The Rho-Guanine Nucleotide Exchange Factor Domain of Obscurin Activates RhoA Signaling in Skeletal Muscle
The Rho-Guanine Nucleotide Exchange Factor Domain of Obscurin Activates RhoA Signaling in Skeletal Muscle
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DOI：
10.1091/mbc.e08-10-1029
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发表时间：
2009-09-01
影响因子：
3.3
通讯作者：
Bloch，Robert J.
中科院分区：
文献类型：
--
作者：
Ford-Speelman，Diana L.;Roche，Joseph A.;Bloch，Robert J.
暗蛋白是横纹肌的一种大的（约800-kDa）模块蛋白，集中在每个肌节的M带和Z盘周围，在那里它被很好地定位以感知收缩活动。Obscurin含有几个信号传导结构域，包括rho-鸟嘌呤核苷酸交换因子（rhoGEF）结构域和串联普列克底物蛋白同源结构域，与肌肉中rho信号传导中的作用一致。我们研究了obscurin的rhoGEF结构域与小GTP酶相互作用并激活小GTP酶的能力。使用体外和体内方法的组合，我们发现，obscurin的rhoGEF结构域选择性地结合到rhoA，和rhoA与obscurin在骨骼肌中的M带共定位。其他小GTP酶，包括rac 1和cdc 42，既不与obscurin的rhoGEF结构域相关，也不集中在M带水平。此外，在成人骨骼肌中obscurin的rhoGEF结构域的过表达选择性地增加了该组织中rhoA的表达和活性。obscurin的rhoGEF结构域的过表达及其对rhoA的影响改变了rho激酶和citron激酶的表达，这两者都可以在其他组织中被rhoA激活。大应变延长收缩引起的啮齿动物后肢肌肉损伤增加了rhoA活性，并将其从M带转移到Z盘，类似于obscurin的rhoGEF结构域过表达的影响。我们的研究结果表明，obscurin的rhoGEF结构域信号至少部分通过诱导rhoA的表达和激活，并在体外和体内改变下游激酶的表达。
Obscurin is a large (∼800-kDa), modular protein of striated muscle that concentrates around the M-bands and Z-disks of each sarcomere, where it is well positioned to sense contractile activity. Obscurin contains several signaling domains, including a rho-guanine nucleotide exchange factor (rhoGEF) domain and tandem pleckstrin homology domain, consistent with a role in rho signaling in muscle. We investigated the ability of obscurin's rhoGEF domain to interact with and activate small GTPases. Using a combination of in vitro and in vivo approaches, we found that the rhoGEF domain of obscurin binds selectively to rhoA, and that rhoA colocalizes with obscurin at the M-band in skeletal muscle. Other small GTPases, including rac1 and cdc42, neither associate with the rhoGEF domain of obscurin nor concentrate at the level of the M-bands. Furthermore, overexpression of the rhoGEF domain of obscurin in adult skeletal muscle selectively increases rhoA expression and activity in this tissue. Overexpression of obscurin's rhoGEF domain and its effects on rhoA alter the expression of rho kinase and citron kinase, both of which can be activated by rhoA in other tissues. Injuries to rodent hindlimb muscles caused by large-strain lengthening contractions increases rhoA activity and displaces it from the M-bands to Z-disks, similar to the effects of overexpression of obscurin's rhoGEF domain. Our results suggest that obscurin's rhoGEF domain signals at least in part by inducing rhoA expression and activation, and altering the expression of downstream kinases in vitro and in vivo.