Evolution of the deep-sea gulper eel mitochondrial genomes: large-scale gene rearrangements originated within the eels

Recent studies have demonstrated that deviations from the typical vertebrate mitochondrial gene order are more frequent than initially thought. Such deviations, however, are minor, with inversions and/or translocations of a few genes being involved and tandem duplication of the gene regions followed by deletions of genes having been invoked as mechanisms originating in such novel gene order. During the course of molecular phylogenetic studies on the Elopomorpha (eels and their allies), we found that mitochondrial genomes (mitogenomes) from the two deep-sea gulper eels, Eurypharynx pelecanoides (Eurypharyngidae) and Saccopharynx lavenbergi (Saccopharyngidae), exhibit an identical gene order which greatly differs from that of any other vertebrates. Phylogenetic analysis using the mitogenomic data from 59 species of fish not only confirmed a single origin of such a gene order with confidence but also indicated that it had been derived from the typical vertebrate gene order. Detailed comparisons of the gulper eel gene order with that of typical vertebrates suggested that occurrence of a single step, large-scale duplication of gene region extending >12 kb, followed by deletions of genes in a common ancestor of the two species, most parsimoniously accounts for this unusual gene arrangement.     

Effects of serum deprivation on myofibrillar proteolysis in chick myotube cultures

We examined the effects of serum deprivation on myofibrillar proteolysis in chick myotubes. Myotubes were incubated with serum-free medium for 24 hours. N(tau)-methylhistidine release, as an index of myofibrillar proteolysis, as well as protease activities such as calpain, proteasome, and cathepsins (B+L and D) activities were increased by serum deprivation. These results indicate that serum deprivation induces calpain, proteasome, and cathepsins activities, resulting in an increase in myofibrillar proteolysis in chick myotubes.     

Morphological and functional changes in cell junctions during secretory stimulation in the perfused rat submandibular gland

To examine the influence of cholinergic and beta-adrenergic agents on paracellular transport, we applied confocal microscopy and freeze-fracture to the isolated, perfused submandibular gland of the rat. By confocal microscopy, perfusion of lucifer yellow through an arterial catheter, revealed a bright fluorescence in the basolateral spaces of acini, but not in the intercellular canaliculi. However, addition of isoproterenol on carbachol stimulation, induced lucifer yellow fluorescence in intercellular canaliculi. This finding indicates that isoproterenol is capable of opening the paracellular route. The tight junction strands surrounding intercellular canaliculi were visualized using freeze replicas. Fixation was carried out both by vascular perfusion with Karnovsky's solution and by metal contact rapid freezing with liquid helium. In the chemically-fixed specimens, the strand particles of tight junctions formed 2-5 lines at the P-face along most of the apical portion at rest. With carbachol/isoproterenol stimulation, the strand particles rearranged with free ends and terminal loops. In the rapidly frozen specimens, the strand particles were arranged more irregularly even in the resting state. The meshwork of strands became more disheveled and interrupted during carbachol/ isoproterenol stimulation. The present findings led us to conclude that: 1) the beta-adrenergic agent, isoproterenol, can open the paracellular transport. 2) in the rapidly frozen specimen, the tight junction strand particles are arranged roughly and become disheveled and interrupted during stimulation by carbachol/isoproterenol. These findings may be related to rearrangement of subcellular structures, especially of the actin filament network.     

Structure-activity relationship of benzo[b]thiophene-2-sulfonamide derivatives as novel human chymase inhibitors

We have identified a new class of chymase inhibitor through a substituent analysis of MWP00965, which we previously discovered by in silico screening. TY-51076 (7) showed high potency (IC(50)=56 nM) and excellent selectivity for chymase compared to chymotrypsin and cathepsin G (>400-fold). The synthesis and structure-activity relationship of this class are described.     

How meaningful are Bayesian support values?

In this study, we used an empirical example based on 100 mitochondrial genomes from higher teleost fishes to compare the accuracy of parsimony-based jackknife values with Bayesian support values. Phylogenetic analyses of 366 partitions, using differential taxon and character sampling from the entire data matrix of 100 taxa and 7,990 characters, were performed for both phylogenetic methods. The tree topology and branch-support values from each partition were compared with the tree inferred from all taxa and characters. Using this approach, we quantified the accuracy of the branch-support values assigned by the jackknife and Bayesian methods, with respect to each of 15 basal clades. In comparing the jackknife and Bayesian methods, we found that (1) both measures of support differ significantly from an ideal support index; (2) the jackknife underestimated support values; (3) the Bayesian method consistently overestimated support; (4) the magnitude by which Bayesian values overestimate support exceeds the magnitude by which the jackknife underestimates support; and (5) both methods performed poorly when taxon sampling was increased and character sampling was not increases. These results indicate that (1) the higher Bayesian support values are inappropriate (in magnitude), and (2) Bayesian support values should not be interpreted as probabilities that clades are correctly resolved. We advocate the continued use of the relatively conservative bootstrap and jackknife approaches to estimating branch support rather than the more extreme overestimates provided by the Markov Chain Monte Carlo-based Bayesian methods.     

Expressions of adrenomedullin mRNA and protein in rats with hypobaric hypoxia-induced pulmonary hypertension

Experimental pulmonary hypertension induced in a hypobaric hypoxic environment (HHE) is characterized by structural remodeling of the heart and pulmonary arteries. Adrenomedullin (AM) has diuretic, natriuretic, and hypotensive effects. To study the possible effects of HHE on the AM synthesis system, 150 male Wistar rats were housed in a chamber at the equivalent of a 5,500-m altitude level for 21 days. After 14 days of exposure to HHE, pulmonary arterial pressure (PAP) was significantly increased (compared with control rats). The plasma AM protein level was significantly increased on day 21 of exposure to HHE. In the right ventricle (RV), right atrium, and left atrium of the heart, the expressions of AM mRNA and protein were increased in the middle to late phase (5-21 days) of HHE, whereas in the brain and lung they were increased much earlier (0.5-5 days). In situ hybridization and immunohistochemistry showed AM mRNA and protein staining to be more intense in the RV in animals in the middle to late phase of HHE exposure than in the controls. During HHE, these changes in AM synthesis, which occurred strongly in the RV, occurred alongside the increase in PAP. Conceivably, AM may play a role in modulating pulmonary hypertension in HHE.     

Calmodulin reverses rundown of L-type Ca(2+) channels in guinea pig ventricular myocytes

Calmodulin (CaM) is implicated in regulation of Ca²⁺ channels as a Ca²⁺ sensor. The effect of CaM on rundown of L-type Ca²⁺ channels in inside-out patch form was investigated in guinea pig ventricular myocytes. Ca²⁺ channel activity disappeared within 1–3 min and did not reappear when the patch was excised and exposed to an artificial intracellular solution. However, application of CaM (0.03, 0.3, 3 µM) + 3 mM ATP to the intracellular solution within 1 min after patch excision resulted in dose-dependent activation of channel activity. Channel activity averaged 11.2%, 94.7%, and 292.9%, respectively, of that in cell-attached mode. Channel activity in inside-out patch mode was induced by CaM + ATP at nanomolar Ca²⁺ concentrations ([Ca²⁺]); however, increase to micromolar [Ca²⁺] rapidly inactivated the channel activity induced, revealing that the effect of CaM on the channel was Ca²⁺ dependent. At the 2nd, 4th, 6th, 8th, and 10th minutes after patch excision, CaM (0.75 µM) + ATP induced Ca²⁺ channel activity to 150%, 100%, 96.9%, 29.3%, and 16.6%, respectively, revealing a time-dependent action of CaM on the channel. CaM added with adenosine 5'-(,-imido)triphosphate (AMP-PNP) also induced channel activity, although with much lower potency and shorter duration. Protein kinase inhibitors KN-62, CaM-dependent protein kinase (CaMK)II 281-309, autocamtide-related CaMKII inhibitor peptide, and K252a (each 1–10 µM) did not block the effect of CaM, indicating that the effect of CaM on the Ca²⁺ channel was phosphorylation independent. Neither CaM nor ATP alone induced Ca²⁺ channel activity, showing a cooperative effect of CaM and ATP on the Ca²⁺ channel. These results suggest that CaM is a crucial regulatory factor of Ca²⁺ channel basal activity. cardiac myocyte; calcium channel; patch clamp