Formation of junctions involved in excitation-contraction coupling in skeletal and cardiac muscle.

During excitation-contraction (e-c) coupling of striated muscle, depolarization of the surface membrane is converted into Ca2+ release from internal stores. This process occurs at intracellular junctions characterized by a specialized composition and structural organization of membrane proteins. The coordinated arrangement of the two key junctional components--the dihydropyridine receptor (DHPR) in the surface membrane and the ryanodine receptor (RyR) in the sarcoplasmic reticulum--is essential for their normal, tissue-specific function in e-c coupling. The mechanisms involved in the formation of the junctions and a potential participation of DHPRs and RyRs in this process have been subject of intensive studies over the past 5 years. In this review we discuss recent advances in understanding the organization of these molecules in skeletal and cardiac muscle, as well as their concurrent and independent assembly during development of normal and mutant muscle. From this information we derive a model for the assembly of the junctions and the establishment of the precise structural relationship between DHPRs and RyRs that underlies their interaction in e-c coupling.

Increased mRNA levels for components of the lysosomal, Ca2+-activated, and ATP-ubiquitin-dependent proteolytic pathways in skeletal muscle from head trauma patients.

The cellular mechanisms responsible for enhanced muscle protein breakdown in hospitalized patients, which frequently results in lean body wasting, are unknown. To determine whether the lysosomal, Ca2+-activated, and ubiquitin-proteasome proteolytic pathways are activated, we measured mRNA levels for components of these processes in muscle biopsies from severe head trauma patients. These patients exhibited negative nitrogen balance and increased rates of whole-body protein breakdown (assessed by [13C]leucine infusion) and of myofibrillar protein breakdown (assessed by 3-methylhistidine urinary excretion). Increased muscle mRNA levels for cathepsin D, m-calpain, and critical components of the ubiquitin proteolytic pathway (i.e., ubiquitin, the 14-kDa ubiquitin-conjugating enzyme E2, and proteasome subunits) paralleled these metabolic adaptations. The data clearly support a role for multiple proteolytic processes in increased muscle proteolysis. The ubiquitin proteolytic pathway could be activated by altered glucocorticoid production and/or increased circulating levels of interleukin 1beta and interleukin 6 observed in head trauma patients and account for the breakdown of myofibrillar proteins, as was recently reported in animal studies.

Structural interpretation of the mutations in the beta-cardiac myosin that have been implicated in familial hypertrophic cardiomyopathy.

In 10-30% of hypertrophic cardiomyopathy kindreds, the disease is caused by > 29 missense mutations in the cardiac beta-myosin heavy chain (MYH7) gene. The amino acid sequence similarity between chicken skeletal muscle and human beta-cardiac myosin and the three-dimensional structure of the chicken skeletal muscle myosin head have provided the opportunity to examine the structural consequences of these naturally occurring mutations in human beta-cardiac myosin. This study demonstrates that the mutations are related to distinct structural and functional domains. Twenty-four are clustered around four specific locations in the myosin head that are (i) associated with the actin binding interface, (ii) around the nucleotide binding site, (iii) adjacent to the region that connects the two reactive cysteine residues, and (iv) in close proximity to the interface of the heavy chain with the essential light chain. The remaining five mutations are in the myosin rod. The locations of these mutations provide insight into the way they impair the functioning of this molecular motor and also into the mechanism of energy transduction.

Abnormal junctions between surface membrane and sarcoplasmic reticulum in skeletal muscle with a mutation targeted to the ryanodine receptor.

Junctions that mediate excitation-contraction (e-c) coupling are formed between the sarcoplasmic reticulum (SR) and either the surface membrane or the transverse (T) tubules in normal skeletal muscle. Two structural components of the junctions, the feet of the SR and the tetrads of T tubules, have been identified respectively as ryanodine receptors (RyRs, or SR calcium-release channels), and as groups of four dihydropyridine receptors (DHPRs, or voltage sensors of e-c coupling). A targeted mutation (skrrm1) of the gene for skeletal muscle RyRs in mice results in the absence of e-c coupling in homozygous offspring of transgenic parents. The mutant gene is expected to produce no functional RyRs, and we have named the mutant mice "dyspedic" because they lack feet--the cytoplasmic domain of RyRs anchored in the SR membrane. We have examined the development of junctions in skeletal muscle fibers from normal and dyspedic embryos. Surprisingly, despite the absence of RyRs, junctions are formed in dyspedic myotubes, but the junctional gap between the SR and T tubule is narrow, presumably because the feet are missing. Tetrads are also absent from these junctions. The results confirm the identity of RyRs and feet and a major role for RyRs and tetrads in e-c coupling. Since junctions form in the absence of feet and tetrads, coupling of SR to surface membrane and T tubules appears to be mediated by additional proteins, distinct from either RyRs or DHPRs.