Embryonic and post-embryonic utilization and subcellular localization of the nuclear receptor SpSHR2 in the sea urchin

SpSHR2 (Strongylocentrotus purpuratus steroid hormone receptor 2) is a nuclear receptor, encoded by a maternal RNA in the sea urchin embryo. These maternal SpSHR2 transcripts, which are present in all cells, persist until the blastula stage and then are rapidly turned over. A small fraction of the embryonic SpSHR2 protein is maternal, but the majority of this nuclear receptor in the embryo is the product of new synthesis, presumably from the maternal RNA after fertilization. In agreement with the mRNA distribution, the SpSHR2 protein is also detected in all embryonic cells. Contrary to the RNA though, the SpSHR2 protein persists throughout embryonic development to the pluteus stage, long after the mRNA is depleted. Following fertilization and as soon as the 2-cell stage, the cytoplasmic SpSHR2 protein enters rapidly into the embryonic nuclei where it appears in the form of speckles. During subsequent stages (from fourth cleavage onward), SpSHR2 resides in speckled form in both the nucleus and the cytoplasm of the embryonic cells. The cytoplasmic localization of SpSHR2 differs between polarized and non-polarized cells, maintaining an apical position in the ectoderm and endoderm versus a uniform distribution in mesenchyme cells. Following the end of embryonic development (pluteus stage), the SpSHR2 protein is depleted from all tissues. During the ensuing four weeks of larval development, the SpSHR2 is not detected in either the larval or the rudiment cells which will give rise to the adult. Just prior to metamorphosis, at about 35 days post-fertilization, the protein is detected again but in contrast to the uniform distribution in the early embryo, the larval SpSHR2 is specifically expressed in cells of the mouth epithelium and the epaulettes. In adult ovaries and testes, SpSHR2 is specifically detected in the myoepithelial cells surrounding the ovarioles and the testicular acini. Nuclear SpSHR2 in blastula extracts binds to the C1R hormone response element in the upstream promoter region of the CyIIIb actin gene indicating that the latter may be a target of this nuclear receptor in the sea urchin embryo

Myosin Binding Protein-C Slow: An Intricate Subfamily of Proteins

Myosin binding protein C (MyBP-C) consists of a family of thick filament associated proteins. Three isoforms of MyBP-C exist in striated muscles: cardiac, slow skeletal, and fast skeletal. To date, most studies have focused on the cardiac form, due to its direct involvement in the development of hypertrophic cardiomyopathy. Here we focus on the slow skeletal form, discuss past and current literature, and present evidence to support that: (i) MyBP-C slow comprises a subfamily of four proteins, resulting from complex alternative shuffling of the single MyBP-C slow gene, (ii) the four MyBP-C slow isoforms are expressed in variable amounts in different skeletal muscles, (iii) at least one MyBP-C slow isoform is preferentially found at the periphery of M-bands and (iv) the MyBP-C slow subfamily may play important roles in the assembly and stabilization of sarcomeric M- and A-bands and regulate the contractile properties of the actomyosin filaments

Myosin Binding Protein-C: A Regulator of Actomyosin Interaction in Striated Muscle

Myosin-Binding protein-C (MyBP-C) is a family of accessory proteins of striated muscles that contributes to the assembly and stabilization of thick filaments, and regulates the formation of actomyosin cross-bridges, via direct interactions with both thick myosin and thin actin filaments. Three distinct MyBP-C isoforms have been characterized; cardiac, slow skeletal, and fast skeletal. Numerous mutations in the gene for cardiac MyBP-C (cMyBP-C) have been associated with familial hypertrophic cardiomyopathy (FHC) and have led to increased interest in the regulation and roles of the cardiac isoform. This review will summarize our current knowledge on MyBP-C and its role in modulating contractility, focusing on its interactions with both myosin and actin filaments in cardiac and skeletal muscles

Myosin binding protein-C slow phosphorylation is altered in Duchenne dystrophy and arthrogryposis myopathy in fast-twitch skeletal muscles

Myosin Binding Protein-C slow (sMyBP-C), encoded by MYBPC1, comprises a family of regulatory proteins of skeletal muscles that are phosphorylated by PKA and PKC. MYBPC1 missense mutations are linked to the development of Distal Arthrogryposis-1 (DA-1). Although structure-function details for this myopathy are evolving, function is undoubtedly driven by sequence variations and post-translational modifications in sMyBP-C. Herein, we examined the phosphorylation profile of sMyBP-C in mouse and human fast-twitch skeletal muscles. We used Flexor Digitorum Brevis (FDB) isolated from young (~2-months old) and old (~14-months old) wild type and mdx mice, and human Abductor Hallucis (AH) and gastrocnemious muscles carrying the DA-1 mutations. Our results indicate both constitutive and differential phosphorylation of sMyBP-C in aged and diseased muscles. We report a 7–35% reduction in the phosphorylation levels of select sites in old wild type and young or old mdx FDB mouse muscles, compared to young wild type tissue. Similarly, we observe a 30–70% decrease in the phosphorylation levels of all PKA and PKC phospho-sites in the DA-1 AH, but not gastrocnemius, muscle. Overall, our studies show that the phosphorylation pattern of sMyBP-C is differentially regulated in response to age and disease, suggesting that phosphorylation plays important roles in these processes

Alterations in cytoskeletal and Ca2+ cycling regulators in atria lacking the obscurin Ig58/59 module

IntroductionObscurin (720–870 kDa) is a giant cytoskeletal and signaling protein that possesses both structural and regulatory functions in striated muscles. Immunoglobulin domains 58/59 (Ig58/59) of obscurin bind to a diverse set of proteins that are essential for the proper structure and function of the heart, including giant titin, novex-3, and phospholamban (PLN). Importantly, the pathophysiological significance of the Ig58/59 module has been further underscored by the discovery of several mutations within Ig58/59 that are linked to various forms of myopathy in humans. We previously generated a constitutive deletion mouse model, Obscn-ΔIg58/59, that expresses obscurin lacking Ig58/59, and characterized the effects of this deletion on cardiac morphology and function through aging. Our findings demonstrated that Obscn-ΔIg58/59 male animals develop severe arrhythmia, primarily manifesting as episodes of junctional escape and spontaneous loss of regular p-waves, reminiscent of human atrial fibrillation, accompanied by significant atrial enlargement that progresses in severity with aging.Methods and ResultsTo comprehensively characterize the molecular alterations responsible for these pathologies, we performed proteomic and phospho-proteomic analyses in aging Obscn-ΔIg58/59 atria. Our studies revealed extensive and novel alterations in the expression and phosphorylation profile of major cytoskeletal proteins, Ca2+ regulators, and Z-disk associated protein complexes in the Obscn-ΔIg58/59 atria through aging.DiscussionThese studies implicate obscurin, particularly the Ig58/59 module, as an essential regulator of the Z-disk associated cytoskeleton and Ca2+ cycling in the atria and provide new molecular insights into the development of atrial fibrillation and remodeling