuRF-1 expression was significantly elevated in denervated TA muscle compared to innervated TA muscle (p < 0.001). Second, there was a progressive increase in the G/F-actin ratio in TA myocytes from day 3 to 14 after RLNI (p < 0.01). Furthermore, colocalization of MuRF-1 and G-actin in denervated TA myocytes was observed. Moreover, the upregulation of MuRF-1 was closely associated with the ubiquitination of G-actin in denervated TA myocytes and muscle tissues. Knockdown of MuRF-1 decelerated the degree of TA muscle atrophy compared with that in the Blank and NC groups (p < 0.001) but seemed to promote the compensatory movement of the healthy side.
Collectively, we illustrate a novel molecular mechanism underlying MuRF-1-mediated internal laryngeal muscle denervated atrophy in that MuRF-1 could promote disequilibrium of the G/F-actin ratio by regulating G-actin ubiquitination.
NA Laryngoscope, 134:855-864, 2024.
肌肉环指蛋白-1（MuRF-1）是一种 E3 泛素连接酶，已被报道通过介导多种蛋白质的泛素化降解来加重骨骼肌失神经萎缩，然而 MuRF-1 介导的喉内肌失神经萎缩的分子机制尚不清楚。
建立单侧大鼠喉返神经（RLN）切断模型，以评估喉的失神经萎缩。通过免疫荧光和 Western blot 分析 RLN 损伤前后甲杓肌（TA）肌细胞中 MuRF-1、G-和 F-肌动蛋白的表达。共免疫沉淀实验检测 MuRF-1 与 G-肌动蛋白之间的分子相互作用。免疫沉淀试验检测失神经和神经支配的 TA 肌肉组织中 MuRF-1 介导的 G-肌动蛋白泛素化。使用 shRNA-MuRF-1 AAV 在体内抑制失神经 TA 肌肉中的 MuRF-1 表达。
首先，与神经支配的 TA 肌肉相比，失神经 TA 肌肉中的 MuRF-1 表达显著升高（p<0.001）。其次，RLNI 后第 3 天至 14 天，TA 肌细胞中的 G/F-肌动蛋白比值逐渐增加（p<0.01）。此外，在失神经 TA 肌细胞中观察到 MuRF-1 与 G-肌动蛋白的共定位。此外，MuRF-1 的上调与失神经 TA 肌细胞和肌肉组织中 G-肌动蛋白的泛素化密切相关。与空白组和 NC 组相比，MuRF-1 敲低组 TA 肌肉萎缩程度减慢（p<0.001），但似乎促进了健侧的代偿运动。
总的来说，我们阐明了 MuRF-1 介导的喉内肌失神经萎缩的新分子机制，即 MuRF-1 可以通过调节 G-肌动蛋白泛素化来促进 G/F-肌动蛋白比值的失衡。
NA Laryngoscope，134：855-864，2024。
