Master of Science

thesisRSC is a multiprotein complex that remodels the structure of chromatin and in turn regulates processes like transcription. The mechanism by which RSC remodels a nucleosome is still unclear. Although bulk solution experiments have added a growing body of evidence to the mechanism by which RSC remodels a nucleosome, the mechanistic properties of the remodeler is still under debate. In this work, we have constructed a system to study the mechanistic properties of a single molecule of RSC on a naked DNA template in real time and obtain its properties like rate of translocation, processivity, force generated by the translocase and rate of ATP turnover. The SWI/SNF family of chromatin remodeling complexes contains conserved Actin-related proteins (ARPs). Genetic and biochemical evidence suggests that Sth1 might be regulated by two Actin-Related Proteins, ARP7 and ARP9. Genome wide suppressors of ARPs mapped to two domains in Sth1 called the post-HSA (Helicase/ Sant associated) domain and ATPase domain. In this work, dominant negatives of Sth1 were obtained that also localized to the post-HSA (Helicase/ Sant associated) domain. This suggests that the post-HSA domain might be involved in the regulation of Sth1. Sth1 (301-1097) construct containing the suppressor mutations co-purified with ARPs were compared for their ATPase activity and KM with wild type Sth1 (301-1097) Arps complex. The mutants showed higher ATPase activity than wild type, which was not due to their increased affinity for DNA. iv Based on the fact that the loop formed by the action of a remodeler is negatively supercoiled, in this work, we also developed an assay to assess the torsion generated by ATP dependent chromatin remodelers on naked DNA. TetR-Sth1 (301-1146) fusion co-purified with ARPs successfully produced torsion in an ATP dependent manner as assayed by this assay. This assay also depends on the action of by E. coli topoisomerase I, which specifically relaxes negative supercoils and on TetR-TetO interaction which serves to anchor the motor protein and thus constrains the loop formed by it

Condom tamponade in the management of atonic postpartum hemorrhage

Background: Postpartum hemorrhage (PPH) remains a leading direct cause of maternal death in both developed and developing countries. Millennium development goal of reducing maternal mortality rate cannot be achieved unless the prevention and treatment of PPH is prioritized. Objectives of the study were to study the efficacy of condom tamponade to arrest bleeding in cases of atonic PPH unresponsive to uterotonics.Methods: Prospective study conducted in the Kottayam Medical College where mothers who develop atonic PPH following childbirth unresponsive to uterotonics (after ruling out traumatic PPH) were included in study. Condom tamponade was applied in them and inflated with normal saline until the bleeding stops. Intrauterine drain was fitted with this to find out ongoing bleeding with tamponade in situ. The Condom tamponade was kept for 12-24 hrs and gradually deflated when bleeding ceased. Outcome measures were (1) Ability of condom catheter to stop bleeding; (2) Cases requiring further intervention; (3) Time required to stop bleeding and (4) Subsequent morbidity in terms of infection.Results: Of the 487 cases of atonic PPH, 61 required condom tamponade; bleeding controlled in 59 cases (96.7%); two patients (3.3%) needed further surgical intervention. No cases of maternal mortality due to PPH. No clinical evidence of intrauterine infection.Conclusions: The hydrostatic condom tamponade controls PPH quickly and effectively. It is simple, inexpensive, easily available, needs less expertise and life saving. Any healthcare provider may use this procedure as a timely measure to save the life of patients especially during referral to decrease ongoing loss

Allosteric interactions of DNA and nucleotides with S. cerevisiae RSC

RSC, Remodel the Structure of Chromatin, is an essential chromatin remodeler of Saccharomyces cerevisiae that has been shown to have DNA translocase properties. We studied the DNA binding properties of a ‘trimeric minimal RSC’ (RSCt) of the RSC chromatin remodeling complex and the effect of nucleotides on this interaction using fluorescence anisotropy. RSCt binds to 20 bp fluorescein labeled double stranded DNA with a Kd of approximately 100 nM. The affinity of RSCt for DNA is reduced in the presence of AMP-PNP and ADP in a concentration dependent manner with the addition of AMP-PNP having the more pronounced effect. These differences in the magnitude at which the binding of ADP and AMP-PNP affect the affinity of DNA binding by RSCt suggests that the physical movement of the enzyme along DNA begins between the binding of ATP and its subsequent hydrolysis. Furthermore, the fact that the highest affinity for DNA binding by RSCt occurs in the absence of bound nucleotide offers a mechanistic explanation for the low apparent processivity of DNA translocation by the enzyme.This work was supported by National Institutes of Health Grant P20 RR017708 to C.J.F. and NIH GM60415 to B.R.C. Evan Rich was supported, in part, by an Undergraduate Research Award provided by the Kansas University Honors Program

Computerized music-based intervention module for auditory processing and working memory in older adults

IntroductionThe contribution of technology to the field of health is vast, both in diagnosis and management. More so, the use of computer-based intervention has become increasingly widespread over the past decade. Human beings experience a decline in auditory processing and cognitive skills as they age, consistent with deterioration of other bodily functions. In addition, speech perception abilities in both quiet and in the presence of noise are impacted by auditory processing abilities and cognitive skills such as working memory. This pilot study explored the use of music as an intervention for improving these skills and employed a computerized delivery of the intervention module.MethodA battery of tests was carried out to assess the baseline auditory processing and working memory skills in eight older adults between the ages of 56 and 79 years, all of whom had normal hearing. Following the assessment, a short-term computerized music-based intervention was administered. The style of music chosen was Carnatic classical music, a genre widely practiced in Southern India. The intervention module involved note and tempo discrimination and was carried out for a maximum of 10 half-hour sessions. The multi-level intervention module was constructed and administered using Apex software. Following the intervention, the auditory processing and cognitive skills of the participants were reassessed to study any changes in their auditory processing and working memory skills.Results and discussionThere were positive changes observed in all the auditory processing and some of the working memory abilities. This paper discusses in detail the systematic structuring of the computerized music-based intervention module and its effects on the auditory processing and cognitive skills in older adults

Interaction between TCF7L2 polymorphism and dietary fat intake on high density lipoprotein cholesterol

Recent evidence suggests that lifestyle factors influence the association between the Melanocortin 4 receptor (MC4R) and Transcription Factor 7-Like 2 (TCF7L2) gene variants and cardio-metabolic traits in several populations; however, the available research is limited among the Asian Indian population. Hence, the present study examined whether the association between the MC4R single nucleotide polymorphism (SNP) (rs17782313) and two SNPs of the TCF7L2 gene (rs12255372 and rs7903146) and cardio-metabolic traits is modified by dietary factors and physical activity. This cross sectional study included a random sample of normal glucose tolerant (NGT) (n = 821) and participants with type 2 diabetes (T2D) (n = 861) recruited from the urban part of the Chennai Urban Rural Epidemiology Study (CURES). A validated food frequency questionnaire (FFQ) was used for dietary assessment and self-reported physical activity measures were collected. The threshold for significance was set at P = 0.00023 based on Bonferroni correction for multiple testing [(0.05/210 (3 SNPs x 14 outcomes x 5 lifestyle factors)]. After Bonferroni correction, there was a significant interaction between the TCF7L2 rs12255372 SNP and fat intake (g/day) (Pinteraction = 0.0001) on high-density lipoprotein cholesterol (HDL-C), where the 'T' allele carriers in the lowest tertile of total fat intake had higher HDL-C (P = 0.008) and those in the highest tertile (P = 0.017) had lower HDL-C compared to the GG homozygotes. In a secondary analysis of SNPs with the subtypes of fat, there was also a significant interaction between the SNP rs12255372 and polyunsaturated fatty acids (PUFA, g/day) (Pinteraction<0.0001) on HDL-C, where the minor allele carriers had higher HDL-C in the lowest PUFA tertile (P = 0.024) and those in the highest PUFA tertile had lower HDL-C (P = 0.028) than GG homozygotes. In addition, a significant interaction was also seen between TCF7L2 SNP rs12255372 and fibre intake (g/day) on HDL-C (Pinteraction<0.0001). None of the other interactions between the SNPs and lifestyle factors were statistically significant after correction for multiple testing. Our findings indicate that the association between TCF7L2 SNP rs12255372 and HDL-C may be modified by dietary fat intake in this Asian Indian population

Circulating adiponectin mediates the association between omentin gene polymorphism and cardiometabolic health in Asian Indians

Background: Plasma omentin levels have been shown to be associated with circulating adiponectin concentrations and cardiometabolic disease-related outcomes. In this study, we aim to examine the association of omentin gene polymorphism with serum adiponectin levels and cardiometabolic health status using a genetic approach and investigate whether these associations are modified by lifestyle factors. Methods: The study included 945 normal glucose tolerant and 941 unrelated individuals with type 2 diabetes randomly selected from the Chennai Urban Rural Epidemiology Study (CURES), in southern India. Study participants were classified into cardiometabolically healthy and unhealthy, where cardiometabolically healthy were those without hypertension, diabetes, and dyslipidemia. Fasting serum adiponectin levels were measured by radioimmunoassay. The omentin A326T (rs2274907) single nucleotide polymorphism (SNP) was screened by polymerase chain reaction-restriction fragment length polymorphism and direct sequencing. Results: The ‘A’ allele of the omentin SNP was significantly associated with lower adiponectin concentrations after adjusting for age, sex, body mass index (BMI), waist circumference (WC) and cardiometabolic health status (p=1.90 x 10-47). There was also a significant association between circulating adiponectin concentrations and cardiometabolic health status after adjusting for age, sex, BMI, WC and Omentin SNP (p=7.47x10-10). However, after adjusting for age, sex, BMI, WC and adiponectin levels, the association of ‘A’ allele with cardiometabolic health status disappeared (p=0.79) suggesting that adiponectin serves as a mediator of the association between omentin SNP and cardiometabolic health status. There were no significant interactions between the SNP and dietary factors on adiponectin levels and cardiometabolic health status (p>0.25, for all comparisons). Conclusions: Our findings show that adiponectin might function as a mechanistic link between omentin SNP and increased risk of cardiometabolic diseases independent of common and central obesity in Asian Indians. Further studies are required to confirm the molecular mechanisms involved in this triangular relationship between omentin gene, adiponectin and cardiometabolic diseases

Interaction between FTO gene variants and lifestyle factors on metabolic traits in an Asian Indian population

Background Lifestyle factors such as diet and physical activity have been shown to modify the association between fat mass and obesity–associated (FTO) gene variants and metabolic traits in several populations; however, there are no gene-lifestyle interaction studies, to date, among Asian Indians living in India. In this study, we examined whether dietary factors and physical activity modified the association between two FTO single nucleotide polymorphisms (rs8050136 and rs11076023) (SNPs) and obesity traits and type 2 diabetes (T2D). Methods The study included 734 unrelated T2D and 884 normal glucose-tolerant (NGT) participants randomly selected from the urban component of the Chennai Urban Rural Epidemiology Study (CURES). Dietary intakes were assessed using a validated interviewer administered semi-quantitative food frequency questionnaire (FFQ). Physical activity was based upon the self-report. Interaction analyses were performed by including the interaction terms in the linear/logistic regression model. Results There was a significant interaction between SNP rs8050136 and carbohydrate intake (% energy) (Pinteraction = 0.04), where the ‘A’ allele carriers had 2.46 times increased risk of obesity than those with ‘CC’ genotype (P = 3.0 × 10−5) among individuals in the highest tertile of carbohydrate intake (% energy, 71 %). A significant interaction was also observed between SNP rs11076023 and dietary fibre intake (Pinteraction = 0.0008), where individuals with AA genotype who are in the 3rd tertile of dietary fibre intake had 1.62 cm lower waist circumference than those with ‘T’ allele carriers (P = 0.02). Furthermore, among those who were physically inactive, the ‘A’ allele carriers of the SNP rs8050136 had 1.89 times increased risk of obesity than those with ‘CC’ genotype (P = 4.0 × 10−5). Conclusions This is the first study to provide evidence for a gene-diet and gene-physical activity interaction on obesity and T2D in an Asian Indian population. Our findings suggest that the association between FTO SNPs and obesity might be influenced by carbohydrate and dietary fibre intake and physical inactivity. Further understanding of how FTO gene influences obesity and T2D through dietary and exercise interventions is warranted to advance the development of behavioral intervention and personalised lifestyle strategies, which could reduce the risk of metabolic diseases in this Asian Indian population

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

DEVELOPMENT OF RESTRICTION ENZYME-BASED ASSAYS FOR CONCURRENT DETECTION OF MULTIPLE CYTOSINE MODIFICATIONS AND GENOMIC ABERRATIONS

Ph.DDOCTOR OF PHILOSOPHY (FOE

An Enhanced Approach for Character Recognition Based On Discriminative Semi-MarkovModel

Abstract- A methodology for extracting text from Video obtaining from frames, Text that appears in these images contains important and useful information. Text extraction in images has been used in large variety of applications such as mobile robot navigation, document retrieving, object identification, detection the vehicle license plate, etc. In this paper, we employ discrete wavelet transform (DWT) for extracting text information from complex images. The input image may be a color image or a grayscale image. If the image is color image, then preprocessing is required. For extracting text edges, the sobel edge detector is applied on each sub image. The resultant edges so obtained are used to form an edge map. Morphological operations are applied on the processed edge map and further thresholding is applied to improve the performance. Keywords-- Edge detector, discrete wavelets transform (DWT), Image segmentation, Text extraction and Text recognition. I