RESUMO
Transcription elongation requires elaborate coordination between the transcriptional machinery and chromatin regulatory factors to successfully produce RNA while preserving the epigenetic landscape. Recent structural and genomic studies have highlighted that suppressor of Ty 6 (Spt6), a conserved histone chaperone and transcription elongation factor, sits at the crux of the transcription elongation process. Other recent studies have revealed that Spt6 also promotes DNA replication and genome integrity. Here, we review recent studies of Spt6 that have provided new insights into the mechanisms by which Spt6 controls transcription and have revealed the breadth of Spt6 functions in eukaryotic cells.
Assuntos
Histonas , Humanos , Replicação do DNA/genética , Instabilidade Genômica/genética , Chaperonas de Histonas/genética , Chaperonas de Histonas/química , Chaperonas de Histonas/metabolismo , Histonas/genética , Histonas/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Fatores de Transcrição/genética , Transcrição Gênica , Fatores de Elongação da Transcrição/genética , Fatores de Elongação da Transcrição/química , Fatores de Elongação da Transcrição/metabolismo , AnimaisRESUMO
Membrane repair is essential to cell survival. In skeletal muscle, injury often associates with plasma membrane disruption. Additionally, muscular dystrophy is linked to mutations in genes that produce fragile membranes or reduce membrane repair. Methods to enhance repair and reduce susceptibility to injury could benefit muscle in both acute and chronic injury settings. Annexins are a family of membrane-associated Ca2+-binding proteins implicated in repair, and annexin A6 was previously identified as a genetic modifier of muscle injury and disease. Annexin A6 forms the repair cap over the site of membrane disruption. To elucidate how annexins facilitate repair, we visualized annexin cap formation during injury. We found that annexin cap size positively correlated with increasing Ca2+ concentrations. We also found that annexin overexpression promoted external blebs enriched in Ca2+ and correlated with a reduction of intracellular Ca2+ at the injury site. Annexin A6 overexpression reduced membrane injury, consistent with enhanced repair. Treatment with recombinant annexin A6 protected against acute muscle injury in vitro and in vivo. Moreover, administration of recombinant annexin A6 in a model of muscular dystrophy reduced serum creatinine kinase, a biomarker of disease. These data identify annexins as mediators of membrane-associated Ca2+ release during membrane repair and annexin A6 as a therapeutic target to enhance membrane repair capacity.
Assuntos
Anexina A6/farmacologia , Cálcio/metabolismo , Membrana Celular/metabolismo , Músculo Esquelético/lesões , Distrofia Muscular Animal/prevenção & controle , Animais , Anexina A6/genética , Membrana Celular/patologia , Feminino , Masculino , Camundongos , Camundongos Knockout , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Distrofia Muscular Animal/genética , Distrofia Muscular Animal/metabolismo , Distrofia Muscular Animal/patologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacologiaRESUMO
The adult myocardium relies on oxidative metabolism. In ischemic myocardium, such as the embryonic heart, glycolysis contributes more prominently as a fuel source. The sulfonylurea receptor 2 (SUR2) was previously implicated in the normal myocardial transition from glycolytic to oxidative metabolism that occurs during adaptation to postnatal life. This receptor was now selectively deleted in adult mouse myocardium resulting in protection from ischemia reperfusion injury. SUR2-deleted cardiomyocytes had enhanced glucose uptake, and SUR2 forms a complex with the major glucose transporter. These data identify the SUR2 receptor as a target to shift cardiac metabolism to protect against myocardial injury.
RESUMO
Genetic disruption of the dystrophin complex produces muscular dystrophy characterized by a fragile muscle plasma membrane leading to excessive muscle degeneration. Two genetic modifiers of Duchenne Muscular Dystrophy implicate the transforming growth factor ß (TGFß) pathway, osteopontin encoded by the SPP1 gene and latent TGFß binding protein 4 (LTBP4). We now evaluated the functional effect of these modifiers in the context of muscle injury and repair to elucidate their mechanisms of action. We found that excess osteopontin exacerbated sarcolemmal injury, and correspondingly, that loss of osteopontin reduced injury extent both in isolated myofibers and in muscle in vivo. We found that ablation of osteopontin was associated with reduced expression of TGFß and TGFß-associated pathways. We identified that increased TGFß resulted in reduced expression of Anxa1 and Anxa6, genes encoding key components of the muscle sarcolemma resealing process. Genetic manipulation of Ltbp4 in dystrophic muscle also directly modulated sarcolemmal resealing, and Ltbp4 alleles acted in concert with Anxa6, a distinct modifier of muscular dystrophy. These data provide a model in which a feed forward loop of TGFß and osteopontin directly impacts the capacity of muscle to recover from injury, and identifies an intersection of genetic modifiers on muscular dystrophy.
Assuntos
Genes Modificadores , Proteínas de Ligação a TGF-beta Latente/fisiologia , Músculo Esquelético/fisiologia , Distrofia Muscular Animal/genética , Osteopontina/metabolismo , Animais , Anexina A1/genética , Anexina A1/metabolismo , Anexina A6/genética , Anexina A6/metabolismo , Feminino , Regulação da Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos DBA , Camundongos Knockout , Músculo Esquelético/lesões , Distrofia Muscular Animal/metabolismo , Distrofia Muscular Animal/patologia , Osteopontina/genética , Receptores de Fatores de Crescimento Transformadores beta/genética , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Recuperação de Função Fisiológica , Sarcolema/fisiologiaRESUMO
The muscular dystrophies are genetically diverse. Shared pathological features among muscular dystrophies include breakdown, or loss of muscle, and accompanying fibrotic replacement. Novel strategies are needed to enhance muscle repair and function and to slow this pathological remodeling. Glucocorticoid steroids, like prednisone, are known to delay loss of ambulation in patients with Duchenne muscular dystrophy but are accompanied by prominent adverse effects. However, less is known about the effects of steroid administration in other types of muscular dystrophies, including limb-girdle muscular dystrophies (LGMDs). LGMD 2B is caused by loss of dysferlin, a membrane repair protein, and LGMD 2C is caused by loss of the dystrophin-associated protein, γ-sarcoglycan. Herein, we assessed the efficacy of steroid dosing on sarcolemmal repair, muscle function, histopathology, and the regenerative capacity of primary muscle cells. We found that in murine models of LGMD 2B and 2C, daily prednisone dosing reduced muscle damage and fibroinflammatory infiltration. However, daily prednisone dosing also correlated with increased muscle adipogenesis and atrophic remodeling. Conversely, intermittent dosing of prednisone, provided once weekly, enhanced muscle repair and did not induce atrophy or adipogenesis, and was associated with improved muscle function. These data indicate that dosing frequency of glucocorticoid steroids affects muscle remodeling in non-Duchenne muscular dystrophies, suggesting a positive outcome associated with intermittent steroid dosing in LGMD 2B and 2C muscle.
Assuntos
Glucocorticoides/farmacologia , Músculo Esquelético/efeitos dos fármacos , Distrofia Muscular do Cíngulo dos Membros/tratamento farmacológico , Animais , Distrofina/efeitos dos fármacos , Distrofina/metabolismo , Glucocorticoides/administração & dosagem , Proteínas de Membrana/metabolismo , Camundongos , Músculo Esquelético/patologia , Distrofia Muscular do Cíngulo dos Membros/patologia , Distrofia Muscular de Duchenne/tratamento farmacológico , Distrofia Muscular de Duchenne/metabolismo , Prednisona/administração & dosagem , Prednisona/farmacologia , Sarcoglicanas/efeitos dos fármacos , Sarcoglicanas/metabolismoRESUMO
Glucocorticoid steroids such as prednisone are prescribed for chronic muscle conditions such as Duchenne muscular dystrophy, where their use is associated with prolonged ambulation. The positive effects of chronic steroid treatment in muscular dystrophy are paradoxical because these steroids are also known to trigger muscle atrophy. Chronic steroid use usually involves once-daily dosing, although weekly dosing in children has been suggested for its reduced side effects on behavior. In this work, we tested steroid dosing in mice and found that a single pulse of glucocorticoid steroids improved sarcolemmal repair through increased expression of annexins A1 and A6, which mediate myofiber repair. This increased expression was dependent on glucocorticoid response elements upstream of annexins and was reinforced by the expression of forkhead box O1 (FOXO1). We compared weekly versus daily steroid treatment in mouse models of acute muscle injury and in muscular dystrophy and determined that both regimens provided comparable benefits in terms of annexin gene expression and muscle repair. However, daily dosing activated atrophic pathways, including F-box protein 32 (Fbxo32), which encodes atrogin-1. Conversely, weekly steroid treatment in mdx mice improved muscle function and histopathology and concomitantly induced the ergogenic transcription factor Krüppel-like factor 15 (Klf15) while decreasing Fbxo32. These findings suggest that intermittent, rather than daily, glucocorticoid steroid regimen promotes sarcolemmal repair and muscle recovery from injury while limiting atrophic remodeling.
Assuntos
Glucocorticoides/administração & dosagem , Músculo Esquelético/efeitos dos fármacos , Prednisona/administração & dosagem , Animais , Anexina A6/genética , Anexina A6/metabolismo , Células Cultivadas , Esquema de Medicação , Avaliação Pré-Clínica de Medicamentos , Expressão Gênica , Glucocorticoides/efeitos adversos , Masculino , Camundongos da Linhagem 129 , Camundongos Endogâmicos DBA , Camundongos Endogâmicos mdx , Músculo Esquelético/fisiopatologia , Atrofia Muscular/induzido quimicamente , Distrofia Muscular de Duchenne/tratamento farmacológico , Prednisona/efeitos adversos , Ligação Proteica , Receptores de Glucocorticoides/metabolismo , Regeneração , Sarcolema/efeitos dos fármacos , Sarcolema/fisiologia , Ativação Transcricional/efeitos dos fármacosRESUMO
Dysferlin is a membrane-associated protein implicated in membrane resealing; loss of dysferlin leads to muscular dystrophy. We examined the same loss-of-function Dysf mutation in two different mouse strains, 129T2/SvEmsJ (Dysf(129)) and C57BL/6J (Dysf(B6)). Although there are many genetic differences between these two strains, we focused on polymorphisms in Anxa6 because these variants were previously associated with modifying a pathologically distinct form of muscular dystrophy and increased the production of a truncated annexin A6 protein. Dysferlin deficiency in the C57BL/6J background was associated with increased Evan's Blue dye uptake into muscle and increased serum creatine kinase compared to the 129T2/SvEmsJ background. In the C57BL/6J background, dysferlin loss was associated with enhanced pathologic severity, characterized by decreased mean fiber cross-sectional area, increased internalized nuclei, and increased fibrosis, compared to that in Dysf(129) mice. Macrophage infiltrate was also increased in Dysf(B6) muscle. High-resolution imaging of live myofibers demonstrated that fibers from Dysf(B6) mice displayed reduced translocation of full-length annexin A6 to the site of laser-induced sarcolemmal disruption compared to Dysf(129) myofibers, and impaired translocation of annexin A6 associated with impaired resealing of the sarcolemma. These results provide one mechanism by which the C57BL/6J background intensifies dysferlinopathy, giving rise to a more severe form of muscular dystrophy in the Dysf(B6) mouse model through increased membrane leak and inflammation.