Nonsynonymous substitution rate (Ka) is a relatively consistent parameter for defining fast-evolving and slow-evolving protein-coding genes

Background  

Mammalian genome sequence data are being acquired in large quantities and at enormous speeds. We now have a tremendous opportunity to better understand which genes are the most variable or conserved, and what their particular functions and evolutionary dynamics are, through comparative genomics.     

Accumulation of β-catenin by lithium chloride in porcine myoblast cultures accelerates cell differentiation

The Wnt/β-catenin signaling pathway regulates cell proliferation and differentiation to determine cell fate during embryogenesis. Lithium chloride (LiCl) is known to activate canonical Wnt signaling by inhibiting glycogen synthetase kinase-3β and consequently stabilizing free cytosolic β-catenin. To understand the role of the Wnt/β-catenin pathway in the regulation of porcine myoblast differentiation, we studied the effects of LiCl on cultured porcine myoblasts and β-catenin expression. A supplementation of 25 mM LiCl induced myoblast differentiation into myotubes over 3 days of culture. By semi-quantitative RT-PCR analyses, levels of mRNA encoding MyoD, Myogenin, Myf5 and several Wnt-responsive genes in the cultured myoblast cells were significantly increased after LiCl treatment. Using Western blotting and immunofluorescence analysis, we found that the protein levels of β-catenin were consistently increased by LiCl. Meanwhile, phosphorylated GSK-3β at Ser9 levels were also increased as an indicator of GSK-3β inactivation. Additionally, the nuclear staining of endogenous β-catenin was also significantly increased in porcine myoblasts 48 h after LiCl treatment. These results provided additional evidence that Wnt/β-catenin is a significant pathway that regulates myogenic differentiation. An enhanced level of β-catenin plays a positive role in porcine myoblast differentiation.     

Molecular characterization of glycogen synthase 1 and its tissue expression profile with type II hexokinase and muscle-type phosphofructokinase in horses

Muscle glycogen synthase (GYS1) is the rate-limiting enzyme in glycogen synthesis, and its activity is regulated by the phosphorylation states of certain amino acid residues encoded by the GYS1 gene. In the present study, the authors molecularly characterized the full-length equine GYS1 (eGYS1) cDNA and found that it contains a less common polyadenylation signal (AATACA). An amino acid alignment with other mammalian GYS1 showed that the phosphorylation sites in eGYS1 are completely conserved. Genomic DNA analysis revealed that the equine-specific substitutions (Glu 16 Asp and Ala 252 Thr) were completely conserved among six equine species. The tissue expression profiles of eGYS1, equine type II hexokinase (eHKII) and muscle-type phosphofructokinase (ePFKM) were determined by real-time PCR and western blot analysis. The mRNA expression level of eGYS1 was significantly higher in the cervical muscle as compared to other tissues. The cervical muscle and heart tissue samples contained a broad range of eGYS1 protein bands that appeared to reflect multiple phosphorylation states. eHKII was predominately expressed only in the cervical muscle; unlike its expression in other mammals, eHKII was not substantially expressed in the insulin-responsive heart or adipose tissue of horse. The expression level of ePFKM mRNA was significantly higher in the heart than in the cervical muscle, which differs from the PFKM expression pattern of other mammals. These tissue expression profiles are fundamental for the understanding of equine glucose metabolism.     

Optimisation of plantlet regeneration from leaf and nodal derived callus of <Emphasis Type="Italic">Vanilla planifolia</Emphasis> Andrews

An indirect in vitro plant regeneration protocol for Vanilla planifolia has been established. Juvenile leaf and nodal segments from V. planifolia were used as explants to initiate callus. Nodal explants showed better callus initiation than juvenile leaf explants, with 35.0% of explants forming callus when cultured on Murashige and Skoog (MS) basal medium supplemented with 2.0 mg/l 1-naphthylacetic acid (NAA) and 1.0 mg/l 6-benzyladenine (BA). Almost 10.0% of juvenile leaf explants were induced to form callus on the MS basal medium containing 2.0 mg/l NAA and 2.0 mg/l BA, whereas no callus formed in the presence of any concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) and BA. After 8 weeks, callus generated was transferred to MS basal medium containing 1.0 mg/l BA and 0.5 mg/l NAA. A mean number of 4.2 shoots per callus was produced on this medium, with a mean length of 3.8 cm after 8 weeks of culture. Roots formed on 88.3% of plantlets when they were cultured on MS medium supplemented with 1.0 mg/l NAA, with a mean length of 4.4 cm after 4 weeks of culture. Of the rooted plantlets, 90.0% survived acclimatisation and were making new growth after 4 weeks.     

Molecular variation and evolution of the tyrosine kinase domains of insulin receptor <Emphasis Type="Italic">IRa</Emphasis> and <Emphasis Type="Italic">IRb</Emphasis> genes in Cyprinidae

The insulin receptor (IR) gene plays an important role in regulating cell growth, differentiation and development. In the present study, DNA sequences of insulin receptor genes, IRa and IRb, were amplified and sequenced from 37 representative species of the Cyprinidae and from five outgroup species from non-cyprinid Cypriniformes. Based on coding sequences (CDS) of tyrosine kinase regions of IRa and IRb, molecular evolution and phylogenetic relationships were analyzed to better understand the characteristics of IR gene divergence in the family Cyprinidae. IRa and IRb were clustered into one lineage in the gene tree of the IR gene family, reconstructed using the unweighted pair group method with arithmetic mean (UPGMA). IRa and IRb have evolved into distinct genes after IR gene duplication in Cyprinidae. For each gene, molecular evolution analyses showed that there was no significant difference among different groups in the reconstructed maximum parsimony (MP) tree of Cyprinidae; IRa and IRb have been subjected to similar evolutionary pressure among different lineages. Although the amino acid sequences of IRa and IRb tyrosine kinase regions were highly conserved, our analyses showed that there were clear sequence variations between the tyrosine kinase regions of IRa and IRb proteins. This indicates that IRa and IRb proteins might play different roles in the insulin signaling pathway.     

MiR-204 regulates cardiomyocyte autophagy induced by ischemia-reperfusion through LC3-II

Background  

Autophagy plays a significant role in myocardial ischemia-reperfusion (IR) injury. So it is important to inhibit autophagy to protect cardiomyocytes besides anti-apoptosis. MiRNA has been demonstrated to protect cardiomyocytes against apoptosis during IR, while whether it has anti-autophagy effect has not been known. The aim of this study was to investigate whether miR-204 regulated autophagy by regulating LC3-II protein, which is the marker of autophagosome during myocardial IR injury.     

The impact of founder events and introductions on genetic variation in the muskox <Emphasis Type="Italic">Ovibos moschatus</Emphasis> in Sweden

The muskox Ovibos moschatus (Zimmerman 1780) is a specialised arctic mammal with a highly fragmented circumpolar distribution, with native populations in Canada and east Greenland and introduced populations in west Greenland, Alaska, Siberia and Eurasia. In 1971, five O. moschatus individuals from an introduced population in Norway migrated to Sweden. After a peak population of 36 individuals in the mid-1980s, the Swedish population now numbers seven individuals, making it vulnerable to both demographic and genetic stochasticity (i.e. inbreeding). Here, we analyse genetic variation among native and introduced populations of O. moschatus to evaluate the genetic effect of sequential founder events in this species. Our results show that genetic variation among native and introduced O. moschatus populations do not conform entirely to the expectations from sequential founder events, most likely because of random processes associated with introduction. In the Swedish population, a calf resulting from the mating of a wild cow and a captive Greenlandic bull contributes significantly to the current genetic variation. Thus, even a single outbreeding event may, at least momentarily, increase the genetic variation and potentially prevent inbreeding depression. Our results should aid the long-term preservation of O. moschatus in Sweden and Europe.     

Methyl jasmonate counteracts boron toxicity by preventing oxidative stress and regulating antioxidant enzyme activities and artemisinin biosynthesis in <Emphasis Type="Italic">Artemisia annua</Emphasis> L.

Boron is an essential plant micronutrient, but it is phytotoxic if present in excessive amounts in soil for certain plants such as Artemisia annua L. that contains artemisinin (an important antimalarial drug) in its areal parts. Artemisinin is a sesquiterpene lactone with an endoperoxide bridge. It is quite expensive compound because the only commercial source available is A. annua and the compound present in the plant is in very low concentration. Since A. annua is a major source of the antimalarial drug and B stress is a deadly threat to its cultivation, the present research was conducted to determine whether the exogenous application of methyl jasmonate (MeJA) could combat the ill effects of excessive B present in the soil. According to the results obtained, the B toxicity induced oxidative stress and reduced the stem height as well as fresh and dry masses of the plant remarkably. The excessive amounts of soil B also lowered the net photosynthetic rate, stomatal conductance, internal CO₂ concentration and total chlorophyll content in the leaves. In contrast, the foliar application of MeJA enhanced the growth and photosynthetic efficiency both in the stressed and non-stressed plants. The excessive B levels also increased the activities of antioxidant enzymes, such as catalase, peroxidase and superoxide dismutase. Endogenous H₂O₂ and O₂⁻ levels were also high in the stressed plants. However, the MeJA application to the stressed plants reduced the amount of lipid peroxidation and stimulated the synthesis of antioxidant enzymes, enhancing the content and yield of artemisinin as well. Thus, it was concluded that MeJA might be utilized in mitigating the B toxicity and improving the content and yield of artemisinin in A. annua plant.     

Distinct behavior of claudin-3 and -4 around lactation period in mammary alveolus in mice

Tight junctions (TJs) are the most apical junctional complexes and restrict the fluid flux through the paracellular pathway. In the mammary glands, the tightness of TJs occurs shortly after parturition to prevent the leakage of milk components from the lumen and the loosening of TJs is induced immediately after weaning. Claudins are transmembrane proteins, and their composition at the apical-most regions determines the permeability of TJs. In this study, we investigated the localization and expression patterns of claudin-3 and -4 in the mammary glands around the lactation period because it is unclear how claudins construct mammary TJs in the apical-most regions. Our results showed that claudin-3 and -4 change not only their level of expression but also their localization in the processes of parturition, lactation, and weaning. Claudin-3 was concentrated in the apical-most regions during lactation, whereas claudin-4 gradually decreased at the beginning of lactation and increased drastically immediately after weaning. The qualitative change of claudin-3 was also identified by western blotting analysis as an additional band around the lactation period. In addition, parts of the mammary epithelial cells showed intensive positive reactions to claudin-4 in the lateral membrane and cytoplasm after weaning, concurrently with the involution mammary glands. These results indicate that claudin-3 in the apical-most regions maintains the impermeable TJs during lactation, and claudin-4 contributes to the permeability changes of TJs immediately after parturition and weaning.