Rapamycin inhibition of baculovirus recombinant (BVr) ribosomal protein S6 kinase (S6K1) is mediated by an event other than phosphorylation

Background

Ribosomal protein S6 kinase 1(S6K1) is an evolutionary conserved kinase that is activated in response to growth factors and viral stimuli to influence cellular growth and proliferation. This downstream effector of target of rapamycin (TOR) signaling cascade is known to be directly activated by TOR- kinase mediated hydrophobic motif (HM) phosphorylation at Threonine 412 (T412). Selective loss of this phosphorylation by inactivation of TOR kinase or activation/recruitment of a phosphatase has accordingly been implicated in mediating inhibition by rapamycin.

Findings

We present evidence that baculovirus driven expression of S6K1 in insect cells (Sf9) fails to activate the enzyme and instead renders it modestly active representing 4-6 folds less activity than its fully active mammalian counterpart. Contrary to the contention that viral infection activates TOR signaling pathway, we report that BVr enzyme fails to exhibit putative TOR dependent phosphorylation at the HM and the resultant phosphorylation at the activation loop (AL) of the enzyme, correlating with the level of activity observed. Surprisingly, the BVr enzyme continued to exhibit sensitivity to rapamycin that remained unaffected by mutations compromised for TOR phosphorylation (T412A) or deletions compromised for TOR binding (ΔNH _2-46/ΔCT₁₀₄).

Conclusions

These data together with the ability of the BVr enzyme to resist inactivation by phosphatases indicate that inhibition by rapamycin is not mediated by any phosphorylation event in general and TOR dependent phosphorylation in particular.     

Integrating prevention of mother-to-child HIV transmission programs to improve uptake: a systematic review

Background

We performed a systematic review to assess the effect of integrated perinatal prevention of mother-to-child transmission of HIV interventions compared to non- or partially integrated services on the uptake in low- and middle-income countries.

Methods

We searched for experimental, quasi-experimental and controlled observational studies in any language from 21 databases and grey literature sources.

Results

Out of 28 654 citations retrieved, five studies met our inclusion criteria. A cluster randomized controlled trial reported higher probability of nevirapine uptake at the labor wards implementing HIV testing and structured nevirapine adherence assessment (RRR 1.37, bootstrapped 95% CI, 1.04–1.77). A stepped wedge design study showed marked improvement in antiretroviral therapy (ART) enrolment (44.4% versus 25.3%, p<0.001) and initiation (32.9% versus 14.4%, p<0.001) in integrated care, but the median gestational age of ART initiation (27.1 versus 27.7 weeks, p = 0.4), ART duration (10.8 versus 10.0 weeks, p = 0.3) or 90 days ART retention (87.8% versus 91.3%, p = 0.3) did not differ significantly. A cohort study reported no significant difference either in the ART coverage (55% versus 48% versus 47%, p = 0.29) or eight weeks of ART duration before the delivery (50% versus 42% versus 52%; p = 0.96) between integrated, proximal and distal partially integrated care. Two before and after studies assessed the impact of integration on HIV testing uptake in antenatal care. The first study reported that significantly more women received information on PMTCT (92% versus 77%, p<0.001), were tested (76% versus 62%, p<0.001) and learned their HIV status (66% versus 55%, p<0.001) after integration. The second study also reported significant increase in HIV testing uptake after integration (98.8% versus 52.6%, p<0.001).

Conclusion

Limited, non-generalizable evidence supports the effectiveness of integrated PMTCT programs. More research measuring coverage and other relevant outcomes is urgently needed to inform the design of services delivering PMTCT programs.     

Prevalence of type 2 diabetes in the States of the co-operation council for the arab States of the gulf: a systematic review

Aims

The recent and ongoing worldwide expansion in prevalence of Type 2 Diabetes (T2DM) is a considerable risk to individuals, health systems and economies. The increase in prevalence has been particularly marked in the states of the Co-operation Council for the Arab States of the Gulf (GCC), and these trends are set to continue. We aimed to systematically review the current prevalence of T2DM within these states, and also within particular sub-populations.

Methods

We identified 27 published studies for review. Studies were identified by systematic database searches. Medline and Embase were searched using terms such as diabetes mellitus, non-insulin-dependent, hyperglycemia, prevalence, epidemiology and Gulf States. Our search also included scanning reference lists, contacting experts and hand-searching key journals. Studies were judged against pre-determined inclusion and exclusion criteria, and where suitable for inclusion, data extraction and quality assessment was achieved using a specifically-designed tool. All studies where prevalence of diabetes was investigated were eligible for inclusion. The inclusion criteria required that the study population be of a GCC country, but otherwise all ages, sexes and ethnicities were included, resident and migrant populations, urban and rural, of all socioeconomic and educational backgrounds. No limitations on publication type, publication status, study design or language of publication were imposed. However, we did not include secondary reports of data, such as review articles without novel data synthesis.

Conclusions

The prevalence ofT2DM is an increasing problem for all GCC states. They may therefore benefit to a relatively high degree from co-ordinated implementation of broadly consistent management strategies. Further study of prevalence in children and in national versus expatriate populations would also be useful.     

Enhancing protein crystallization through precipitant synergy

Suitable conditions for protein crystallization are commonly identified by screening combinations of independent factors that affect crystal formation. Because precipitating agents are prime determinants of crystallization, we investigated whether a systematic exploration of combinations of mechanistically distinct precipitants would enhance crystallization. A crystallization screen containing 64 precipitant mixtures was devised. Tests with ten HIV envelope-related proteins demonstrated that use of precipitant mixtures significantly enhanced both the probability of crystallization as well as the quality of optimized crystals. Tests with hen egg white lysozyme generated a novel C2 crystal from a salt/organic solvent mixture; structure solution at 2 A resolution revealed a lattice held together by both hydrophobic and electrostatic dyad interactions. The results indicate that mechanistically distinct precipitants can synergize, with precipitant combinations adding unique dimensions to protein crystallization.     

Comparative genomics of Helicobacter pylori isolates recovered from ulcer disease patients in England

Background  

Genomic diversity of H. pylori from many different human populations is largely unknown. We compared genomes of 65 H. pylori strains from Nottingham, England. Molecular analysis was carried out to identify rearrangements within and outside the cag-pathogenicity-island (cag PAI) and DNA sequence divergence in candidate genes. Phylogenetic analysis was carried out based on various high-resolution genotyping techniques.     

Molecular identification and phylogenetic analyses of multiple nucleopolyhedrovirus isolated from <Emphasis Type="Italic">Lymantria obfuscata</Emphasis> (Lepidoptera: Lymantriidae) in India

The Lymantria obfuscata Walker (Lyob) multiple (M) nucleopolyhedrovirus (NPV) (LyobMNPV) has been isolated and successfully applied for the management of the Indian gypsy moth, L. obfuscata in Jammu and Kashmir (J&K), India. The present work aimed to investigate the variability of LyobMNPV isolates from six localities of J&K through molecular [amplification of the polyhedrin (polh), late expression factor-8 (lef-8) and late expression factor-9 (lef-9) genes] and biological (bioassays) characterization. To identify the position of LyobMNPV in the phylogenetic tree of baculoviruses, partial sequences of the polh, lef-8 and lef-9 genes were determined by using the DNA sequences within their coding regions by optimizing the polymerase chain reaction with degenerate primers. The sequence alignment revealed that LyobMNPV isolates exhibited seven, five and eleven single nucleotide polymorphic sites in the case of polh, lef-8 and lef-9, respectively. The phylogenetic analyses supported placing LyobMNPV with the Lymantria dispar L. MNPV (LdMNPV) isolates from different countries, and showed that it was more closely related to LdMNPV than to Lymantria xylina Swinhoe NPV and Lymantria monacha L. NPV. The contaminated diet plug bioassays using 2nd instar larvae indicated that the median lethal dose (LD₅₀) and median survival time (ST₅₀) of different isolates of LyobMNPV against L. obfuscata were lower than those of LdMNPV against L. dispar. LyobMNPV was more closely related to LdMNPV but its LD₅₀ and ST₅₀ were lower than those of LdMNPV. The study provides novel information on the position of LyobMNPV in the phylogenetic tree of baculoviruses and about biological and genetic variation of Lymantria species’ NPV isolates from different parts of the world.     

Heavy metal detoxification mechanisms in halophytes: an overview

Heavy metals are among the major pollutants from anthropogenic inputs that reach mangrove ecosystem by urban and agricultural runoff, industrial effluents, boating, mining and other processes. To minimize the detrimental effects of heavy metal exposure and their accumulations, plants in general have evolved biological detoxification mechanisms, which include avoidance or exclusion, excretion and accumulation. To protect the cellular components from oxidative damage by heavy metal contamination, biological systems have developed enzymatic and non-enzymatic antioxidant mechanisms. Another detoxification mechanisms produced in plants are osmoprotectants, which are the compatible solutes which maintain a favourable water potential gradient and protect cellular structures from toxic ions. Besides these mechanisms, another heavy metal detoxification system in plants involves the chelation of metals by metal binding molecules like metallothioneins (MTs) and phytochelatins (PCs). To limit the heavy metal toxicity from mangrove ecosystem, it was found that phytoremediation is a most useful technology where in plants are used to remove pollutants from the environment and it is considered as a comparatively new, low-cost and highly promising technology for the remediation of heavy metal. Rhizofiltration, phytovolatilization, phytoextraction and phytostabilization are the important phytoremediation techniques. Among these phytoextraction and phytostabilization are found highly important in the case of mangroves and are promising means of phytoremediation.     

Clathrin promotes centrosome integrity in early mitosis through stabilization of centrosomal ch-TOG

Clathrin depletion by ribonucleic acid interference (RNAi) impairs mitotic spindle stability and cytokinesis. Depletion of several clathrin-associated proteins affects centrosome integrity, suggesting a further cell cycle function for clathrin. In this paper, we report that RNAi depletion of CHC17 (clathrin heavy chain 17) clathrin, but not the CHC22 clathrin isoform, induced centrosome amplification and multipolar spindles. To stage clathrin function within the cell cycle, a cell line expressing SNAP-tagged clathrin light chains was generated. Acute clathrin inactivation by chemical dimerization of the SNAP-tag during S phase caused reduction of both clathrin and ch-TOG (colonic, hepatic tumor overexpressed gene) at metaphase centrosomes, which became fragmented. This was phenocopied by treatment with Aurora A kinase inhibitor, suggesting a centrosomal role for the Aurora A-dependent complex of clathrin, ch-TOG, and TACC3 (transforming acidic coiled-coil protein 3). Clathrin inactivation in S phase also reduced total cellular levels of ch-TOG by metaphase. Live-cell imaging showed dynamic clathrin recruitment during centrosome maturation. Therefore, we propose that clathrin promotes centrosome maturation by stabilizing the microtubule-binding protein ch-TOG, defining a novel role for the clathrin-ch-TOG-TACC3 complex.