A cross-sectional study to compare levels of physical activity among adolescents in rural and urban areas of Western Maharashtra.

Background: Childhood and adolescent overweight and obesity has been associated with long term health deterioration. Lack of physical activity in this age group is an important contributing factor towards increase in overweight and obesity. Due to vast socio-cultural differences across India, it is important to assess the patterns of physical activity in different age groups and regions, to understand the differences among adolescents in different socio-geographic regions. However, the studies from India are inconsistent due to use of varied parameters and methodology and a need was felt of incorporating easy to use measures of quantifying physical activity. Methods: The study was carried out on 416 adolescents (aged 10 to 17 years) from 2 schools each from urban and rural area of Western Maharashtra. The study populations were stratified by gender and age. The data on physical activity was collected using a pre-tested, researcher-administered questionnaire adapted from the Global Physical Activity Questionnaire (GPAQ) and the Physical Activity Questionnaire- Adolescent (PAQ-A). Results: 16.8 percent of rural adolescents did not meet the defined criteria for adequate physical activity as compared to 33.2 percent in the urban population. Urban adolescent population was found to have significantly higher anthropometric measures of adiposity. Conclusion: Higher proportion of adolescents performing adequate physical activity is likely to be a key factor in keeping the BMI and TMI lower in rural as compared to urban adolescent population.

In Vivo Cross-Linking and Co-Immunoprecipitation Procedure to Analyze Nuclear tRNA Export Complexes in Yeast Cells.

tRNAs are small noncoding RNAs that are predominantly known for their roles in protein synthesis and also participate in numerous other functions ranging from retroviral replication to apoptosis. In eukaryotic cells, all tRNAs move bidirectionally, shuttling between the nucleus and the cytoplasm. Bidirectional nuclear-cytoplasmic tRNA trafficking requires a complex set of conserved proteins. Here, we describe an in vivo biochemical methodology in Saccharomyces cerevisiae to assess the ability of proteins implicated in tRNA nuclear export to form nuclear export complexes with tRNAs. This method employs tagged putative tRNA nuclear exporter proteins and co-immunoprecipitation of tRNA-exporter complexes using antibody-conjugated magnetic beads. Because the interaction between nuclear exporters and tRNAs may be transient, this methodology employs strategies to effectively trap tRNA-protein complexes in vivo. This pull-down method can be used to verify and characterize candidate proteins and their potential interactors implicated in tRNA nuclear-cytoplasmic trafficking.

Three tRNA nuclear exporters in S. cerevisiae: parallel pathways, preferences, and precision.

tRNAs that are transcribed in the nucleus are exported to the cytoplasm to perform their iterative essential function in translation. However, the complex set of tRNA post-transcriptional processing and subcellular trafficking steps are not completely understood. In particular, proteins involved in tRNA nuclear export remain unknown since the canonical tRNA nuclear exportin, Los1/Exportin-t, is unessential in all tested organisms. We previously reported that budding yeast Mex67-Mtr2, a mRNA nuclear exporter, co-functions with Los1 in tRNA nuclear export. Here we employed in vivo co-purification of tRNAs with endogenously expressed nuclear exporters to document that Crm1 also is a bona fide tRNA nuclear exporter. We document that Los1, Mex67-Mtr2 and Crm1 possess individual tRNA preferences for forming nuclear export complexes with members of the 10 families of intron-containing pre-tRNAs. Remarkably, Mex67-Mtr2, but not Los1 or Crm1, is error-prone, delivering tRNAs to the cytoplasm prior to 5' leader removal. tRNA retrograde nuclear import functions to monitor the aberrant leader-containing spliced tRNAs, returning them to the nucleus where they are degraded by 3' to 5' exonucleases. Overall, our work identifies a new tRNA nuclear exporter, uncovers exporter preferences for specific tRNA families, and documents contribution of tRNA nuclear import to tRNA quality control.

Efficacy of group intervention on tobacco cessation among male employees in health-care setting: A randomized controlled trial.

BACKGROUND: The tobacco epidemic is one of the biggest public health threats the world has ever faced. World Health Organization has estimated that tobacco use (smoking and smokeless) is currently responsible for the death of about 7 million people across the world each year. The objective of the study was not only to find the effect of group intervention on tobacco cessation but also to describe certain epidemiological factors associated with tobacco cessation and make suitable recommendations to tackle this epidemic. METHODS: A randomized controlled trial was carried out among male employees who were tobacco users in health-care setting in Western Maharashtra. In the study, 60 subjects each in intervention and control arm were taken. Pretested validated questionnaires were used for the study. The intervention comprised of two sessions delivered 5 weeks apart. Control arm received self-help material (Booklet) immediately after baseline data collection. The outcomes were measured using structured interview schedule. The data were analyzed using SPSS, version 20. RESULTS: Overall, 13.3% of the study subjects had quit tobacco use post intervention. In the intervention group 21.7% of the participants had quit tobacco since past one month and 5% in the control group (relative risk (RR) = 4.33). Low to moderate nicotine dependence (p = 0.023, RR = 6.46) and stage of contemplation (p = 0.018) were found to be important predictors of abstinence. CONCLUSION: Community-based group intervention for tobacco cessation is the way forward to tackle the tobacco epidemic.

tRNA dynamics between the nucleus, cytoplasm and mitochondrial surface: Location, location, location.

Although tRNAs participate in the essential function of protein translation in the cytoplasm, tRNA transcription and numerous processing steps occur in the nucleus. This subcellular separation between tRNA biogenesis and function requires that tRNAs be efficiently delivered to the cytoplasm in a step termed "primary tRNA nuclear export". Surprisingly, tRNA nuclear-cytoplasmic traffic is not unidirectional, but, rather, movement is bidirectional. Cytoplasmic tRNAs are imported back to the nucleus by the "tRNA retrograde nuclear import" step which is conserved from budding yeast to vertebrate cells and has been hijacked by viruses, such as HIV, for nuclear import of the viral reverse transcription complex in human cells. Under appropriate environmental conditions cytoplasmic tRNAs that have been imported into the nucleus return to the cytoplasm via the 3rd nuclear-cytoplasmic shuttling step termed "tRNA nuclear re-export", that again is conserved from budding yeast to vertebrate cells. We describe the 3 steps of tRNA nuclear-cytoplasmic movements and their regulation. There are multiple tRNA nuclear export and import pathways. The different tRNA nuclear exporters appear to possess substrate specificity leading to the tantalizing possibility that the cellular proteome may be regulated at the level of tRNA nuclear export. Moreover, in some organisms, such as budding yeast, the pre-tRNA splicing heterotetrameric endonuclease (SEN), which removes introns from pre-tRNAs, resides on the cytoplasmic surface of the mitochondria. Therefore, we also describe the localization of the SEN complex to mitochondria and splicing of pre-tRNA on mitochondria, which occurs prior to the participation of tRNAs in protein translation. This article is part of a Special Issue entitled: SI: Regulation of tRNA synthesis and modification in physiological conditions and disease edited by Dr. Boguta Magdalena.

Sharing the load: Mex67-Mtr2 cofunctions with Los1 in primary tRNA nuclear export.

Eukaryotic transfer RNAs (tRNAs) are exported from the nucleus, their site of synthesis, to the cytoplasm, their site of function for protein synthesis. The evolutionarily conserved ß-importin family member Los1 (Exportin-t) has been the only exporter known to execute nuclear export of newly transcribed intron-containing pre-tRNAs. Interestingly, LOS1 is unessential in all tested organisms. As tRNA nuclear export is essential, we previously interrogated the budding yeast proteome to identify candidates that function in tRNA nuclear export. Here, we provide molecular, genetic, cytological, and biochemical evidence that the Mex67-Mtr2 (TAP-p15) heterodimer, best characterized for its essential role in mRNA nuclear export, cofunctions with Los1 in tRNA nuclear export. Inactivation of Mex67 or Mtr2 leads to rapid accumulation of end-matured unspliced tRNAs in the nucleus. Remarkably, merely fivefold overexpression of Mex67-Mtr2 can substitute for Los1 in los1Δ cells. Moreover, in vivo coimmunoprecipitation assays with tagged Mex67 document that the Mex67 binds tRNAs. Our data also show that tRNA exporters surprisingly exhibit differential tRNA substrate preferences. The existence of multiple tRNA exporters, each with different tRNA preferences, may indicate that the proteome can be regulated by tRNA nuclear export. Thus, our data show that Mex67-Mtr2 functions in primary nuclear export for a subset of yeast tRNAs.

Association between intimate partner violence & HIV/AIDS: Exploring the pathways in Indian context.

BACKGROUND & OBJECTIVES: Violence against women cutting across diverse socio-economic classes is an under-recognized human rights violation in the world. This analysis was undertaken to examine the prevalence along with predictors of intimate partner violence (IPV) and its association with HIV/AIDS and sexually transmitted infections (STIs) in Indian ever-married women. METHODS: The data obtained from 2005 to 2006 third round of National Family Health Survey-3 (NFHS-3) were used in this study. Analyses were conducted on ever-married women by linking individual women data including violence information and HIV test results. RESULTS: The analyses indicated all forms of violence to be prevalent in India. The prevalence of lifetime IPV reported was 35.3 per cent. Multivariate analysis using logistic regression identified younger age of women, higher number of children, low level of education of women as well as her partner, working status of women, higher spousal age, rural residence, alcohol consumption by husband, childhood witness of violence among parents, nuclear household and lower standard of living to be positively associated with the experience of IPV by the women (P<0.05). HIV-positive status of women, as well as women from high HIV prevalent State, were at increased odds of IPV (P<0.05). INTERPRETATION & CONCLUSIONS: Significantly higher reporting of HIV/STIs by women experiencing IPV hints at new pathways that link violence and HIV. Further, our analysis showed a high prevalence of IPV in India.

Genome-wide screen uncovers novel pathways for tRNA processing and nuclear-cytoplasmic dynamics.

Transfer ribonucleic acids (tRNAs) are essential for protein synthesis. However, key gene products involved in tRNA biogenesis and subcellular movement remain to be discovered. We conducted the first comprehensive unbiased analysis of the role of nearly an entire proteome in tRNA biology and describe 162 novel and 12 previously known Saccharomyces cerevisiae gene products that function in tRNA processing, turnover, and subcellular movement. tRNA nuclear export is of particular interest because it is essential, but the known tRNA exporters (Los1 [exportin-t] and Msn5 [exportin-5]) are unessential. We report that mutations of CRM1 (Exportin-1), MEX67/MTR2 (TAP/p15), and five nucleoporins cause accumulation of unspliced tRNA, a hallmark of defective tRNA nuclear export. CRM1 mutation genetically interacts with los1Δ and causes altered tRNA nuclear-cytoplasmic distribution. The data implicate roles for the protein and mRNA nuclear export machineries in tRNA nuclear export. Mutations of genes encoding actin cytoskeleton components and mitochondrial outer membrane proteins also cause accumulation of unspliced tRNA, likely due to defective splicing on mitochondria. Additional gene products, such as chromatin modification enzymes, have unanticipated effects on pre-tRNA end processing. Thus, this genome-wide screen uncovered putative novel pathways for tRNA nuclear export and extensive links between tRNA biology and other aspects of cell physiology.

Role of forefinger and thumb loops in production of Ψ54 and Ψ55 in tRNAs by archaeal Pus10.

Pseudouridines (Ψ) are found in structurally and functionally important regions of RNAs. Six families of Ψ synthases, TruA, TruB, TruD, RsuA, RluA, and Pus10 have been identified. Pus10 is present in Archaea and Eukarya. While most archaeal Pus10 produce both tRNA Ψ54 and Ψ55, some produce only Ψ55. Interestingly, human PUS10 has been implicated in apoptosis and Crohn's and Celiac diseases. Homology models of archaeal Pus10 proteins based on the crystal structure of human PUS10 reveal that there are subtle structural differences in all of these Pus10 proteins. These observations suggest that structural changes in homologous proteins may lead to loss, gain, or change of their functions, warranting the need to study the structure-function relationship of these proteins. Using comparison of structural models and a series of mutations, we identified forefinger loop (reminiscent of that of RluA) and an Arg and a Tyr residue of archaeal Pus10 as critical determinants for its Ψ54, but not for its Ψ55 activity. We also found that a Leu residue, in addition to the catalytic Asp, is essential for both activities. Since forefinger loop is needed for both rRNA and tRNA Ψ synthase activities of RluA, but only for tRNA Ψ54 activity of Pus10, archaeal Pus10 proteins must use a different mechanism of recognition for Ψ55 activity. We propose that archaeal Pus10 uses two distinct mechanisms for substrate uridine recognition and binding. However, since we did not observe any mutation that affected only Ψ55 activity, both mechanisms for archaeal Pus10 activities must share some common features.

The archaeal COG1901/DUF358 SPOUT-methyltransferase members, together with pseudouridine synthase Pus10, catalyze the formation of 1-methylpseudouridine at position 54 of tRNA.

The methylation of pseudouridine (Ψ) at position 54 of tRNA, producing m(1)Ψ, is a hallmark of many archaeal species, but the specific methylase involved in the formation of this modification had yet to be characterized. A comparative genomics analysis had previously identified COG1901 (DUF358), part of the SPOUT superfamily, as a candidate for this missing methylase family. To test this prediction, the COG1901 encoding gene, HVO_1989, was deleted from the Haloferax volcanii genome. Analyses of modified base contents indicated that while m(1)Ψ was present in tRNA extracted from the wild-type strain, it was absent from tRNA extracted from the mutant strain. Expression of the gene encoding COG1901 from Halobacterium sp. NRC-1, VNG1980C, complemented the m(1)Ψ minus phenotype of the ΔHVO_1989 strain. This in vivo validation was extended with in vitro tests. Using the COG1901 recombinant enzyme from Methanocaldococcus jannaschii (Mj1640), purified enzyme Pus10 from M. jannaschii and full-size tRNA transcripts or TΨ-arm (17-mer) fragments as substrates, the sequential pathway of m(1)Ψ54 formation in Archaea was reconstituted. The methylation reaction is AdoMet dependent. The efficiency of the methylase reaction depended on the identity of the residue at position 55 of the TΨ-loop. The presence of Ψ55 allowed the efficient conversion of Ψ54 to m(1)Ψ54, whereas in the presence of C55, the reaction was rather inefficient and no methylation reaction occurred if a purine was present at this position. These results led to renaming the Archaeal COG1901 members as TrmY proteins.

Pseudouridine formation in archaeal RNAs: The case of Haloferax volcanii.

Pseudouridine (Ψ), the isomer of uridine, is commonly found at various positions of noncoding RNAs of all organisms. Ψ residues are formed by a number of single- or multisite specific Ψ synthases, which generally act as stand-alone proteins. In addition, in Eukarya and Archaea, specific ribonucleoprotein complexes, each containing a distinct box H/ACA guide RNA and four core proteins, can produce Ψ at many sites of different cellular RNAs. Cbf5 is the core Ψ synthase in these complexes. Using Haloferax volcanii as an archaeal model organism, we show that, contrary to eukaryotes, the Cbf5 homolog (HVO_2493) is not essential in this archaeon. The Cbf5-deleted strain of H. volcanii completely lacks Ψ at positions 1940, 1942, 2605, and 2591 (Escherichia coli positions 1915, 1917, 2572, and 2586) of its 23S rRNA, and contains reduced steady-state levels of some box H/ACA RNAs. Archaeal Cbf5 is known to have tRNA Ψ55 synthase activity in vitro but we could not confirm this activity in vivo in H. volcanii. Conversely, the Pus10 (previously PsuX) homolog (HVO_1979), which can produce tRNA Ψ55, as well as Ψ54 in vitro, is shown here to be essential in H. volcanii, whereas the corresponding tRNA Ψ55 synthases, Pus4 and TruB, are not essential in yeast and E. coli, respectively. Finally, we demonstrate that HVO_1852, the TruA/Pus3 homolog, is responsible for the pseudouridylation of position 39 in H. volcanii tRNAs and that the corresponding gene is not essential.

Bellary, India achieves negligible case fatality due to Japanese encephalitis despite no vaccination: an outbreak investigation in 2004.

OBJECTIVE: To confirm the existence of the outbreak of suspected Japanese encephalitis, identify the source, to understand the circumstances due to which the outbreak was taking place and to suggest measures for its control. METHODS: The team visited Bellary from 4th to 10th Sept, 2004. The team interviewed the key persons and analyzed the records at District Surveillance Unit and Entomological Surveillance Unit and case records of suspected JE cases admitted in Encephalitis ward in Vijay Nagar Institute of Medical Sciences (VIMS). Eco-entomological survey was done in houses and surroundings of 3 randomly selected cases of Encephalitis in rural and urban areas of District Bellary. Their family members and neighbors were also asked for the awareness and presence of disease. Data was analyzed for epidemiological and clinical profiles. RESULTS: The suspected JE cases were being reported from end of June 2004. The cases were sporadic and out of 34 cases reported to VIMS (till 10th of September), 32 were from Bellary district and 2 were from adjoining Andhra Pradesh. Among these 32, 22 were from Bellary Taluk, which in turn were mainly concentrated (10 were reported) in urban Bellary. The case fatality rate was zero as no death was reported. Entomological surveillance (done by District Surveillance Unit) revealed a high outdoor presence of Culex tritaeniorhynchus as well as an indoor rising density of this mosquito from 2 per man hour catch in January to 22 in the month of August in the affected villages. On the contrary, the investigations on 7th and 8th September revealed high densities of An.subpictus and An. peditaenatus and nil of Culex species in the urban areas. Amplifier host of pigs and water birds were occasionally sighted in the area. CONCLUSION: A good community awareness of encephalitis, a prompt referral system and a good supportive treatment for the patients and a good surveillance system and response were observed. Very close proximity with amplifying hosts of pigs was avoided by the community, though piggeries were still not very far away (1-3 Km). These may explain the reduction in cases, deaths and disabilities due to this disease in this district over the years. Possibilities of mutant strain which is less virulent and/or a better immune status of at risk population may also need to be explored. The impact of the mass vaccination with SA 14-14-2, imported from China in Bellary during July, 2006 remains to be evaluated. This will further decrease the case load.