The giant sarcomeric protein titin contains a protein kinase domain (TK) ideally positioned to sense mechanical load. We identified a signaling complex where TK interacts with the zinc-finger protein nbr1 through a mechanically inducible conformation. Nbr1 targets the ubiquitin-associated p62/SQSTM1 to sarcomeres, and p62 in turn interacts with MuRF2, a muscle-specific RING-B-box E3 ligase and ligand of the transactivation domain of the serum response transcription factor (SRF). Nuclear translocation of MuRF2 was induced by mechanical inactivity and caused reduction of nuclear SRF and repression of transcription. A human mutation in the titin protein kinase domain causes hereditary muscle disease by disrupting this pathway.During muscle differentiation, a specific program of gene expression leads to the translation of myofibrillar proteins and their assembly into contractile units, the sarcomeres, which are constantly remodeled to adapt to changes in mechanical load. The giant protein titin (also known as connectin) acts as a molecular blueprint for sarcomere assembly by providing specific attachment sites for numerous sarcomeric proteins, as well as acting as a molecular spring (1, 2). Titin also contains a catalytic serine-threonine kinase domain (TK), which is inhibited by a specific dual mechanism (3). However, the upstream elements controlling TK activation, its range of cellular substrates, and particularly the role of TK in mature muscle are largely unknown. Spanning half sarcomeres from Z disk to M band, titin is in a unique position to sense mechanical strain along the sarcomere (1). The elastic properties of the titin molecule and the mechanical deformation of the M band during stretch and contraction (4) suggest that the signaling properties of TK might be modulated by mechanically induced conformational changes. Molecular dynamics simulations suggest that mechanical strain can induce a catalytically active conformation of TK (5).The catalytic kinase domain of titin interacts with nbr1. We searched for further elements of a putative signaling pathway that might recognize mechanically induced conformational intermediates of titin's catalytic domain. In a systematic two-hybrid screening approach with various structure-based open states of the catalytic site [kin1, kin2, and kin3 (6)], we identified the zinc-finger protein nbr1 (7) as a TK ligand, which interacted via its Nterminal PB1 domain with the semiopened construct kin3 (Fig. 1, A and B). This interaction was also seen in precipitation experiments with nbr1 and TK-kin3 ( fig. S1A). Kin1, where the complete regulatory domain closes the active site, and kin2, where the a helix R1 (3) is deleted, did not interact. Thus, aR1 was necessary but not sufficient for nbr1 binding, which also required a semiopened catalyt-

The patterning of vertebrate somitic muscle is regulated by signals from neighboring tissues. We examined the generation of slow and fast muscle in zebrafish embryos and show that Sonic hedgehog (Shh) secreted from the notochord can induce slow muscle from medial cells of the somite. Slow muscle derives from medial adaxial myoblasts that differentiate early, whereas fast muscle arises later from a separate myoblast pool. Mutant fish lacking shh expression fail to form slow muscle but do form fast muscle. Ectopic expression of shh, either in wild-type or mutant embryos, leads to ectopic slow muscle at the expense of fast. We suggest that Shh acts to induce myoblasts committed to slow muscle differentiation from uncommitted presomitic mesoderm.

Skeletal muscle fibres are multinucleate syncitial cells that change size during adult life depending on functional demand. The relative contribution of change in nuclear number and/or cell growth to fibre size change is unclear. We report that nuclei/unit length decreases in larger fibres during skeletal muscle ageing. This leads to an increased size of nuclear domain (quantity of cytoplasm/number of nuclei within that cytoplasm). Initially, larger fibres have more satellite cells than small fibres, but this advantage is lost as satellite cells decline with age. These changes are accompanied by an overall decline in fibre size, returning domain size to the normal range. Exacerbated loss of fibre nuclei per unit length during ageing of myoD-null mice provides the first experimental support for the hypothesis that a satellite cell defect causes inadequate nuclear replacement. We propose a model in which a decline in satellite cell function and/or number during ageing leads to a loss of nuclei from large fibres and an associated domain size increase that triggers cytoplasmic atrophy through the normal cell-size-regulating machinery.