Annual Report 2005-2006

Science & Engineering Research Council

Life Sciences

Animal Sciences

• Four projects were sanctioned to unravel the mysteries of biological rhythms such as, Molecular mechanisms involved in neural regulation of pacemaker entrainment (biological clock) in a diurnal rodent; Role of food in regulation of circadian and seasonal responses in birds; Role of melatonin in regulation of clock genes; and Circadian physiology of the high altitude himalayan strains of drosophila;
• Two projects were sanctioned to study the behavioural ecology of primates viz, Niche separation in sympatric primates in the rainforests of the northern western ghats; and Demography and behavioural ecology of the Indian Bonnet Macaque (Macacradiata): a comparative account of the two subspecies;
• In Biodiversity, projects such as, Extremophilic free living ciliates (Protozoa, Ciliophora) from biodiversity hotspots for the Indian subcontinent; Screening of indigenous species of Trichogramma westwood & Trichogrammatoidea Girault (Hymenoptera : Trichogrammatidae) from central India and their utilization against important forest insect pests; Litter insect diversity and trophic interactions in relation to the substrate biochemical profile in the major forest vegetations of Kerala, were sanctioned.
• Other projects sanctioned included Development of a protocol for the restoration of a fish species using its post-mortem preserved sperm and genome inactivated eggs of intra and interspecific females; Reproductive effects of deltamethrin in the fish, Labeo rohita; Nitrifying bioreactors integrated into prawn hatchery systems: Biofilm structure, population dynamics and kinetic modeling; Biology, population dynamics and nutritional values of Bosmina Tripurae: a new candidate of micro crustacea in aquaculture; Cryobiology and nutritional quality of rotifers (Brachionus plicatilis and Brachionus calyciflorus); Gastro-intestinal nutritional grouping of microbes integrated with carotinoid pigments on growth kinetics and colouration of ornamental fishes; In vivo effect of vitellogenesis on the gonadotropin and estradiol-17B in the catfish, Clarias batrachus, etc.
• Toxicology studies launched include, Tissue concentrations of pesticide and its impact on reproductive potential in food fishes of river Ganga, Gomti and Sai of Eastern Uttar pradesh; Investigations on the mechanisms of success of Mytilopsis sallei (Recluz) in managing toxic load arising out of biodeterioration control measures; Antioxidants as modulators of oxidative stress and oxidative signals in mammalian erythrocyte; Isolation of bioactive molecules from selected plants against the armyworm Spodoptera litura (Fab.) and cotton bollworm, Helicoverpa armigera (Hub.,) (Lepidoptera : Noctuidae);
• In Reproductive endocrinology projects were launched to study, Immune system of sperm autoantigens by epididymal epithelium; The function and molecular regulation of schwann cell insulin receptors; LH/HCG receptor gene expression in mouse and human uterus and its relevance in protein, steroid hormones and prostaglandin production; Mechanisms related to oxidant-mediated testicular mitochondrial dysfunctions; Application of parthenogenetic engineering in the development of superior breeds/hybrids of the mulberry silkworm, Bombyx mori L.

Research highlights of ongoing projects are as follows:

The amphibian diversity of Dibrusaikhowa National Park, Assam has been compiled and compared with the adjacent mainland forest areas in order to study the effect of flood and change in vegetation of the park. Some new species, such as Bufo melanostictus and Rana humeralis have been added to the amphibian fauna of India.

The projects related to endocrinology has reported effect of thyroid hormones on lipid peroxidation and antioxidant enzyme activities in fish that has revealed rapid action of tri-iodothyronine (T3) in vivo and in vitro on antioxidant enzyme activities in teleost Anabas testudineus. It has been indicated for the first time that the muscarinic M1 receptors are also involved in regulation of arylalkaline Nacetyltransferace (AA-NAT) activity in the photoreceptive pineal of the fish mainly via both A1and Badrenergic receptors.

Electronic micro-motility meter has been fabricated with spectacular advancement and automation to record the motility of the parasites in vitro. The pharmacodynamic effect of azadirachtin, A.concinna. A. Arabica, C.fistula and A.sativul have shown that these plant resources are the effective antihelminthic drugs and could be used to eradicate paramphistome infection in livestock. An important microbial assay has been developed for the detection of Chloramphenicol (CAP) residues in shrimp muscle using luminescent bacteria, Photobacterium leiognathi with a minimum detection limit of 1 ng/ml (1 ppb).

An interaction meeting in animal physiology was organized to generate high quality proposals in the area.

Health Sciences

Support to Bio-Medical Sciences to strengthen the knowledge, generate baseline data, develop newer diagnostic methods, devices, processes, and drug delivery systems, study mechanism of action, to develop manpower and motivate them to engage in R&D activities in emerging and frontline areas was continued. The new projects were sanctioned for generation of DNA and peptide based vaccines, to study the genetic basis of diseases, cell cycle regulation and apoptosis, use of molecular biology techniques to understand patho-physiology of non-communicable diseases - CVD, cancer, etc., identifying new targets for drug delivery, identification, characterization of immunomodulating antitumor/ anti-diabetic/ antioxidant/ hypocholesteriemic/anticancer potential of plant products and development of drug molecules using genomics/proteomics.

With the increasing occurrence of cancer in the country, several new projects have been sanctioned to study various face of the disease e.g. tumor cell apoptosis via modulation of protein kinase C and supression of telomerase by natural polyphenols and isothiocyanates; Galectins and dietary galectininhibitors in tumor spread/metastasis and prevention; Genetic polymorphism of drug metabolizing enzymes and risk of urological and gastroenterological cancers; Targeted inhibition of cell cycle phase specific signalling molecules to induce selective apoptosis in cancer cells; Assessment of caveolin-1 mutations as possible risk factors in development of human breast carcinoma.

Life-style diseases like diabetes, cardio-vascular disorder, obesity, bone-related disorders, nephrology/urology, reproductive biology, ageing, neurological disorder, etc. are increasing. New projects in the area of non-communicable diseases e.g. Development & evaluation of phytosomes - a value added herbal drug delivery system; Expression of glial specific genes and their role in development and aging of the brain of mice; Molecular genetic basis of hypercalciuric nephrolihiasis in northern India; Rapid diagnosis of myocardial cell damage by a novel radiotracer; Development of anti-ulcer herbal formulation; Phenotypic and functional characterization of T cells in synovial fluid of patients with juvenile rheumatoid arthritis; Biossay development of quality control and standardization of an ayurvedic drug, abhrak bhasmas; Mucoadhesive buccal drug delivery for synthetic and peptide drugs: A promising option to conventional therapy; Precision grip as tool for quantification of sensory motor deficit in patients with spasticity; Porin of Shigella dysenteriae-mediated regulation of Toll-Llike receptor and costimulatory molecule of immunoglobolin and interleukin; Biochemical markers for the early diagnosis of osteoporosis were also sanctioned.

Pathogen based diseases are a major health hazard. Studies have been initiated to understand Immunological evaluation of non-porin outer membrane protein of Salmonella for protection against experimental salmonellosis and their potential for development of a candidate vacinne against typhoid; Liposol momensin in malaria chemotherapy; Identification of Mycobacterial tuberculosis by in-situ hybridisation and demostration of mycobacterial antigen in the CSF cytospin smears by an immunocytochemical for the early laboratory diagnosis of tuberculous meningitis (TMB); Understanding mechanisms of drug resistance in Vibrio cholerae; Immunochemical characterization of filarial glutathione-s-transferase and its protective potential in experimental filariasis

Environment plays a vital role in health management. Projects are sanctioned to study Development of a therapy for reversal of harmful effects of environmental toxic chemicals on the respiratory systems; Studies on the screening of chromosome abnormalities of blood lymphocytes of occupationally exposed textile dye unit workers; Study on the mechanism of hepatic damage with the degree of arsenic exposure; Role of intracranial self-stimulation and enriched environmental in the amelioration of stress induced cognitive deficits; Characterization of repetitive environmental stress conditions: Role of antioxidant defense, heat shock proteins and signal transduction network.

Major Research Highlights and Achievements

The role of integrin 􊼟 in regulation of MMP-2 activity may be of immense importance in understanding integrin-MMP inter-relationship and in clinical management of invasive potential of tumor cells according to the project results of integrin receptors role in invasive potential of tumor cells. The evaluation of mutation and expression of FHIT gene in Indian female breast cancer patients showed a change in exon 8 of the gene due to a point mutation (C T) at -98 codon. In a molecular genetic study on the prevalence of Hypercalcinuric nephrolithiasis, it was found to be more in North Indian population. The autosomal mode of inheritance was found to be most predominant. A study was carried out for the first time in Asian Indian children with congenital uropathies to determine whether I/ D polymorphism in the ACE gene are predictive of renal parenchymal damage. The ACE I/D genotype were found to be predictive of renal parenchymal damage independent of other known risk factors. Renin angiotensin System has a central role in the development of end-stage renal disease. Plasma Renin Activity (PRA) studies provide evidence that there is ongoing renal damage in patients with Posterior Urethral Valve (PUVs) even after valve ablation. Plasma renin activity (PRA) is one such marker that reflects the beginning of RAS activation, and thus, early recognition of children with ongoing renal damage. Decrease in GFR is considered to be the most sensitive marker of early renal damage. The findings of the study on chromosomal and molecular genetic evaluation of infertile man revealed that the genes on p arm of chromosome 13 have association with the intellectual ability and fertility status. Many autosomal genes are also involved in testicular development and spermatogenesis apart from the Y chromosomal genes. Studies are under progress for developing of anti-angiogenic therapy in retinal neo-vascularisation. In a study on molecular mechanism of thyroid hormone-induced cardiac hypertrophy an impairment of glucose transport in hyperthyroid cardiomyocetes was found, meanwhile the SOD, collagens, cytochrome P450 isoforms, matrix metalloproteins, etc., showing some new aspects of hyperthyroid heart to assess the hypertrophied cardiomycetes. The study on spatial and temporal sequestered sex and species-specific transcript from the spermatogenesis pathway of mouse lead to the isolation of the new cDNA from the heterochromatin long arm of the Y chromosome. Studies also found that the content of lamin decreasing during diabetes and ameliorated by feeding dietary fibers like wheat braun and gaur gum. The ultra-structural studies in type II diabetes for amyloid formation suspects that amyloid fibrils at various stages may affect the cell function in terms of insulin secretion.

The projects on communicable diseases have come out with various leads such as :-The lectins in invertebrates and in mosquitoes exposed to bio-larvicides might have been enhanced probably functioning as humoral immunoglobulins or binding agents in the gut membrane inhibiting microfilarial migration to the thoracic muscles; The priming of BALB/C neonates with a low dose of irradiated Leishmania major promastigotes establishes an age and IL-12 dependent host-protective Th1 response; Regarding the development of novel anti-HIV molecules, 10 organo-metallic ruthenium drugs are synthesized and screened for their activity, one involves inhibitory action of drug on soluble Topoisomerase II containing both isoforms and other to monitor isoform-specific inhibition activity of the drug; The HEV replicase and helicase-RdRp fusion constructs confirmed localization in membranes of endoplasmic reticulum and membrane bound vesicles respectively. The HEV replicase domain is found to start from 3913 nt position and 3511.

Plant Sciences

New projects funded during the year were: Characterization of Cyanobacterial Phycobiliproteins for Potential Applications, Biochemical & Molecular Basis of Cyanobacterial Adaptation to UV-B and Copper, Finger printing of Cyanobacterial strains Isolated from North Eastern Region Shillong, Diversity of Chrysophytes in fresh water Aquatic bodies of India, Microbial function and Diversity in Biogeochemical Cycling of Iron, Impact of industrial pollution on microbial diversity in region around Narmada estuary of Gujarat, Fungal Diversity and leaf litter decomposition in the Running fresh water bodies of Kumaun Himalaya, Community structure and dynamics of wood-rotting Basidiomycetes (Fungi) on decomposing trunks in temperature forests, Correlating the commensal attributes of rhizobacteria to plant growth promotion, Studies on the Photometabolism of anilines by Rhodobacter sphaeroids OU5: A detoxification process coupled to production of value added indoles, Ecology of reduced tillage and mulcing in the Central Himalayan cropfield, Conservation strategies and pharmacological evaluation of Utleria salicifolia an endangered ethonomedicinal plant of Western Ghats, Reproductive Biology of Cardamoms & Amomum, Screening of Aloe vera L. germplasms for cosmetic gel and micropropagation of elite clones, Incorporation of fungal disease resistance in Indian mustard (Brassica juncea [L.] Czern and Coss) through in vitro techniques, A molecular study of cytokinin signal transduction pathway in cucumber seedling towards understanding the downstream elements, Herbicidal property of invasive and noxious weed lantana (Lantana camara L.) constituents, Study of interaction between excitation energy (high light) oxidant stress and protective mechanisms on photosynthesis and productivity in rice seedings, Regulation of chlorophy II Biosynthesis in higher plants, Molecular approaches in understanding the organization of novel chromosome etc.

Some important results in ongoing projects were as follows :

In a project to understand light perception and signal transduction mechanisms operative in plants, Arabidopsis has been used as a model system. Arabidopsis seedling development follows two distinct pathways, i.e. skotomorphogenesis (in dark) and photomorphogenesis (in light). In the study a bHLH transcription factor (ZBF1/MYC2) has been identified and functionally characterized that acts as a negative regulator of blue light mediated photomorphogenesis (Figure 2.4). The promoter deletion analysis in Arabidopsis transgenics revealed that light and ethylene interact to regulate the spatial expression of HLS1/PHO1 through cis-regulatory elements, LRE and ERE, respectively, residing in the upstream promoter region.

Figure 2.4: The role of ZBF1/MYC2 as a negative regulator in blue light mediated seedling growth in Arabidopsis. The seedlings were grown for six days in dark (A), red (B), blue (C) or far-infrared light (D). In each panel, the wild type and zbf1 mutant are shown on the left and right respectively. It can be noted that blue light has a more dramatic effect on seedling growth inhibition in the zbf1 mutant in panel C

In another project, in the area of induced systemic resistance (ISR) against viruses, it was observed that a susceptible plant can be induced to resist virus infections through the application of proteins of plant origin, both at site of treatment and at untreated remote sites. CIP-29 is one such resistance inducer protein isolated from Clerodendrum inerme, treatment with which causes the synthesis/ activation of a new virus inhibitory agent (VIA), that appears to be directly antiviral. Purified VIA was also antiviral (Figure 2.5 and 2.6).

Figure 2.5: Systemic resistance induction by CIP-29		Figure 2.6: Single peak of VIA activity on sizeexclusion chromatography

In another project, a number of plant growth promoting rhizobacterial strains were studied of which Pseudomonas strains RRLJ 134 and RRLJ 04 were selected as potential strains through in vitro antibiosis study and field experiments. Application of RRLJ 134 enhanced growth of some cole crops and RRLJ 04 improved plant growth of pigeon pea under gnotobiotic and field condition besides suppressing wilt disease caused by Fusarium udum. Two fractions namely, diethyl ether and chloroform fractions, were extracted. Under gnotobiotic condition, respective fractions showed growth promoting and disease suppressing activity against fusarial wilt of pigeon pea. Bioactive fractions were further purified by thin layer chromatography and 6 compounds, 3 each from respective fractions were separated viz., E1, E2 & E3 (diethyl ether) and C1, C2 & C3 (chloroform) of which C1, C3 and E1 were found to be phenazine analogues. C1, C3 & E1 showed in vitro antibiosis against F. udum (Figure 2.7). Structural elucidation of C1 has been done and found to be a phenazine derivative.

Yellow fraction (E 1)                                                      Yellow fraction (C 1)
Figure2.7: In-vitro antibiosis of identified bioactive fractions from RRLJ 04 (25mg/100ml methanol)

In one of the ongoing project on "Biological control of a wilt disease complex of pigeon pea (caused by Heterodera cajani, Meloidogyne sp. and Furarium udum) by fluorescent Pseudomonads and Bacillus Spp", sixty isolates of Bacillus and fluorescent Pseudomonads were isolated and characterized using Bergey's Manual of Determinative Bacteriology. In a project on Botanical pesticides the mixture of flavonoids isolated from leaves of C. procera showed excellent adulticidal, oviposition deterrent and ovicidal action against pulse beetle, C. chinensis and was also found to affect the longevity, life cycle duration and weight both of male and female insects.

Biochemistry, Biophysics, Molecular Biology and Microbiology

New projects sanctioned were:

Mechanism of action of anaphase promoting complex in gene silencing fission yeast; Chromatin transcription of some unusual tRNA genes of the yeast Saccharomyces cerevisiae; Characterization of the positive regulatory region of the first exon of human cathepsin L gene; Xylitol production by candida sp; A study on the transforming growth factor-B signalling intermediate, Smad3 in human cervical cancer; Functional studies on Escherichia Coli PepN, a metallo aminoendopeptidase involved in cytosolic protein degradation; Regulation of platelet cytoskeletal organisation by Rho family GTPases and associated molecules; Studies on isolation and mechanism of action of the antihyperglycemic and hypolipidemic compound (s) from Cassia auriculata leaf extract; Role of two proteins in mitrochondrial tRNA import; Inhibition of Gene expression in HIV-1 using peptides libraries and amino acid analogues; Cyclic peptides as proteinase inhibitors: purification, molecular cloning and expression; In vitro and in situ dynamics and mechanism of glycoprotein substrate recognition and folding by the endoplasmic reticulum chaperones calreticulin and calnexin; Molecular basis of multidrug resistance gene regulation in candida albicans; Design, synthesis and evaluation of inhibitors of lipooxygenase as anti-inflammatory agents; Genome-wide analysis of Serine/Threonine-Specific protein kinases of Arabidopsis and experimental determination of their function; Investigations on the possible involvement of the bgl genes of Escherichia coli in stationary phase;

Research Highlights and achievements in completed and on-going projects are as follows:

• Embryonic stem cell (ES) based model of cardiovascular development was successfully used to identify and analyze genes required for cardiovascular development. Two novel genes asrij and rudhira were identified. Their importance in cardiovascular development was demonstrated.
• Candida drug resistance gene (CDR1) was shown as a major drug efflux pump contributing towards multi drug resistance in Candida. The studies will help in designing strategies to combat commonality occurring antifungal resistance.
• Rab 11 gene, apart from its known role during oogenesis, was also shown be essential during ommatidial differentiation in Drosophila eye development and for viability and maintenance of photoreceptor cells and associated bristles in Drosophila.
• Structural studies on peanut lectin, legume lectins, Erythina corollodendron lectin, Artocarpin lectin were carried out. Detailed computational analysis of cross linking of garlic lectin molecule by oligomannosides was carried out. Role of aromatic residues and loop length in generating carbohydrate specificity was elucidated.
• The calcium binding molecular chaperon 'calreticulin' has a unique oligosaccharide binding site. It functions by binding to its substrate through its lectin function only and that protein-protein interactions do not stabilize these reactions. It act as a scaffold for binding of other endoplasmic reticulum chaperons and folding catalysts.
• A large number of genes showed altered expression under the short exposure of rice seedlings to salinity stress. These novel stress responsive genes could be useful in studies related to genomics.
• The yeast mini chromosome maintenance genes MCM12 and MCM13 genes were cloned and characterised and none of them was found to be essential for growth. MCM12 was found to be same as chl1p, a gene earlier shown to be required for chromosome segregation was also shown to be required for transcriptional silencing, stress response and aging. It also plays role in sister chromatid cohesion, chromatin structure and DNA damage or DNA replication block check point function.
• A protein was identified from crude extract of rose which showed unusual thermal stability and specificity for oligosaccharides. This protein could provide useful information on its applicability as a thermal stabilizing agent in food industry and clinical laboratory. It could also find potential use as a tissue marker or in purification of glycoconjugates.
• An alternate pathway, C5 pathway, for formation of ALA (crucial precursor for heme biosynthesis) from glutamate, was shown to be functional in malarial parasite apicoplast and was therefore proposed as a novel drug target.
• Role of various amino acids in structure-function and stability was traced in 19 types of collagen. Among other findings, Proline and charged amino acids were proposed to play a pivotal role in stabilization of various types of collagen. The findings will be useful in understanding molecular basis for various genetic disorder of collagen, collagen-collagenase interaction and design of collagenase inhibitors.
• Studies on Functional and structural characteristics of Saccharomyces cerevisiae Meiosis - specific Hopl protein showed that the HOP1 via its zinc-finger motif is likely to play a direct role in meiotic chromosome pairing and recombination by its ability to promote synapsis between double-strand DNA helices from yeast to any other organism. This is a first demonstration of pairing of doublestranded DNA helices promoted by a component involved in chromosome synapsis and recombination.
• Under the studies on inhibition of telomerase activity in vivo and biological consequences in cultured mammlian cells, the telomerase activity was inhibited by a number of agents including PKC inhibitors and reverse transcriptase inhibitors Two new extracts of medicinal plants were also found to inhibit the telomerase activity.