Deletion of caveolin scaffolding domain alters cancer cell migration

Caveolin-1 (Cav-1) is an integral membrane protein that plays an important role in proliferative and terminally differentiated cells. As a structural component of Caveolae, Cav-1 interacts with signaling molecules via a caveolin scaffolding domain (CSD) regulating cell signaling. Recent reports have shown that Cav-1 is a negative regulator in tumor metastasis. Therefore, we hypothesize that Cav-1 inhibits cell migration through its CSD. HeLa cells were engineered to overexpress Cav-1 (Cav-1 OE), Cav-1 without a functional CSD (?CSD), or enhanced green fluorescent protein (EGFP) as a control. HeLa cell migration was suppressed in Cav-1 OE cells while ?CSD showed increased migration, which corresponded to a decrease in the tight junction protein, zonula occludens (ZO-1). The migration phenotype was confirmed in multiple cancer cell lines. Phosphorylated STAT-3 was decreased in Cav-1 OE cells compared to control and ?CSD cells; reducing STAT-3 expression alone decreased cell migration. ?CSD blunted HeLa proliferation by increasing the number of cells in the G2/M phase of the cell cycle. Overexpressing the CSD peptide alone suppressed HeLa cell migration and inhibited pSTAT3. These findings suggest that Cav-1 CSD may be critical in controlling the dynamic phenotype of cancer cells by facilitating the interaction of specific signal transduction pathways, regulating STAT3 and participating in a G2/M checkpoint. Modulating the CSD and targeting specific proteins may offer potential new therapies in the treatment of cancer metastasis.     

Investigation of tRNA<Superscript>Leu/Lys</Superscript> and ATPase 6 Genes Mutations in Huntington’s Disease

Huntington disease (HD) is a genetically dominant condition caused by expanded CAG repeats which code for glutamine in the HD gene product, huntingtin. Huntingtin is expressed in almost all tissues, so abnormalities outside the brain can also be expected. Involvement of nuclei and mitochondria in HD pathophysiology has been suggested. In fact mitochondrial dysfunction is reported in brains of patients suffering from HD. The tRNA gene mutations are one of hot spots that can cause mitochondrial disorders. In this study, possible mitochondrial DNA (mtDNA) damage was evaluated by screening for mutations in the tRNA^leu/lys and ATPase 6 genes of 20 patients with HD, using PCR and automated DNA sequencing. Mutations including an A8656G mutation in one patient were observed, which may be causal to the disease. Understanding the role of mitochondria in the pathogenesis of neurodegenerative diseases could potentially be important for the development of therapeutic strategies in HD.     

The c.42_52del11 Mutation in TPRN and Progressive Hearing Loss in a Family from Pakistan

The DFNB79 locus harbors TPRN mutations in which have been reported in a few families with deafness. Four frameshift mutations in TPRN have been described to cause severe or severe-to-profound hearing loss in Moroccan and Pakistani families, and a single frameshift mutation was associated with progressive hearing loss in deaf individuals in a Dutch family. We identified a Pakistani family in which the affected individuals were homozygous for a pathogenic mutation, c.42_52del11, in TPRN (p.G15Afs150X). In contrast to the previously reported individuals affected by the same mutation, hearing loss is likely to be progressive in this family. Thus the same mutation of TPRN can be associated with different thresholds of hearing as well as differences in the stability of the phenotype.     

VEGF gene mRNA expression in patients with coronary artery disease

To assess the expression of vascular endothelium growth factor (VEGF) mRNA in unstimulated peripheral blood mononuclear cells of patients with and without coronary artery disease (CAD). We also studied whether the functional VEGF -2,578C/A polymorphism may influence the level of VEGF mRNA expression in individuals undergoing coronary angiography because chest pain. We assessed 50 consecutive patients with angiographically confirmed CAD (CAD+). Also, 50 consecutive individuals with normal coronary studies were included in the study for comparison. VEGF mRNA expression was examined using quantitative real-time PCR and genotyping for VEGF -2,578C/A was performed using ARMS-PCR technique. VEGF mRNA expression was significantly decreased in CAD+ patients when compared to CAD- individuals (p = 0.01). The frequency of VEGF -2578 allele C and genotype CC was increased in CAD+ patients. In this regard, homozygosity for the CC genotype was more commonly observed in CAD+ (30 %) than in those without CAD disease (18 %). However, the difference was slightly out of the range of significance (p = 0.1). In addition, a trend for reduction in the expression of VEGF mRNA was observed when patients carrying the VEGF -2,578AA genotype were compared with those VEGF -2,578AC heterozygous or those homozygous for the VEGF -2,578CC genotype. VEGF gene expression is decreased in individuals with CAD+ disease. The VEGF -2,578C/A polymorphism may influences the expression of VEGF.     

Molecular docking analysis of fluoroquinolones and other natural and synthetic compounds with the HCV NS3 helicase

It is of an interest to document the molecular docking analysis of fluoroquinolones and other natural and synthetic compounds with the HCV NS3 helicase. Data shows that three fluoroquinolones interacted with the NS3 helicase in the catalytic region, targeting some of the amino acids known to play a crucial role in NS3 helicase activity. Similarly, binding energy shows that the fluoroquinolones were comparable to the thiazolpiperazinyl derivatives, while superior to several of the synthetic and natural derivatives. The results show three fluoroquinolones to be potent helicase inhibitors that can be repurposed as supplemental therapy against HCV especially in cases non-responsive to DAAAs.     

Evaluation of Prader‐Willi Syndrome Gene MAGEL2 in Severe Childhood‐Onset Obesity

MAGEL2 is one of the five genes inactivated in Prader‐Willi Syndrome, a neurodevelopmental chromosome microdeletion disorder modified by genomic imprinting. By early childhood, individuals with Prader‐Willi Syndrome exhibit hypothalamic dysfunction, including hyperphagia, and become obese in the absence of behavioral intervention. Murine Magel2 is highly expressed in the hypothalamus during development. We screened the MAGEL2 open reading frame for mutations in genomic DNA samples from hyperphagic but non‐dysmorphic individuals with severe childhood‐onset obesity. Although no mutations likely to affect gene function were identified, we identified three variant alleles. We conclude that severe childhood‐onset obesity is not commonly caused by MAGEL2 mutations.     

Production enhancement and refolding of caprine growth hormone expressed in Escherichia coli

This study describes comparison between IPTG and lactose induction on expression of caprine growth hormone (cGH), enhancing cell densities of Escherichia coli cultures and refolding the recombinant cGH, produced as inclusion bodies, to biologically active state. 2–3 times higher cell densities were obtained in shake flask cultures when induction was done with lactose showing almost same level of expression as in case of IPTG induction. With lactose induction highest cell densities were achieved in TB (OD₆₀₀ 16.3) and M9NG (OD₆₀₀ 16.1) media, producing 885 and 892 mg cGH per liter of the culture, respectively. Lactose induction done at mid-exponential stage resulted in a higher cell density and thus higher product yield. cGH over-expressed as inclusion bodies was solubilized in 50 mM Tris–Cl buffer (pH 12.5) containing 2 M urea, followed by dilution and lowering the pH in a step-wise manner to obtain the final solution in 50 mM Tris–Cl (pH 9.5). The cGH was purified by Q-Sepharose chromatography followed by gel filtration with a recovery yield of 39% on the basis of total cell proteins. The product thus obtained showed a single band by SDS–PAGE analysis. MALDI-TOF analysis showed a single peak with a mass of 21,851 dalton, which is very close to its calculated molecular weight. A bioassay based on proliferation of Nb2 rat lymphoma cells showed that the purified cGH was biologically active.     

Genetics of obesity

Considerable attention is currently being paid to the secular changes in food intake and physical activity that underlie the increase in the prevalence of obesity that is apparent in many societies. While this is laudable it would be unwise to view these environmental factors in isolation from the biological factors that normally control body weight and composition and the compelling evidence that inter-individual differences in susceptibility to obesity have strong genetic determinants. This is particularly important, as it is only in the past decade that we have begun to obtain substantive information regarding the molecular constituents of pathways controlling mammalian energy balance and therefore, for the first time, are in a position to achieve a better mechanistic understanding of this disease. Population-based association and linkage studies have highlighted a number of loci at which genetic variation is associated with obesity and related phenotypes and the identification and characterization of monogenic obesity syndromes has been particularly fruitful. While there is widespread acceptance that hereditary factors might predispose to human obesity, it is frequently assumed that such factors would influence metabolic rate or the selective partitioning of excess calories into fat. However, it is notable that, thus far, all monogenic defects causing human obesity actually disrupt hypothalamic pathways and have a profound effect on satiety and food intake. To conclude, the evidence we have to date suggests that the major impact of genes on human obesity is just as likely (or perhaps more likely) to directly impact on hunger, satiety and food intake rather than metabolic rate or nutrient partitioning. At the risk of oversimplification, it seems that from an aetiological/genetic standpoint, human obesity appears less a metabolic than a neuro-behavioural disease.     

Evolutionary trends in Tetrastigma (Vitaceae): Morphological diversity and taxonomic implications

Tetrastigma (Miq.) Planch. (Vitaceae) is a genus with ca. 100 species showing great morphological diversity. Previous molecular phylogenetic studies suggested that traditional classification systems are not consistent with the molecular phylogeny, and Tetrastigma is undergoing further systematic investigation. We traced the evolutionary trends of 20 morphological characters within a robust phylogenetic framework. Our results revealed that many morphological characters show either multiple transitions or few state changes, however, some characters show distinct variation. The two subgenera in Tetrastigma (subgen. Tetrastigma and subgen. Palmicirrata) based on unbranched/bifurcate versus digitately branched tendrils are not supported because subgen. Tetrastigma is paraphyletic. However, the unbranched versus bifurcate/digitately branched tendril is of taxonomic utility to characterize some of the major clades. Inflorescences in Tetrastigma appear axillary, but are leaf‐opposed on a compressed axillary shoot. We found most of the species in Tetrastigma retained the ancestral compound dichasial inflorescence, except those of clade IV that have derived pseudo‐umbellate inflorescences. Other characters including habit, leaf organization, and berry shape provide additional morphological support for the major clades. Our morphological analysis and recent molecular study suggest each of the five major clades within Tetrastigma be treated as distinct taxonomic sections (five sections in the genus).     

A molecular evaluation of dengue virus pathogenesis and its latest vaccine strategies

More than one third of the world’s population living in tropical and subtropical areas of the world is at risk of dengue infections and as many as 100 million people are yearly infected. This disease has reemerged during the past 20 years in the form of an epidemic. Dengue is caused by one of four related serotypes of dengue virus and often leads to severe forms of the disease, resulting commonly from secondary infections. Dengue virus is a mosquito borne virus, belongs to the family Flaviviridae and consists of a single stranded positive sense RNA genome. Like other RNA viruses it escapes defense mechanisms and neutralization attempts by mutations, which make it more resistant and adaptable to its environment. Antiviral strategies and vaccine development is thus impaired and hence to date there is no licensed vaccine available for dengue virus. Here we discuss various efforts made towards the identification of potential vaccine targets for dengue as well as various strategies employed by research groups/pharmaceutical companies towards the development of a successful dengue vaccine.