Evolution and structural analysis of Glossina morsitans (Diptera; Glossinidae) Tetraspanins

Tetraspanins are important conserved integral membrane proteins expressed in many organisms. Although there is limited knowledge about the full repertoire, evolution and structural characteristics of individual members in various organisms, data obtained so far show that tetraspanins play major roles in membrane biology, visual processing, memory, olfactory signal processing, and mechanosensory antennal inputs. Thus, these proteins are potential targets for control of insect pests. Here, we report that the genome of the tsetse fly, Glossina morsitans (Diptera: Glossinidae) encodes at least seventeen tetraspanins (GmTsps), all containing the signature features found in the tetraspanin superfamily members. Whereas six of the GmTsps have been previously reported, eleven could be classified as novel because their amino acid sequences do not map to characterized tetraspanins in the available protein data bases. We present a model of the GmTsps by using GmTsp42Ed, whose presence and expression has been recently detected by transcriptomics and proteomics analyses of G. morsitans. Phylogenetically, the identified GmTsps segregate into three major clusters. Structurally, the GmTsps are largely similar to vertebrate tetraspanins. In view of the exploitation of tetraspanins by organisms for survival, these proteins could be targeted using specific antibodies, recombinant large extracellular loop (LEL) domains, small-molecule mimetics and siRNAs as potential novel and efficacious putative targets to combat African trypanosomiasis by killing the tsetse fly vector

Pitx2 Expression Defines a Left Cardiac Lineage of Cells: Evidence for Atrial and Ventricular Molecular Isomerism in the iv/iv Mice

AbstractThe homeobox gene Pitx2 has been characterized as a mediator of left-right signaling in heart, gut, and lung morphogenesis. However, the relationship between the developmental role of Pitx2 and its expression pattern at the organ level has not been explored. In this study we focus on the role of Pitx2 in heart morphogenesis. Chicken Pitx2 transcripts are present in the left portion of the cardiac crescent and in the left side of the heart tube. Through looping Pitx2 is present in the left atrium, in the ventral portion of the ventricles and in the left-ventral part of the outflow tract. Mouse Pitx2 shows a similar developmental profile of expression. To test whether Pitx2 represents a lineage marker we have tagged the left portion of the chicken cardiac tube with fluorescent DiD. Labeled cells were found at HH16 in the left atrium and in the ventral region of the ventricles and the outflow tract. In the iv/iv mouse model of cardiac heterotaxia Pitx2 was abnormally expressed in the atrial and in the ventricular chambers. Furthermore, altered Pitx2 expression correlated with the occurrence of DORV. Our data reveal the existence of molecular isomerism not only in the atrial, but also in the ventricular compartment of the heart

Chamber Formation and Morphogenesis in the Developing Mammalian Heart

AbstractIn this study we challenge the generally accepted view that cardiac chambers form from an array of segmental primordia arranged along the anteroposterior axis of the linear and looping heart tube. We traced the spatial pattern of expression of genes encoding atrial natriuretic factor, sarcoplasmic reticulum calcium ATPase, Chisel, Irx5, Irx4, myosin light chain 2v, and β-myosin heavy chain and related these to morphogenesis. Based on the patterns we propose a two-step model for chamber formation in the embryonic heart. First, a linear heart forms, which is composed of “primary” myocardium that nonetheless shows polarity in phenotype and gene expression along its anteroposterior and dorsoventral axes. Second, specialized ventricular chamber myocardium is specified at the ventral surface of the linear heart tube, while distinct left and right atrial myocardium forms more caudally on laterodorsal surfaces. The process of looping aligns these primordial chambers such that they face the outer curvature. Myocardium of the inner curvature, as well as that of inflow tract, atrioventricular canal, and outflow tract, retains the molecular signature originally found in linear heart tube myocardium. Evidence for distinct transcriptional programs which govern compartmentalization in the forming heart is seen in the patterns of expression of Hand1 for the dorsoventral axis, Irx4 and Tbx5 for the anteroposterior axis, and Irx5 for the distinction between primary and chamber myocardium

Epigenetic state changes underlie metabolic switch in mouse post-infarction border zone cardiomyocytes

Myocardial infarction causes ventricular muscle loss and formation of scar tissue. The surviving myocardium in the border zone, located adjacent to the infarct, undergoes profound changes in function, structure and composition. How and to what extent these changes of border zone cardiomyocytes are regulated epigenetically is not fully understood. Here, we obtained transcriptomes of PCM-1-sorted mouse cardiomyocyte nuclei of healthy left ventricle and 7 days post myocardial infarction border zone tissue. We validated previously observed downregulation of genes involved in fatty acid metabolism, oxidative phosphorylation and mitochondrial function in border zone-derived cardiomyocytes, and observed a modest induction of genes involved in glycolysis, including Slc2a1 (Glut1) and Pfkp. To gain insight into the underlying epigenetic regulatory mechanisms, we performed H3K27ac profiling of healthy and border zone cardiomyocyte nuclei. We confirmed the switch from Mef2-to AP-1 chromatin association in border zone cardiomyocytes, and observed, in addition, an enrichment of PPAR/RXR binding motifs in the sites with reduced H3K27ac signal. We detected downregulation and accompanying epigenetic state changes at several key PPAR target genes including Ppargc1a (PGC-1α), Cpt2, Ech1, Fabpc3 and Vldrl in border zone cardiomyocytes. These data indicate that changes in epigenetic state and gene regulation underlie the maintained metabolic switch in border zone cardiomyocytes

Neuregulin-1 enhances cell-cycle activity, delays cardiac fibrosis, and improves cardiac performance in rat pups with right ventricular pressure load

Objectives: Right ventricular (RV) failure is a leading cause of death in patients with congenital heart disease. RV failure is kept at bay during childhood. Limited proliferation of cardiomyocytes is present in the postnatal heart. We propose that cardiomyocyte proliferation improves RV adaptation to pressure load (PL). We studied adaptation in response to increased RV PL and the role of increased cardiomyocyte cell cycle activity (CCA) in rat pups growing into adulthood. Methods: We induced RV PL at day of weaning in rats (3 weeks; 30-40 g) by pulmonary artery banding and followed rats into adulthood (300 g). We performed histological analyses and RNA sequencing analysis. To study the effects of increased cardiomyocyte cell cycle activity, we administered neuregulin-1 (NRG1), a growth factor involved in cardiac development. Results: PL induced an increase in CCA, with subsequent decline of CCA (sham/PL at 4 weeks: 0.14%/0.83%; P = .04 and 8 weeks: 0.00%/0.00%; P = .484) and cardiac function (cardiac index: control/PL 4 weeks: 4.41/3.29; P = .468 and 8 weeks: 3.57/1.44; P = .024). RNA sequencing analysis revealed delayed maturation and increased CCA pathways. NRG1 stimulated CCA (PL vehicle/NRG1 at 2 weeks: 0.62%/2.28%; P = .003), improved cardiac function (cardiac index control vs vehicle/NRG1 at 2 weeks: 4.21 vs 3.07/4.17; P = .009/.705) and postponed fibrosis (control vs vehicle/NRG1 at 4 weeks: 1.66 vs 4.82%/2.97%; P = .009/.078) in RV PL rats during childhood. Conclusions: RV PL during growth induces a transient CCA increase. Further CCA stimulation improves cardiac function and delays fibrosis. This proof-of-concept study shows that stimulation of CCA can improve RV adaptation to PL in the postnatal developing heart and might provide a new approach to preserve RV function in patients with congenital heart disease.</p

Neuregulin-1 enhances cell-cycle activity, delays cardiac fibrosis, and improves cardiac performance in rat pups with right ventricular pressure load

Objectives: Right ventricular (RV) failure is a leading cause of death in patients with congenital heart disease. RV failure is kept at bay during childhood. Limited proliferation of cardiomyocytes is present in the postnatal heart. We propose that cardiomyocyte proliferation improves RV adaptation to pressure load (PL). We studied adaptation in response to increased RV PL and the role of increased cardiomyocyte cell cycle activity (CCA) in rat pups growing into adulthood. Methods: We induced RV PL at day of weaning in rats (3 weeks; 30-40 g) by pulmonary artery banding and followed rats into adulthood (300 g). We performed histological analyses and RNA sequencing analysis. To study the effects of increased cardiomyocyte cell cycle activity, we administered neuregulin-1 (NRG1), a growth factor involved in cardiac development. Results: PL induced an increase in CCA, with subsequent decline of CCA (sham/PL at 4 weeks: 0.14%/0.83%; P = .04 and 8 weeks: 0.00%/0.00%; P = .484) and cardiac function (cardiac index: control/PL 4 weeks: 4.41/3.29; P = .468 and 8 weeks: 3.57/1.44; P = .024). RNA sequencing analysis revealed delayed maturation and increased CCA pathways. NRG1 stimulated CCA (PL vehicle/NRG1 at 2 weeks: 0.62%/2.28%; P = .003), improved cardiac function (cardiac index control vs vehicle/NRG1 at 2 weeks: 4.21 vs 3.07/4.17; P = .009/.705) and postponed fibrosis (control vs vehicle/NRG1 at 4 weeks: 1.66 vs 4.82%/2.97%; P = .009/.078) in RV PL rats during childhood. Conclusions: RV PL during growth induces a transient CCA increase. Further CCA stimulation improves cardiac function and delays fibrosis. This proof-of-concept study shows that stimulation of CCA can improve RV adaptation to PL in the postnatal developing heart and might provide a new approach to preserve RV function in patients with congenital heart disease.</p

Tbx2 and Tbx3 induce atrioventricular myocardial development and endocardial cushion formation

A key step in heart development is the coordinated development of the atrioventricular canal (AVC), the constriction between the atria and ventricles that electrically and physically separates the chambers, and the development of the atrioventricular valves that ensure unidirectional blood flow. Using knock-out and inducible overexpression mouse models, we provide evidence that the developmentally important T-box factors Tbx2 and Tbx3, in a functionally redundant manner, maintain the AVC myocardium phenotype during the process of chamber differentiation. Expression profiling and ChIP-sequencing analysis of Tbx3 revealed that it directly interacts with and represses chamber myocardial genes, and induces the atrioventricular pacemaker-like phenotype by activating relevant genes. Moreover, mutant mice lacking 3 or 4 functional alleles of Tbx2 and Tbx3 failed to form atrioventricular cushions, precursors of the valves and septa. Tbx2 and Tbx3 trigger development of the cushions through a regulatory feed-forward loop with Bmp2, thus providing a mechanism for the co-localization and coordination of these important processes in heart development

Pioneering disadvantage : consumer reactions to marketing mix strategies / M. Sadiq Sohail and Mohamad Farid Mahmood

Prior research has focused on the early entrant advantage, although a growing body of knowledgde suggests suggests that late movers have been outselling pioneers. In this research, the authors examine the issue of a pay phone service provider in Malaysia. In 1990, the company launches its product and creates an industry, but by the turn of the century, it has a meager 17 per cent of the market share. The Paper examines the specific issue of how a market pioneer has been eclipsed. It is hypothesized that the main cause of the declining sales is the in appropriate marketing mix strategies. The major findings based on a survey include consumers' acceptance level of the product, price, place, promotion, people, process and physical evidence strategies of the company

TGFBR1 Variants Can Associate with Non-Syndromic Congenital Heart Disease without Aortopathy

BACKGROUND: Congenital heart diseases (CHD) are the most common congenital malformations in newborns and remain the leading cause of mortality among infants under one year old. Molecular diagnosis is crucial to evaluate the recurrence risk and to address future prenatal diagnosis. Here, we describe two families with various forms of inherited non-syndromic CHD and the genetic work-up and resultant findings. METHODS: Next-generation sequencing (NGS) was employed in both families to uncover the genetic cause. In addition, we performed functional analysis to investigate the consequences of the identified variants in vitro. RESULTS: NGS identified possible causative variants in both families in the protein kinase domain of the TGFBR1 gene. These variants occurred on the same amino acid, but resulted in differently substituted amino acids (p.R398C/p.R398H). Both variants co-segregate with the disease, are extremely rare or unique, and occur in an evolutionary highly conserved domain of the protein. Furthermore, both variants demonstrated a significantly altered TGFBR1-smad signaling activity. Clinical investigation revealed that none of the carriers had (signs of) aortopathy. CONCLUSION: In conclusion, we describe two families, with various forms of inherited non-syndromic CHD without aortopathies, associated with unique/rare variants in TGFBR1 that display altered TGF-beta signaling. These findings highlight involvement of TGFBR1 in CHD, and warrant consideration of potential causative TGFBR1 variants also in CHD patients without aortopathies

Variation in a Left Ventricle–Specific Hand1 Enhancer Impairs GATA Transcription Factor Binding and Disrupts Conduction System Development and Function

Rationale The ventricular conduction system (VCS) rapidly propagates electrical impulses through the working myocardium of the ventricles to coordinate chamber contraction. Genome-wide association studies (GWAS) have associated nucleotide polymorphisms, most are located within regulatory intergenic or intronic sequences, with variation in VCS function. Two highly correlated polymorphisms (r2>0.99) associated with VCS functional variation (rs13165478 and rs13185595) occur 5’ to the gene encoding the bHLH transcription factor HAND1. Objective Here, we test the hypothesis that these polymorphisms influence HAND1 transcription thereby influencing VCS development and function. Methods and Results We employed transgenic mouse models to identify an enhancer that is sufficient for left ventricle (LV) cis-regulatory activity. Two evolutionarily conserved GATA transcription factor cis-binding elements within this enhancer are bound by GATA4 and are necessary for cis-regulatory activity, as shown by in vitro DNA binding assays. CRISPR/Cas9-mediated deletion of this enhancer dramatically reduces Hand1 expression solely within the LV but does not phenocopy previously published mouse models of cardiac Hand1 loss-of-function. Electrophysiological and morphological analyses reveals that mice homozygous for this deleted enhancer display a morphologically abnormal VCS, and a conduction system phenotype consistent with right bundle branch block. Using 1000 Genomes Project data, we identify three additional SNPs, located within the Hand1 LV enhancer, that compose a haplotype with rs13165478 and rs13185595. One of these SNPs, rs10054375, overlaps with a critical GATA cis-regulatory element within the Hand1 LV enhancer. This SNP, when tested in electrophoretic mobility shift assays (EMSA), disrupts GATA4 DNA-binding. Modeling two of these SNPs in mice causes diminished Hand1 expression and mice present with abnormal VCS function. Conclusions Together, these findings reveal that SNP rs10054375, which is located within a necessary and sufficient LV-specific Hand1 enhancer, exhibits reduces GATA DNA-binding in EMSA and this enhancer in total, is required for VCS development and function in mice and perhaps humans