RESUMEN
Mechanical properties of the jaw-closing muscles of the cat are poorly understood. These muscles are known to differ in myosin and fibre type compositions from limb muscles. This work aims to correlate mechanical properties of single fibres in cat jaw and limb muscles with their myosin subunit compositions. The stiffness minimum frequency, f(min), which reflects isometric cross-bridge kinetics, was measured in Ca(2+)-activated glycerinated fast and slow fibres from cat jaw and limb muscles for temperatures ranging between 15 and 30 degrees C by mechanical perturbation analysis. At 15 degrees C, f(min) was 0.5 Hz for limb-slow fibres, 4-6 Hz for jaw-slow fibres, and 10-13 Hz for limb-fast and jaw-fast fibres. The activation energy for f(min) obtained from the slope of the Arrhenius plot for limb-slow fibres was 30-40% higher than values for the other three types of fibres. SDS-PAGE and western blotting using highly specific antibodies verified that limb-fast fibres contained IIA or IIX myosin heavy chain (MyHC). Jaw-fast fibres expressed masticatory MyHC while both jaw-fast and jaw-slow fibres expressed masticatory myosin light chains (MLCs). The nucleotide sequences of the 3' ends of the slow MyHC cDNAs isolated from cat masseter and soleus cDNA libraries showed identical coding and 3'-untranslated regions, suggesting that jaw-slow and limb-slow fibres express the same slow MyHC gene. We conclude that the isometric cross-bridge cycling kinetics of jaw-fast and limb-fast fibres detected by f(min) are indistinguishable in spite of differences in MyHC and light chain compositions. However, jaw-slow fibres, in which the same slow MyHCs are found in combination with MLCs of the jaw type, show enhanced cross-bridge cycling kinetics and reduced activation energy for cross-bridge detachment.
Asunto(s)
Fibras Musculares de Contracción Rápida/fisiología , Fibras Musculares de Contracción Lenta/fisiología , Músculo Esquelético/fisiología , Cadenas Pesadas de Miosina/fisiología , Cadenas Ligeras de Miosina/fisiología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Gatos , Extremidades , Maxilares , Cinética , Músculo Masetero/química , Músculo Masetero/fisiología , Datos de Secuencia Molecular , Fibras Musculares de Contracción Rápida/química , Fibras Musculares de Contracción Lenta/química , Músculo Esquelético/química , Cadenas Pesadas de Miosina/química , Cadenas Pesadas de Miosina/genéticaRESUMEN
BACKGROUND/AIMS: Chronic ethanol consumption inhibits liver regeneration. We examined the effects of chronic ethanol consumption on two mitogen-activated protein kinases in relation to induction of cell cycle proteins after partial hepatectomy (PH). METHODS: Male Wistar rats were ethanol-fed (EF) or pair-fed (PF) for 16 weeks before PH. Hepatic activation of extracellular signal regulated kinase (ERK)1/2, p38 kinase and expression of cyclinD1, cyclin-dependent kinase-4 (cdk4) and proliferating cell nuclear antigen (PCNA) were studied. RESULTS: In PF rats, PH-induced p38 activation was evident at 2h and was maximal at 12h. There was a close temporal relationship between p38 activation, cyclin D1 and PCNA expression. Alcohol exposure reduced p38 activation, cyclin D1 and PCNA, each by approximately 50%. ERK1/2 activation occurred during the first 2h post-PH in both EF and PF rats, and there was no later increase in PF rats. In vivo inhibition of p38 suppressed PCNA expression whereas the effect of ERK1/2 inhibition was inconsistent. CONCLUSIONS: p38 kinase activation is linked temporally with cyclin D1 expression after PH and appears to exert cell cycle control in the adult liver. p38 signaling also appears to be a target for the inhibitory effect of chronic alcohol on liver regeneration.
Asunto(s)
Alcoholismo/enzimología , Ciclina D1/genética , Etanol/toxicidad , Hepatectomía , Hígado/enzimología , ARN Mensajero/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Alcoholismo/patología , Animales , Western Blotting , Ciclina D1/biosíntesis , Quinasa 4 Dependiente de la Ciclina/metabolismo , Modelos Animales de Enfermedad , Activación Enzimática/efectos de los fármacos , Expresión Génica , Hígado/efectos de los fármacos , Hígado/patología , Regeneración Hepática/efectos de los fármacos , Masculino , Antígeno Nuclear de Célula en Proliferación/metabolismo , ARN Mensajero/biosíntesis , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
"Superfast" or masticatory myosin is the molecular motor in the powerful and specialized jaw-closing muscles of carnivores, folivores, and frugivores. This myosin presumably underpins the unusual high force and moderate shortening velocity of muscle fibers expressing it. Here, we report the cloning and sequencing of the cDNA encoding the full-length masticatory myosin heavy chain (MyHC) from cat temporalis muscle. This was obtained by immunoscreening a cDNA expression library and RACE-PCR (rapid amplification of cDNA ends-PCR). Sequence comparisons at the DNA and amino acid levels show that masticatory MyHC has less than 70% homology to known striated MyHCs, compared with 87-96% between other mammalian fast isoforms themselves. Nucleotide substitution rates at the nonsynonymous sites between masticatory MyHC and other mammalian striated MyHCs are considerably higher than between these striated MyHCs themselves. Phylogenetic analysis revealed that masticatory MyHC diverged from invertebrate MyHC before the avian cardiac MyHC subclass and the mammalian fast/developmental and slow/cardiac MyHC subclasses. Masticatory MyHC is thus a distinct new subclass of vertebrate striated myosins. The early divergence from invertebrate MyHC, combined with immunochemical evidence of its expression in reptilian and shark jaw-closing muscles, suggests that masticatory MyHC evolved in early gnathostomes, driven by benefits derived from powerful jaw closure. During the mammalian radiation, some taxa continued to express it, while others adapted to new types of food and eating habits by replacing masticatory MyHC with more appropriate isoforms normally found in limb and cardiac muscles.