Proteoglycans are present in the transverse tubule system of skeletal muscle

Abstract

The transverse tubule system (T-tubule, T-system) of skeletal muscle is a membranous network that penetrates the interior of myofibers. The T-system is continuous with the sarcolemma and therefore provides a path for membrane excitation to reach internal myofibrils. In this study we demonstrate that T-tubules in elasmobranch fish, frog, and rat skeletal muscle contain a matrix of chondroitin sulfate proteoglycans. We used anti-T1, a mouse monoclonal antibody that recognizes a rare chondroitin sulfate epitope, for immunolocalization and biochemical studies. First, we find that T1 immunoreactivity colocalizes with a T-tubule marker, the dihydropyridine receptor alpha 2 subunit, in both frog and fish muscle. Secondly, the distribution of T1 immunoreactivity exactly matches the different distribution of T-tubules in rat and frog muscle. In rat muscle, two bands of T1 immunoreactivity are detected per sarcomere, a distribution that corresponds to the T-tubules located at the two A-I junctions of each sarcomere. In frog muscle, we detect one band of T1 immunoreactivity per sarcomere that corresponds to the one T-tubule per sarcomere located at the Z line. Lastly, we have isolated and biochemically characterized T1 antigenicity from fish skeletal muscle. Like extracellular matrix proteoglycans of cartilage, T1 antigenicity requires denaturing conditions to be solubilized. In fish muscle, two chondroitin sulfate proteoglycans bear T1: a heavily glycosylated proteoglycan with a molecular mass of about 1000 kDa, and a smaller proteoglycan that has a mobility on SDS-PAGE like a protein of molecular mass 280 kDa. We propose that proteoglycans function as structural components in the T-system. The proteoglycans may form a matrix, like the one formed by the cartilage proteoglycans they resemble, that can withstand the cytosolic osmotic pressures present in muscle cells and therefore may prevent the T-tubule from collapsing. We present a quantitative argument in support of this hypothesis.