Properties of easily releasable myofilaments: are they the first step in myofibrillar protein turnover?

Abstract

Myofibrillar proteins must be removed from the myofibril before they can be turned over metabolically in functioning muscle cells. It is uncertain how this removal is accomplished without disruption of the contractile function of the myofibril. It has been proposed that the calpains could remove the outer layer of filaments from myofibrils as a first step in myofibrillar protein turnover. Several studies have found that myofilaments can be removed from myofibrils by trituration in the presence of ATP. These easily releasable myofilaments (ERMs) were proposed to be intermediates in myofibrillar protein turnover. It was unclear, however, whether the ERMs were an identifiable entity in muscle or whether additional trituration would remove more myofilaments until the myofibril was gone and whether calpains could release ERMs from intact myofibrils. The present study shows that few ERMs could be obtained from the residue after the first removal of ERMs, and the yield of ERMs from well-washed myofibrils was reduced, probably because some ERMs had been removed by the washing process. Mild calpain treatment of myofibrils released filaments that had a polypeptide composition and were ultrastructurally similar to ERMs. The yield of calpain-released ERMs was two- to threefold greater than the normal yield. Hence, ERMs are an identifiable entity in myofibrils, and calpain releases filaments that are similar to ERMs. The role of ERMs in myofibrillar protein turnover is unclear, because only filaments on the surface of the myofibril would turn over, and changes in myofibrillar protein isoforms during development could not occur via the ERM mechanism.

Label-free Quantitative Protein Profiling of vastus lateralis Muscle During Human Aging

“…Although the bulk of the myofibrillar proteins precipitate, a small percentage of them is easily releasable and found in the LIS extract (17). The three myofibrillar proteins, which we identified in muscle LIS extract as differentially expressed during aging, were less abundant in older women: myosin-1; titin, and myosin light chain 1/3, skeletal muscle isoform.…”

“…Titin is essential for myofibrillar assembly by connecting the Z line to the M line in the sarcomere, and represents also a regulatory node for various transduction pathways (38). The function of the easily releasable proteins remains unknown, although they were suggested to represent intermediate products in the turnover of myofibrillar proteins (39,17). Therefore their reduced levels in old LIS extract may suggest a decreased myofibrillar protein turnover in the old muscle.…”

“…In a shotgun analysis previously performed on whole muscle extracts, the major isoforms of myosin heavy-chain comprise ϳ42% of the total spectra (15). Because these major isoforms may hamper identification of other proteins, we decided to precipitate myofibrils at low ionic strength (16,17) and to focus on the soluble fraction. In this paper, we present the analytical steps of label-free quantitation, which resulted in the identification and quantitation of 255 muscle proteins common to all ten individuals.…”

Inhibition of the Ubiquitin–Proteasome Pathway Does Not Protect against Ventilator-induced Accelerated Proteolysis or Atrophy in the Diaphragm

“…In this regard, myofibrillar proteins constitute 50-60% of muscle protein and are degraded at a faster rate than other classes of muscle proteins during disuse muscle atrophy 41 . In addition, myofibrillar proteins have also been shown to be the principle protein target of the UPP 42 .…”

Titin-based mechanosensing and signaling: role in diaphragm atrophy during unloading?

“…The calpain family is a group of intracellular cysteine proteases requiring calcium for activity; calpain-1 and -2 are expressed ubiquitously, and calpain-3, also known as p94, is striated muscle specific. Evidence indicates that activation of calpains is an initial step in myofilament proteolysis by cleavage of myosin and actin (38). In this way, calpains yield fragments that are degradable by the ubiquitin-proteasome pathway.…”