Comparative proteome analysis reveals pathogen specific outer membrane proteins of Leptospira

Proteomes of pathogenic Leptospira interrogans and L. borgpetersenii and the saprophytic L. biflexa were filtered through computational tools to identify Outer Membrane Proteins (OMPs) that satisfy the required biophysical parameters for their presence on the outer membrane. A total of 133, 130, and 144 OMPs were identified in L. interrogans, L. borgpetersenii, and L. biflexa, respectively, which forms approximately 4% of proteomes. A holistic analysis of transporting and pathogenic characteristics of OMPs together with Clusters of Orthologous Groups (COGs) among the OMPs and their distribution across 3 species was made and put forward a set of 21 candidate OMPs specific to pathogenic leptospires. It is also found that proteins homologous to the candidate OMPs were also present in other pathogenic species of leptospires. Six OMPs from L. interrogans and 2 from L. borgpetersenii observed to have similar COGs while those were not found in any intermediate or saprophytic forms. These OMPs appears to have role in infection and pathogenesis and useful for anti‐leptospiral strategies.     

Lactate dehydrogenase (LDH) gene duplication during chordate evolution: the cDNA sequence of the LDH of the tunicate Styela plicata

L-Lactate dehydrogenase (L-LDH, E.C. 1.1.1.27) is encoded by two or three loci in all vertebrates examined, with the exception of lampreys, which have a single LDH locus. Biochemical characterizations of LDH proteins have suggested that a gene duplication early in vertebrate evolution gave rise to Ldh-A and Ldh-B and that an additional locus, Ldh-C arose in a number of lineages more recently. Although some phylogenetic studies of LDH protein sequences have supported this pattern of gene duplication, others have contradicted it. In particular, a number of studies have suggested that Ldh-C represents the earliest divergence among vertebrate LDHs and that it may have diverged from the other loci well before the origin of vertebrates. Such hypotheses make explicit statements about the relationship of vertebrate and invertebrate LDHs, but to date, no closely related invertebrate LDH sequences have been available for comparison. We have attempted to provide further data on the timing of gene duplications leading to multiple vertebrate LDHs by determining the cDNA sequence of the LDH of the tunicate Styela plicata. Phylogenetic analyses of this and other LDH sequences provide strong support for the duplications giving rise to multiple vertebrate LDHs having occurred after vertebrates diverged from tunicates. The timing of these LDH duplications is consistent with data from a number of other gene families suggesting widespread gene duplication near the origin of vertebrates. With respect to the relationships among vertebrate LDHs, our data are not consistent with previous claims that Ldh-C represented the earliest divergence. However, the precise relationships among some of the main lineages of vertebrate LDHs were not resolved in our analyses.      

Mitochondrial signatures revealed panmixia in <Emphasis Type="Italic">Lutjanus argentimaculatus</Emphasis> (Forsskål 1775)

Mangrove red snapper, Lutjanus argentimaculatus is a commercially important fish. The genetic stock structure of L. argentimaculatus from Indian waters was identified using mitochondrial ATPase 6 and ATPase 8, and cytochrome b (Cytb) genes. A 842 bp region of ATPase 6/8 genes and 1105 bp region of Cytb gene were amplified in 120 samples from six different locations along the Indian coast and obtained 58 and 66 haplotypes, respectively. The high haplotype and low nucleotide diversity values along with mismatch distribution, Tajima’s D and Fu’s \(F_\mathrm{s}\) analysis suggested a genetic bottleneck events or founder effect, with subsequent population expansion in L. argentimaculatus. Coefficient of genetic differentiation \((F_\mathrm{ST})\) values was low and nonsignificant for both ATPase 6/8 gene and Cytb genes indicating low genetic differentiation in L. argentimaculatus which can be managed as a unit stock in Indian waters.     

Histological profiling of gonads depicting protandrous hermaphroditism in Eleutheronema tetradactylum

The fourfinger threadfin Eleutheronema tetradactylum is reported as a protandrous hermaphrodite from Australian waters, while being a gonochorist in reports from Singapore and India, with a single report of protandrous hermaphroditism from the latter. Histological analysis of gonads of fish from Indian waters confirms protandrous hermaphroditism in E. tetradactylum. The study was based on 480 fish examined from eight locations along the Indian coast. Mean total length (L_T) of male fish was 240 mm with the transition to female starting from 280 mm L_T. Specimens confirmed as mature females were >380 mm L_T.     

Involvement of a protein kinase activity inducer in DNA double strand break repair and radioresistance of Deinococcus radiodurans

Transgenic bacteria producing pyrroloquinoline quinone, a known cofactor for dehydrogenases and an inducer of a periplasmic protein kinase activity, show resistance to both oxidative stress and protection from nonoxidative effects of radiation and DNA-damaging agents. Deinococcus radiodurans R1 encodes an active pyrroloquinoline quinone synthase, and constitutive synthesis of pyrroloquinoline quinone occurred in wild-type bacteria. Disruption of a genomic copy of pqqE resulted in cells that lacked this cofactor. The mutant showed a nearly 3-log decrease in gamma radiation resistance and a 2-log decrease in mitomycin C tolerance compared to wild-type cells. The mutant cells did not show sensitivity to UVC radiation. Expression of pyrroloquinoline quinone synthase in trans showed that there was functional complementation of gamma resistance and mitomycin C tolerance in the pqqE mutant. The sensitivity to gamma radiation was due to impairment or slow kinetics of DNA double strand break repair. Low levels of (32)P incorporation were observed in total soluble proteins of mutant cells compared to the wild type. The results suggest that pyrroloquinoline quinone has a regulatory role as a cofactor for dehydrogenases and an inducer of selected protein kinase activity in radiation resistance and DNA strand break repair in a radioresistant bacterium.     

Synthesis and biological evaluation of platensimycin analogs

Platensimycin (1) displays antibacterial activity due to its inhibition of the elongation condensing enzyme (FabF), a novel mode of action that could potentially lead to a breakthrough in developing a new generation of antibiotics. The medicinal chemistry efforts were focused on the modification of the enone moiety of platensimycin and several analogs showed significant activity against FabF and possess antibacterial activity.     

Facilitating the Recruitment of Minority Ethnic People into Research: Qualitative Case Study of South Asians and Asthma

Background

There is international interest in enhancing recruitment of minority ethnic people into research, particularly in disease areas with substantial ethnic inequalities. A recent systematic review and meta-analysis found that UK South Asians are at three times increased risk of hospitalisation for asthma when compared to white Europeans. US asthma trials are far more likely to report enrolling minority ethnic people into studies than those conducted in Europe. We investigated approaches to bolster recruitment of South Asians into UK asthma studies through qualitative research with US and UK researchers, and UK community leaders.

Methods and Findings

Interviews were conducted with 36 researchers (19 UK and 17 US) from diverse disciplinary backgrounds and ten community leaders from a range of ethnic, religious, and linguistic backgrounds, followed by self-completion questionnaires. Interviews were digitally recorded, translated where necessary, and transcribed. The Framework approach was used for analysis. Barriers to ethnic minority participation revolved around five key themes: (i) researchers'' own attitudes, which ranged from empathy to antipathy to (in a minority of cases) misgivings about the scientific importance of the question under study; (ii) stereotypes and prejudices about the difficulties in engaging with minority ethnic populations; (iii) the logistical challenges posed by language, cultural differences, and research costs set against the need to demonstrate value for money; (iv) the unique contexts of the two countries; and (v) poorly developed understanding amongst some minority ethnic leaders of what research entails and aims to achieve. US researchers were considerably more positive than their UK counterparts about the importance and logistics of including ethnic minorities, which appeared to a large extent to reflect the longer-term impact of the National Institutes of Health''s requirement to include minority ethnic people.

Conclusions

Most researchers and community leaders view the broadening of participation in research as important and are reasonably optimistic about the feasibility of recruiting South Asians into asthma studies provided that the barriers can be overcome. Suggested strategies for improving recruitment in the UK included a considerably improved support structure to provide academics with essential contextual information (e.g., languages of particular importance and contact with local gatekeepers), and the need to ensure that care is taken to engage with the minority ethnic communities in ways that are both culturally appropriate and sustainable; ensuring reciprocal benefits was seen as one key way of avoiding gatekeeper fatigue. Although voluntary measures to encourage researchers may have some impact, greater impact might be achieved if UK funding bodies followed the lead of the US National Institutes of Health requiring recruitment of ethnic minorities. Such a move is, however, likely in the short- to medium-term, to prove unpopular with many UK academics because of the added “hassle” factor in engaging with more diverse populations than many have hitherto been accustomed to. Please see later in the article for the Editors'' Summary     

1-Benzyl-3-cetyl-2-methylimidazolium iodide (NH125) induces phosphorylation of eukaryotic elongation factor-2 (eEF2): a cautionary note on the anticancer mechanism of an eEF2 kinase inhibitor

Eukaryotic elongation factor-2 kinase (eEF2K) relays growth and stress signals to protein synthesis through phosphorylation and inactivation of eukaryotic elongation factor 2 (eEF2). 1-Benzyl-3-cetyl-2-methylimidazolium iodide (NH125) is a widely accepted inhibitor of mammalian eEF2K and an efficacious anti-proliferation agent against different cancer cells. It implied that eEF2K could be an efficacious anticancer target. However, eEF2K siRNA was ineffective against cancer cells including those sensitive to NH125. To test if pharmacological intervention differs from siRNA interference, we identified a highly selective small molecule eEF2K inhibitor A-484954. Like siRNA, A-484954 had little effect on cancer cell growth. We carefully examined the effect of NH125 and A-484954 on phosphorylation of eEF2, the known cellular substrate of eEF2K. Surprisingly, NH125 increased eEF2 phosphorylation, whereas A-484954 inhibited the phosphorylation as expected for an eEF2K inhibitor. Both A-484954 and eEF2K siRNA inhibited eEF2K and reduced eEF2 phosphorylation with little effect on cancer cell growth. These data demonstrated clearly that the anticancer activity of NH125 was more correlated with induction of eEF2 phosphorylation than inhibition of eEF2K. Actually, induction of eEF2 phosphorylation was reported to correlate with inhibition of cancer cell growth. We compared several known inducers of eEF2 phosphorylation including AMPK activators and an mTOR inhibitor. Interestingly, stronger induction of eEF2 phosphorylation correlated with more effective growth inhibition. We also explored signal transduction pathways leading to NH125-induced eEF2 phosphorylation. Preliminary data suggested that NH125-induced eEF2 phosphorylation was likely mediated through multiple pathways. These observations identified an opportunity for a new multipathway approach to anticancer therapies.