Building a myofibril from its component proteins requires the interactions of many different proteins in a process whose details are not understood. Several models have been proposed to provide a framework for understanding the increasing data on new myofibrillar proteins and their localizations during muscle development. In this article we discuss four current models that seek to explain how the assembly occurs in vertebrate cross-striated muscles. The models hypothesize: (a) stress fiber-like structures as templates for the assembly of myofibrils, (b) assembly in which the actin filaments and Z-bands form subunits independently from A-band subunits, with the two subsequently joined together to form a myofibril, (c) premyofibrils as precursors of myofibrils, or (d) assembly occurring without any intermediary structures. The premyofibril model, proposed by the authors, is discussed in more detail as it could explain myofibrillogenesis under a variety of different conditions: in ovo, in explants, and in tissue culture studies on cardiac and skeletal muscles.

During myofibril formation, Z-bodies, small complexes of alpha-actinin and associated proteins, grow in size, fuse and align to produce Z-bands. To determine if there were changes in protein dynamics during the assembly process, Fluorescence Recovery after Photobleaching was used to measure the exchange of Z-body and Z-band proteins with cytoplasmic pools in cultures of quail myotubes. Myotubes were transfected with plasmids encoding Yellow, Green or Cyan Fluorescent Protein linked to the Z-band proteins: actin, alpha-actinin, cypher, FATZ, myotilin, and telethonin. Each Z-band protein showed a characteristic recovery rate and mobility. All except telethonin were localized in both Z-bodies and Z-bands. Proteins that were present both early in development in Zbodies and later in Z-bands had faster exchange rates in Z-bodies. These results suggest that during myofibrillogenesis, molecular interactions develop between the Z-band proteins that decrease their mobility and increase the stability of the Z-bands. A truncated construct of alpha-actinin, which localized in Z-bands in myotubes, and exhibited a very low rate of exchange, lead to disruption of myofibrils, suggesting the importance of dynamic, intact alpha-actinin molecules for the formation and maintenance of Z-bands. Our experiments reveal the Z-band to be a much more dynamic structure than its appearance in electron micrographs of cross-striated muscle cells might suggest.

We review some of the problems in determining how myofibrils may be assembled and just as importantly how this contractile structure may be renewed by sarcomeric proteins moving between the sarcomere and the cytoplasm. We also address in this personal review the recent evidence that indicates that the assembly and dynamics of myofibrils are conserved whether the cells are analyzed in situ or in tissue culture conditions. We suggest that myofibrillogenesis is a fundamentally conserved process, comparable to protein synthesis, mitosis, or cytokinesis, whether examined in situ or in vitro.

In this chapter, we present the current knowledge on de novo assembly, growth, and dynamics of striated myofibrils, the functional architectural elements developed in skeletal and cardiac muscle. The data were obtained in studies of myofibrils formed in cultures of mouse skeletal and quail myotubes, in the somites of living zebrafish embryos, and in mouse neonatal and quail embryonic cardiac cells. The comparative view obtained revealed that the assembly of striated myofibrils is a three-step process progressing from premyofibrils to nascent myofibrils to mature myofibrils. This process is specified by the addition of new structural proteins, the arrangement of myofibrillar components like actin and myosin filaments with their companions into so-called sarcomeres, and in their precise alignment. Accompanying the formation of mature myofibrils is a decrease in the dynamic behavior of the assembling proteins. Proteins are most dynamic in the premyofibrils during the early phase and least dynamic in mature myofibrils in the final stage of myofibrillogenesis. This is probably due to increased interactions between proteins during the maturation process. The dynamic properties of myofibrillar proteins provide a mechanism for the exchange of older proteins or a change in isoforms to take place without disassembling the structural integrity needed for myofibril function. An important aspect of myofibril assembly is the role of actin-nucleating proteins in the formation, maintenance, and sarcomeric arrangement of the myofibrillar actin filaments. This is a very active field of research. We also report on several actin mutations that result in human muscle diseases.

The "premyofibril" model of myofibrillogenesis, based on observations in cultured avian muscle cells, proposes that mature myofibrils are preceded by two intermediary structures: premyofibrils and nascent myofibrils. To determine if this model applies to zebrafish skeletal muscle development, we stained developing embryos with antibodies to sarcomeric alpha-actinin and myosin II. In the youngest muscle cells, sarcomeric alpha-actinin and non-muscle myosin II were each localized in linear arrays of small bands that resembled the premyofibrils in avian myocytes. The distribution of muscle-specific myosin II began as scattered short filaments followed in time by overlapping bundles of filaments and organized A-bands in the older somites. Alpha-actinin organization changed from small z-bodies to beaded Z-bands and ordered Z-bands in myofibrils that extended the length of the elongating somites. In older somites with mature myofibrils, premyofibrils were also present at the ends of the mature myofibrils, suggesting that as the cells and somites grew longer, premyofibrils were involved in the elongation of existing mature myofibrils. Fluorescence Recovery After Photobleaching showed that the exchange of proteins (actin, alpha-actinin, FATZ, myotilin and telethonin) between sarcoplasm and the Z-bands of mature myofibrils in zebrafish resembled that seen for the same proteins in cultured avian myotubes, suggesting that myofibril assembly and maintenance in zebrafish share common properties with avian muscle.