Myofibril assembly and the roles of the ubiquitin proteasome system

Abstract

De novo assembly of myofibrils in vertebrate cross-striated muscles progresses in three distinct steps, first from a minisarcomeric alignment of several nonmuscle and muscle proteins in premyofibrils, followed by insertions of additional proteins and increased organization in nascent myofibrils, ending with mature contractile myofibrils. In a search for controls of the process of myofibril assembly, we discovered that the transition from nascent to mature myofibrils could be halted by inhibitors of three distinct functions of the ubiquitin proteasome system (UPS). First, inhibition of pathway to E3 Cullin ligases that ubiquitinate proteins led to an arrest of myofibrillogenesis at the nascent myofibril stage. Second, inhibition of p97 protein extractions of ubiquitinated proteins led to a similar arrest of myofibrillogenesis at the nascent myofibril stage. Third, inhibitors of proteolytic action by proteasomes also blocked nascent myofibrils from transitioning to mature myofibrils. In contrast, inhibitors of autophagy or lysosomes did not affect myofibrillogenesis. To probe for differences in the effects of UPS inhibitors during myofibrillogenesis, we analyzed by fluorescence recovery after photobleaching the exchange rates of two selected sarcomeric proteins (muscle myosin II heavy chains and light chains). In the presence of p97 and proteasomal inhibitors, the dynamics of each of these two myosin proteins decreased in the nascent myofibril stage, but were unaffected in the mature myofibril stage. The increased stability of myofibrils occurring in the transition from nascent to mature myofibril assembly indicates the importance of dynamics and selective destruction in the muscle myosin II proteins for the remodeling of nascent to mature myofibrils.

The Role of Embryonic Chick Muscle Cell Culture in the Study of Skeletal Myogenesis

“…Concomitant with the progressive alignment of Z-line molecules, thick filaments, formed by the muscle isoform of myosin II, are linked to the nascent myofibrils. Lastly, M-lines are assembled with the addition of myomesin and other proteins, including the M-line part of titin ( Sanger et al, 2010 ; Wang et al, 2020 ). In summary, the assembly of sarcomeres involves interaction between two organizing centers, (i) one involves the assembly of thick filaments and their associated proteins, including titin, and (ii) the other involves the assembly of thin filaments and their associated proteins, forming the I-Z-I complex ( Hill et al, 1986 ).…”

Breakdown of Filamentous Myofibrils by the UPS–Step by Step

“…Studies in animal models have indicated an important role for p97/VCP in promoting muscle development and homeostasis. Loss of p97/VCP function in skeletal or cardiac muscles interferes with autophagy [70], and myofibril assembly [71,72]. In addition, knockdown of TER94, the Drosophila ortholog for p97/VCP, perturbs myofibril organization and function in Drosophila hearts, and causes cardiomyopathies [73].…”

Effect of MG‐132 on myofibrillogenesis and the ubiquitination of GAPDH in quail myotubes

“…among the other models of myofibril formation described in the literature, the three-step model for myofibrillogenesis is gaining the most attention (Sanger et al, 2005(Sanger et al, , 2009(Sanger et al, , 2010(Sanger et al, , 2017J. Wang et al, 2020).…”

“…J. Wang et al (2020) have reported that the transition from nascent to mature myofibrils could be halted by inhibitors of three distinct functions of the Ubiquitin Proteasome System (UPS)-inhibition of E3 Cullin ligases that ubiquitinate sarcomeric proteins, or inhibition of p97 extractions of ubiquitinated sarcomeric proteins, or inhibition of proteosomes where old or misfolded sarcomeric proteins would be proteolyzed.…”

“…Cultured living quail muscles have been used to study the maintenance of different muscle proteins in myofibrils (Wang et al, 2005a), and to elucidate how proteasome inhibitors used to treat human cancers in humans have deleterious effects on human muscles (J. Wang et al, 2020). Cultured embryonic quail myotubes, which upon transfection with pYFPα-actinin, reveals the different stages of myofibrils, for example premyofibrils, nascent myofibrils, and mature myofibrils, when coupled with specific anti-sarcomeric protein stainings (J.…”