Loss of DNA repair mechanisms in cardiac myocytes induce dilated cardiomyopathy

Cardiomyopathy is a progressive disease of the myocardium leading to impaired contractility. Genotoxic cancer therapies are known to be potent drivers of cardiomyopathy, whereas causes of spontaneous disease remain unclear. To test the hypothesis that endogenous genotoxic stress contributes to cardiomyopathy, we deleted the DNA repair gene Ercc1 specifically in striated muscle using a floxed allele of Ercc1 and mice expressing Cre under control of the muscle-specific creatinine kinase (Ckmm) promoter or depleted systemically (Ercc1^−/D mice). Ckmm-Cre^+/−;Ercc1^−/fl mice expired suddenly of heart disease by 7 months of age. As young adults, the hearts of Ckmm-Cre^+/−;Ercc1^−/fl mice were structurally and functionally normal, but by 6-months-of-age, there was significant ventricular dilation, wall thinning, interstitial fibrosis, and systolic dysfunction indicative of dilated cardiomyopathy. Cardiac tissue from the tissue-specific or systemic model showed increased apoptosis and cardiac myocytes from Ckmm-Cre^+/-;Ercc1^−/fl mice were hypersensitive to genotoxins, resulting in apoptosis. p53 levels and target gene expression, including several antioxidants, were increased in cardiac tissue from Ckmm-Cre^+/−;Ercc1^−/fl and Ercc1^−/D mice. Despite this, cardiac tissue from older mutant mice showed evidence of increased oxidative stress. Genetic or pharmacologic inhibition of p53 attenuated apoptosis and improved disease markers. Similarly, overexpression of mitochondrial-targeted catalase improved disease markers. Together, these data support the conclusion that DNA damage produced endogenously can drive cardiac disease and does so mechanistically via chronic activation of p53 and increased oxidative stress, driving cardiac myocyte apoptosis, dilated cardiomyopathy, and sudden death.     

Leaf shelter-building caterpillars harness forces generated by axial retraction of stretched and wetted silk

Leaf shelter-building caterpillars generate most of the force required to pull leaf surfaces together by stretching silk strands while spinning. Axially retractive forces produced by columns of stretched strands enabled caterpillars in our study to generate forces as great as 0.3 Newtons (i.e., a 30-g force). We found that caterpillar silk also contracts instantly when wetted, producing an additional, though smaller, axially retractive force. Contraction ratios (final length/ original length) of the wetted silk of 19 species ranged from 0.21 to 0.93 and were smallest among species that use their silk to make leaf shelters. Our study, the first to identify the specific sources of the energy harnessed by caterpillars to tie, roll, or fold leaves, indicates that silk properties and caterpillar behavior have coevolved to facilitate the leaf shelter-building process.     

Patchy deletion of Bmpr1a potentiates proximal pulmonary artery remodeling in mice exposed to chronic hypoxia

Reduced vascular expression of bone morphogenetic protein type IA receptor (Bmpr1a) has been found in patients with pulmonary arterial hypertension. Our previous studies in mice with patchy deletion of Bmpr1a in vascular smooth muscle cells and cardiac myocytes showed decreased distal vascular remodeling despite a similar severity of hypoxic pulmonary hypertension (HPH). We speculate increased stiffness from ectopic deposition of collagen in proximal pulmonary arteries might account for HPH. Pulsatile pressure-flow relationships were measured in isolated, ventilated, perfused lungs of SM22α;TRE-Cre;R26R;Bmpr1a ^flox/flox (KO) mice and wild-type littermates, following 21 days (hypoxia) and 0 days (control) of chronic hypoxia. Pulmonary vascular impedance, which yields insight into proximal and distal arterial remodeling, was calculated. Reduced Bmpr1a expression had no effect on input impedance Z ₀ (P = 0.52) or characteristic impedance Z _C (P = 0.18) under control conditions; it also had no effect on the decrease in Z ₀ via acute rho kinase inhibition. However, following chronic hypoxia, reduced Bmpr1a expression increased Z _C (P < 0.001) without affecting Z ₀ (P = 0.72). These results demonstrate that Bmpr1a deficiency does not significantly alter the hemodynamic function of the distal vasculature or its response to chronic hypoxia but larger, more proximal arteries are affected. In particular, reduced Bmpr1a expression likely decreased dilatation and increased stiffening in response to hypoxia, probably by collagen accumulation. Increased PA stiffness can have a significant impact on right ventricular function. This study illustrates for the first time how proximal pulmonary artery changes in the absence of distal pulmonary artery changes contribute to pulmonary arterial hypertension.     

STING Millennium Suite: integrated software for extensive analyses of 3d structures of proteins and their complexes

Background  

The integration of many aspects of protein/DNA structure analysis is an important requirement for software products in general area of structural bioinformatics. In fact, there are too few software packages on the internet which can be described as successful in this respect. We might say that what is still missing is publicly available, web based software for interactive analysis of the sequence/structure/function of proteins and their complexes with DNA and ligands. Some of existing software packages do have certain level of integration and do offer analysis of several structure related parameters, however not to the extent generally demanded by a user.     

Genome-scale metabolic reconstruction and <Emphasis Type="Italic">in silico</Emphasis> analysis of methylotrophic yeast <Emphasis Type="Italic">Pichia pastoris</Emphasis> for strain improvement

Background  

Pichia pastoris has been recognized as an effective host for recombinant protein production. A number of studies have been reported for improving this expression system. However, its physiology and cellular metabolism still remained largely uncharacterized. Thus, it is highly desirable to establish a systems biotechnological framework, in which a comprehensive in silico model of P. pastoris can be employed together with high throughput experimental data analysis, for better understanding of the methylotrophic yeast's metabolism.     

Residence time and kinetic efficiency analysis of extracellular signal-regulated kinase 2 inhibitors

The RAS/RAF/MEK/ERK signal transduction cascade plays an important role in the regulation of critical cellular processes such as cell proliferation, migration, and differentiation. The up-regulation of this pathway can negatively affect cell homeostasis and is responsible for the development of various forms of cancer and inflammation processes. Therefore, there is a strong interest in pursuing drug programs targeting some of the enzymes involved in this pathway. In addition to the determination of K_i, K_d, IC₅₀, and/or EC_50, a more thorough kinetic analysis can provide useful information for the selection of the best lead series during the early stage of the drug discovery process. This study describes a medium-throughput fluorescent probe displacement assay for the rapid determination of the k_off constant, residence time, and kinetic efficiency for ERK (extracellular signal-regulated kinase) inhibitors. Using this method, we have identified several inhibitors that we have subjected to further kinetic analysis by comparing k_off constants determined for these time-dependent inhibitors using either the active or inactive form of ERK2.     

Combined experimental and computational strategies define an expansive regulon for GcvB small RNA

The bacterial small RNA GcvB has been known as a translational repressor of mRNAs encoding amino acid transporters and has been postulated to limit uptake of unnecessary amino acids under nutrient-rich conditions. In this issue of Molecular Microbiology, Sharma et al. (2011) provide evidence for a much broader role for GcvB as a global regulator of amino acid metabolism. Using a unique combination of experimental and biocomputational approaches, the authors triple the size of the GcvB regulon, making it the largest sRNA regulon defined to date.     

The L-type calcium channel alpha 1C subunit gene undergoes extensive, uncoordinated alternative splicing

The alpha1C subunit is the pore-forming protein for the L-type calcium channel. Previous studies indicate that there is possible tissue-specific alternative splicing of this gene. In this study we cloned the entire open reading frame of the alpha1C subunit cDNA from adult rat cardiac myocytes in a single piece (6.64 kb). Using 75 positive clones that were identified by restriction enzyme mapping, we tested the alternative splicing patterns of the Ca(v) 1.2 gene that encodes the alpha1C subunit protein and focused on five loci: IS6, post-IS6, IIIS2, IVS3, and the c-terminus. The results indicate that: (1) alternative splicing occurs in most of the loci, giving rise to two or three different isoforms at those sites; (2) there is a predominant form for each splicing site, (3) there does not appear to be consistent coordination of splicing at multiple loci of this gene. Alternative splicing is not tissue-specific in most regions.