Poly-Ig tandems from I-band titin share extended domain arrangements irrespective of the distinct features of their modular constituents

The cellular function of the giant protein titin in striated muscle is a major focus of scientific attention. Particularly, its role in passive mechanics has been extensively investigated. In strong contrast, the structural details of this filament are very poorly understood. To date, only a handful of atomic models from single domain components have become available and data on poly-constructs are limited to scarce SAXS analyses. In this study, we examine the molecular parameters of poly-Ig tandems from I-band titin relevant to muscle elasticity. We revisit conservation patterns in domain and linker sequences of I-band modules and interpret these in the light of available atomic structures of Ig domains from muscle proteins. The emphasis is placed on features expected to affect inter-domain arrangements. We examine the overall conformation of a 6Ig fragment, I65-I70, from the skeletal I-band of soleus titin using SAXS and electron microscopy approaches. The possible effect of highly conserved glutamate groups at the linkers as well as the ionic strength of the medium on the overall molecular parameters of this sample is investigated. Our findings indicate that poly-Ig tandems from I-band titin tend to adopt extended arrangements with low or moderate intrinsic flexibility, independently of the specific features of linkers or component Ig domains across constitutively- and differentially-expressed tandems. Linkers do not appear to operate as free hinges so that lateral association of Ig domains must occur infrequently in samples in solution, even that inter-domain sequences of 4-5 residues length would well accommodate such geometry. It can be expected that this principle is generally applicable to all Ig-tandems from I-band titi

Cooperative control of striated muscle mass and metabolism by MuRF1 and MuRF2

The muscle-specific RING finger proteins MuRF1 and MuRF2 have been proposed to regulate protein degradation and gene expression in muscle tissues. We have tested the in vivo roles of MuRF1 and MuRF2 for muscle metabolism by using knockout (KO) mouse models. Single MuRF1 and MuRF2 KO mice are healthy and have normal muscles. Double knockout (dKO) mice obtained by the inactivation of all four MuRF1 and MuRF2 alleles developed extreme cardiac and milder skeletal muscle hypertrophy. Muscle hypertrophy in dKO mice was maintained throughout the murine life span and was associated with chronically activated muscle protein synthesis. During ageing (months 4–18), skeletal muscle mass remained stable, whereas body fat content did not increase in dKO mice as compared with wild-type controls. Other catabolic factors such as MAFbox/atrogin1 were expressed at normal levels and did not respond to or prevent muscle hypertrophy in dKO mice. Thus, combined inhibition of MuRF1/MuRF2 could provide a potent strategy to stimulate striated muscles anabolically and to protect muscles from sarcopenia during ageing

Changes in Titin and Collagen Underlie Diastolic Stiffness Diversity of Cardiac Muscle

International audienceSmall (N2B) and large (N2BA) cardiac titin isoforms are differentially expressed in a species-specific and heart location-specific manner. To understand how differential expression of titin isoforms may influence passive stiffness of cardiac muscle we investigated the mechanical properties of mouse left ventricular (MLV) wall muscle (expressing predominantly the small titin isoform), bovine left atrial (BLA) wall muscle (predominantly the large isoform), and bovine left ventricular (BLV) wall muscle (expressing small and large isoforms at similar levels). Results indicate that the overall passive muscle stiffness of the muscle types varies nearly tenfold , with stiffness increasing in the following order: BLA, BLV and MLV. To investigate the basis of the variation in the overall muscle stiffness, the contributions of titin and collagen to muscle stiffness were determined. Results showed that increased muscle stiffness results from increases in both titin-and collagen-based passive stiffness, indicating that titin and collagen change in a coordinated fashion. The expression level of the small titin isoform correlates with titin's contribution to overall muscle stiffness, suggesting that differential expression of titin isoforms is an effective means to modulate the filling behavior of the heart

A survey of the primary structure and the interspecies conservation of I-band titin's elastic elements in vertebrates

Witt CC, Olivieri N, Centner T, et al. A survey of the primary structure and the interspecies conservation of I-band titin's elastic elements in vertebrates. Journal of structural biology. 1998;122(1-2):206-215.Titin is a >3000-kDa large filamentous protein of vertebrate-striated muscle, and single titin molecules extend from the Z disc to the M line. In its I-band section, titin behaves extensible and is responsible for myofibrillar passive tension during stretch. However; details of the molecular basis of titin's elasticity are not known. We have compared the motif sequences of titin elastic elements from different vertebrate species and from different regions of the molecule. The I-band titin Ig repeats that are expressed in the stiff cardiac muscle and those that are tissue-specifically expressed in more elastic skeletal muscles represent distinct subgroups. Within the tissue-specifically expressed Ig repeats, a super-repeat structure is found. For the PEVK titin sequences, we surveyed interspecies conservation by hybridization experiments. The sequences of the titin gene which code for the C-terminal region of the PEVK domain are conserved in the genomes of a larger variety of vertebrate, whereas the N-terminal PEVK sequences are more divergent. Future comparisons of titin gene sequences from different vertebrates may improve our understanding of how titin contributes to species diversity of myofibrillar elasticity. Within one species, different classes of Ig repeat families may contribute to elastic diversity of the titin spring in different segments. (C) 1998 Academic Press

Nebulin regulates thin filament length, contractility, and Z-disk structure in vivo

The precise assembly of the highly organized filament systems found in muscle is critically important for its function. It has been hypothesized that nebulin, a giant filamentous protein extending along the entire length of the thin filament, provides a blueprint for muscle thin filament assembly. To test this hypothesis, we generated a KO mouse model to investigate nebulin functions in vivo. Nebulin KO mice assemble thin filaments of reduced lengths and ∼15% of their Z-disks are abnormally wide. Our data demonstrate that nebulin functions in vivo as a molecular ruler by specifying pointed- and barbed-end thin filament capping. Consistent with the shorter thin filament length of nebulin deficient mice, maximal active tension was significantly reduced in KO animals. Phenotypically, the murine model recapitulates human nemaline myopathy (NM), that is, the formation of nemaline rods combined with severe skeletal muscle weakness. The myopathic changes in the nebulin KO model include depressed contractility, loss of myopalladin from the Z-disk, and dysregulation of genes involved in calcium homeostasis and glycogen metabolism; features potentially relevant for understanding human NM