Cathepsin B, D and H activities in muscles of chicks of fast and slow growing strains: effect of age and diet

1. Activities of cathepsins B, D and H were measured in leg and breast muscles of fast growing (broiler) and slow growing (layer) chicks at eight time intervals between 1 and 29 days of age. 2. These enzyme activities were also measured in muscles from fast and slow growing chicks given a low protein (125 g/kg crude protein) diet between the ages of 17 and 24 days. 3. Activities of none of these cathepsins differed greatly between muscle type or strain of chick. However in both strains of chick cathepsin D and H in muscles significantly decreased with increasing age (muscle size) of the chick. Cathepsin D activity also increased when muscle proteolytic rates were increased by feeding a low protein diet. This latter effect was significant only in the muscles of fast growing chicks. 4. The results suggest that lysosomal proteases are not responsible for the differences in muscle protein degradation and growth between fast and slow growing strains of chicks, or between muscle types in the chick.     

Comparative study of the DNA-binding HU-type proteins from slow growing and fast growing strains of Rhizobiaceae

The DNA-binding HU-type proteins have been isolated from two very different strains of Rhizobiaceae: Agrobacterium tumefaciens and Rhizobium japonicum. These proteins have been called HAt and HRj respectively. Their electrophoretic mobility on polyacrylamide gel, amino acid composition and crossed immunoreactivity have been compared to that of the homologous protein isolated from Rhizobium meliloti: the protein HRm. The proteins HAt and HRm show close similarities whereas the protein HRj differs markedly from the two others. The physico-chemical characteristics of the HU-type proteins from these Rhizobiaceae are in good agreement with the respective position of these bacteria in the taxonomy.     

Muscle LIM protein: expressed in slow muscle and induced in fast muscle by enhanced contractile activity

To identify earlychanges in gene expression during the fast-to-slow transition inducedby chronic low-frequency stimulation, total RNA wasextracted from 12-h-stimulated tibialis anterior (TA) muscles of ratsand amplified by differential display RT-PCR. Among the signals ofdifferentially expressed mRNAs, a cDNA ~300 bp in length, which wasalmost undetectable in control TA muscles but prominent in stimulatedTA and normal soleus muscles, was identified. This cDNA was cloned andidentified as corresponding to the mRNA of the muscle LIM protein(MLP). Its differential expression in control, stimulated TA, andsoleus muscles was verified by Northern blotting. Antibodies againstMLP were used to identify by immunoblot analysis a protein of 22 kDa,the predicted molecular mass of MLP. Immunohistochemistry revealedstrong reactivity for MLP in all fibers of normal soleus muscle andfaint staining of some type IIA and type I fibers in control TA muscle.These fibers increased in number and staining intensity in4-day-stimulated TA muscle. MLP thus seems to play an essential roleduring the rearrangement of cytoskeletal and/or myofibrillar structuresin transforming adult muscle fibers.     

Potential phasic and tonic muscles express a common set of fast and slow myosin light chains and fast tropomyosin during early development of chick embryo

We investigated the expression of myosin light chains and tropomyosin subunits during chick embryonic development of the anterior (ALD) and posterior (PLD) parts of the latissimus dorsi muscles. As early as day 8 in ovo, both muscles accumulate a common set of myosin light chains (LC) in similar ratios (LC1F: 55 per cent; LC2S: 25 per cent; LC2F: 12 per cent; LC1S: 8 per cent) and a common set of tropomyosin (TM) subunits (beta 2, beta 1, alpha 2F). Later during development, the slow components of the LC regularly disappear in the PLD and the fast components of the LC and the alpha 2FTM disappear in the ALD, so that the adult pattern is almost established at the time of hatching. Thus, early in development, the two muscles accumulate a common set of fast and slow myosin light chains and fast tropomyosin and some isoforms are repressed at a later stage during development. These data might suggest that during development, the regulatory mechanisms of muscle specific isoform expression differ from one contractile protein to another.     

Potential phasic and tonic muscles express a common set of fast and slow myosin light chains and fast tropomyosin during early development of chick embryo

We investigated the expression of myosin light chains and tropomyosin subunits during chick embryonic development of the anterior (ALD) and posterior (PLD) parts of the latissimus dorsi muscles. As early as day 8 in ovo, both muscles accumulate a common set of myosin light chains (LC) in similar ratios (LC1F : 55 per cent; LC2S : 25 per cent; LC2F : 12 per cent ; LC1S : 8 per cent) and a common set of tropomyosin (TM) subunits (β², β¹, α²_F).Later during development, the slow components of the LC regularly disappear in the PLD and the fast components of the LC and the α²_FTM disappear in the ALD, so that the adult pattern is almost established at the time of hatching.Thus, early in development, the two muscles accumulate a common set of fast and slow myosin light chains and fast tropomyosin and some isoforms are repressed at a later stage during development. These data might suggest that during development, the regulatory mechanisms of muscle specific isoform expression differ from one contractile protein to another.     

Role of cAMP-dependent protein kinase during growth and early development of Dictyostelium discoideum

cAMP-dependent protein kinase (PKA) is an essential regulator of gene expression and cell differentiation during multicellular development of Dictyostelium discoideum. Here we show that PKA activity also regulates gene expression during the growth phase and at the transition from growth to development. Overexpression of PKA leads to overexpression of the discoidinIgamma promoter, while expression of the discoidinIgamma promoter is reduced when PKA activity is reduced, either by expression of a dominant negative mutant of the regulatory subunit or by disruption of the gene for the catalytic subunit (PKA-C). The discoidin phenotype of PKA-C null cells is cell autonomous. In particular, normal secretion of discoidin-inducing factors was demonstrated. In addition, PKA-C null cells are able to respond to media conditioned by PSF and CMF. We conclude that PKA is a major activator of discoidin expression. However, it is not required for production or transduction of the inducing extracellular signals. Therefore, PKA-dependent and PKA-independent pathways regulate the expression of the discoidin genes.     

Evolution of slow and fast development in predatory ladybirds

The frequency distribution of the durations of development of 516 larvae of Adalia bipunctata is unimodal, and the fast‐ and slow‐developing larvae can be identified at the beginning of the fourth (=last) instar. To determine the advantages of fast and slow development, the survival, duration of development, growth and number of aphids consumed by fast‐ and slow‐developing fourth instar larvae fed different numbers aphids were recorded. The percentages of fast‐ and slow‐developing fourth instar larvae that survived when fed 0.5, 1 or an excess of aphids per day, surprisingly, did not differ. The slow‐developing larvae of both sexes took longer to complete their development than the fast‐developing larvae when fed 1 or an excess of aphids per day, and although the weights of the fast‐ and slow‐developing fourth instar larvae differed at the beginning of the instar, they did not differ at the end of this instar when fed 1 aphid per day. However, when reared on an excess of aphids per day, the adult weights of the fast‐developing individuals was greater than that of slow‐developing individuals. The average durations for which the larvae in the two groups survived when fed 0.5 aphids/day differed with the larvae of the fast‐developing individuals surviving for 9.8 ± 0.5 days and slow‐developing individuals 17 ± 1.3 days. Assuming that it is the rate of predator biomass increase, which is maximized by evolution, a model of the relationship between the rate of development/growth of a predator and that of its prey indicates that the optimum growth rate of a predator is positively associated with that of its prey. The evolutionary implications of these results are discussed.     

The role of targeted protein degradation in early neural development

As neural stem cells differentiate into neurons during neurogenesis, the proteome of the cells is restructured by de novo expression and selective removal of regulatory proteins. The control of neurogenesis at the level of gene regulation is well documented and the regulation of protein abundance through protein degradation via the Ubiquitin/26S proteasome pathway is a rapidly developing field. This review describes our current understanding of the role of the proteasome pathway in neurogenesis. Collectively, the studies show that targeted protein degradation is an important regulatory mechanism in the generation of new neurons. genesis 52:287–299, 2014. © 2014 Wiley Periodicals, Inc.     

Muscle development in larvae of a fast growing tropical freshwater fish, the curimatã-pacú

The distribution and ultrastructure of myotomal muscle fibres was studied in larvae and early juveniles of the curimatã-pacú Prochilodus marggravii , a tropical freshwater fish endemic to the São Francisco River system, Brazil. At 26°C, larvae hatched 15 h post-fertilization at a relatively early stage of development with the head still curved around the yolk-sac (head-trunk angle greater than 45°), and prior to pigmentation of the eyes and formation of the jaws, gut and pectoral fins. Although motile the swimming muscles of newly-hatched larvae were largely undifferentiated. The myotomes were made up of a single layer of superficial muscle fibres containing six to eight myofibrils and abundant mitochondria, surrounding an inner core of myoblasts, myotubes and immature muscle fibres. The volume densities of mitochondria and myofibrils in the immature inner muscle fibres of 1-day-old lavae were 14.5 and 6.4% respectively. The body axis straightened within 24 h of hatching and the yolk sac was completely absorbed by 72 h. Larval development was rapid with gill filaments, a muscular stomach, liver and swimbladder present after 7 days. The inner muscle fibres were well differentiated in 7-day-old larvae; the volume density of myofibrils had increased to 63.1% whereas the volume density of mitochondria had decreased to 3.5%. In 14-day-old juveniles the superficial muscle had thickened to a layer two to three fibres thick in the region of the lateral line nerve and capillaries were present in the inner muscle. Muscle growth until 14 days was largely due to the hypertrophy of the fibres present at hatching.     

Thyroidal and neural control of myosin transitions during development of rat fast and slow muscles

The uptake of 125I-labeled epidermal growth factor (125I-EGF) by mouse pancreatic acini was inhibited (40-50%) by the secretagogue cholecystokinin octapeptide (CCK8). Analysis of competitive binding data showed that the apparent Kd of EGF binding increased 135% while the binding capacity was only slightly altered (30% increase). That the effect of CCK8 on acini was mediated by intracellular Ca2+ was indicated by the following: (i) Inhibition of 125I-EGF binding to acini was dose-dependent and paralleled the known abilities of CCK8, its analogs, and the cholinergic secretagogue carbachol to induce Ca2+ efflux from acini; and (ii) addition of the Ca2+ ionophore A23187 also inhibited 125I-EGF binding. In addition, EGF association with A431 cells was also inhibited by A23187 in the presence but not the absence of Ca2+.     

Myosin isoform transitions in regeneration of fast and slow muscles during postnatal development of the rat

Regeneration of rat fast (gastrocnemius medialis) and slow (soleus) muscles was examined after degeneration of myofibers had been achieved by injection of cardiotoxin into the hindleg during the first week after birth. Myogenesis in the regenerating muscles was compared to postnatal myogenesis in the contralateral and in control muscles. Synthesis of embryonic and neonatal myosin isoforms was initiated 3 days after injury. These forms were gradually replaced by the intermediate and fast adult isoforms (type II fiber myosins), whose synthesis followed the same curve in regenerating, contralateral, and control muscles. In contrast, synthesis of the slow myosin isoform (type I fiber myosin) was greatly delayed in injured muscles, but eventually became equal to its synthesis in contralateral and control muscles. It therefore appears that synthesis of type II fiber myosins is similarly regulated, probably by thyroid hormone, in developing regenerating and normal muscles, while synthesis of type I fiber myosin depends on other factor(s).     

Physiological mechanism governing slow and fast development in predatory ladybirds

Aphidophagous and coccidophagous ladybirds, similar to their prey, show marked differences in their pace of life (Dixon, 2000), in particular in their rate of development, with all stages of aphidophagous species developing much faster than those of coccidophagous species. Two hypotheses are proposed to account for the large difference in the pace of life of these two groups. These are that differences in the rate of development are a result of differences in lower temperature thresholds for development or the quality of their respective prey as food (Dixon et al., 2011). Analysis of published results on the rates of development of the eggs of ladybirds indicates that the inverse relationships between the number of day‐degrees required for development (K) and the lower temperature threshold for development (td_min) of these two groups are significantly different. In particular, the respective td_min overlap and K of the aphidophagous and coccidophagous species with a similar td_min are, on average, 38 and 117 day‐degrees (D^o). The relationship between the rate of development (R) and temperature (T) for aphids reared on poor‐ or high‐quality foods indicates that, although the value of td_min of a species depends on food quality, K does not, showing that it is unlikely that K is governed by food quality. Thus, there is little support for differences in either the td_min or food quality governing the difference in the pace of life of these two groups of ladybirds. The results indicate that the physiological mechanism that may govern the difference in the pace of life between these two groups is the number of day‐degrees (K) needed to complete their development. The possible evolutionary reason for this is discussed.     

Temporal expression of TnI fast and slow isoforms in biceps femoris and masseter muscle during pig growth

Biceps femoris (BF) and masseter muscle (MM) are the mixture of slow oxidative and fast-twitch fibres. Compared with MM, BF had the significantly higher expression of myosin heavy chain (MyHC) fast IIx and IIb isoforms (MyHCIIx and MyHCIIb), but lower expression of MyHC slow isoform (MyHCI) and fast IIa isoform (MyHCIIa). The objective of this study was to investigate the expression pattern of troponin I (TnI) slow-twitch isoform (TNNI1) and fast-twitch isoform (TNNI2) in BF and MM of Yorkshire and Meishan pigs which differed significantly in the growth rate. The expression of the TNNI1 and TNNI2 peaked at the postnatal 35 days in Yorkshire pigs and postnatal 60 days in Meishan pigs. The expression of TNNI1 and TNNI2 in Meishan pigs was significantly higher than that in Yorkshire pigs at the foetal 60 days, while the opposite occurred at postnatal 35 days. The expression ratio of TNNI1 relative to TNNI2 favoured TNNI2 expression in BF and MM regardless of Yorkshire and Meishan pigs. TNNI1 expression in MM was significantly higher than that in BF at 60, 120 and 180 days in Meishan pigs and at 120 and 180 days in Yorkshire pigs. On the contrary, no significant difference of TNNI2 expression in BF and MM was found except for Yorkshire pigs of 180 days. This study provided the foundation for future research on TnI isoforms as the model gene to study mechanisms of muscle fibre-specific gene regulation in pigs.     

The role of slow and fast protein motions in allosteric interactions

Allostery is fundamentally thermodynamic in nature. Long-range communication in proteins may be mediated not only by changes in the mean conformation with enthalpic contribution but also by changes in dynamic fluctuations with entropic contribution. The important role of protein motions in mediating allosteric interactions has been established by NMR spectroscopy. By using CAP as a model system, we have shown how changes in protein structure and internal dynamics can allosterically regulate protein function and activity. The results indicate that changes in conformational entropy can give rise to binding enhancement, binding inhibition, or have no effect in the expected affinity, depending on the magnitude and sign of enthalpy–entropy compensation. Moreover, allosteric interactions can be regulated by the modulation a low-populated conformation states that serve as on-pathway intermediates for ligand binding. Taken together, the interplay between fast internal motions, which are intimately related to conformational entropy, and slow internal motions, which are related to poorly populated conformational states, can regulate protein activity in a way that cannot be predicted on the basis of the protein’s ground-state structure.