Intermediate Filaments & Costamere Structure & Function

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Costameres are structures at the surface of striated muscle that align the sarcolemma regularly with nearby myofibrils and transmit contractile force laterally, through the membrane to the extracellular matrix. Costameres must therefore be linked to nearby structures firmly enough to withstand the forces of contraction. Here we focus on links between costameres and myofibrils. Desmin-based and keratin-based intermediate filaments (IFs), as well as actin microfilaments, link superficial myofibrils to costameres at the sarcolemma. We postulate that IFs containing keratins 8 and 19 play an important role at costameres because, unlike desmin, they are found at all costameric structures. We have shown that K8/K19 IFs interact with the dystrophin- glycoprotein complex via K19, and that over-expression of K19 or elimination of K19 by homologous recombination disrupts costameres. The K19-null mutation in mice also leads to mitochondrial mislocalization, increased serum creatine kinase levels and reduced contractile force, suggesting that the absence of IFs containing K19 causes a mild skeletal myopathy. We now have mice in the same genetic background as the K19-null (FVB) that lack desmin, and that lack both K19 and desmin. Here we propose 5 aims to test the hypothesis that K19-based IFs together with desmin IFs form distinct but complementary links between the contractile apparatus and costameres in healthy muscle. We further propose that their absence either alone or together leads to a weakening of the links between costameres and the contractile apparatus, compromised muscle physiology, and increased susceptibility to injury induced by lengthening ("eccentric") contractions. Aim1 addresses the morphological changes in costameres, sarcomeres and intracellular organelles, including mitochondria, in K19-null, desmin-null, and double knock-out (DKO) mice. In Aim 2, we will characterize the contractile activity, running ability, and susceptibility to eccentric injury of the individual mutants and the DKO. Aim 3 addresses the biomechanical properties of muscles lacking K19, desmin or both IF proteins, by investigating the strength of connections between myofibrils, and between myofibrils and the sarcolemma. Our results should therefore elucidate the molecular pathways that transmit force in healthy muscle, and how the absence of IFs can lead to muscle disease.