Anju Singh

A bouquet of DNA structures: Emerging diversity

Structural polymorphism of DNA has constantly been evolving from the time of illustration of the double helical model of DNA by Watson and Crick. A variety of non-canonical DNA structures have constantly been documented across the globe. DNA attracted worldwide attention as a carrier of genetic information. In addition to the classical Watson–Crick duplex, DNA can actually adopt diverse structures during its active participation in cellular processes like replication, transcription, recombination and repair. Structures like hairpin, cruciform, triplex, G-triplex, quadruplex, i-motif and other alternative non-canonical DNA structures have been studied at length and have also shown their in vivo occurrence. This review mainly focuses on non-canonical structures adopted by DNA oligonucleotides which have certain prerequisites for their formation in terms of sequence, its length, number and orientation of strands along with varied solution conditions. This conformational polymorphism of DNA might be the basis of different functional properties of a specific set of DNA sequences, further giving some insights for various extremely complicated biological phenomena. Many of these structures have already shown their linkages with diseases like cancer and genetic disorders, hence making them an extremely striking target for structure-specific drug designing and therapeutic applications.

Structural transition from dimeric to tetrameric i-motif, caused by the presence of TAA at the 3′-end of human telomeric C-rich sequence

Widely dispersed in genomic DNA, the tandem C-rich repetitive stretches may fold below physiological pH, into i-motif structures, stabilized by C.C(+) pairing. Herein, structural status of a 9-mer stretch d(CCCTAACCC), [the truncated double repeat of human telomeric sequence], and its extended version, comprising of additional--TAA segment at the 3-end, representing the complete double repeat d(CCCTAACCCTAA), has been investigated. The pH dependent monophasic UV-melting, Gel and CD data suggested that while the truncated version adopts a bimolecular i-motif structure, its complete double repeat (12-mer) sequence exists in two (bimolecular and tetramolecular) forms. A model is proposed for the tetramolecular i-motif with conventional C.C(+) base pairs, additionally stabilized by asymmetric A.A base pairs at the -3 TAA flanking ends and Watson-Crick A.T hydrogen bonding between intervening bases on antiparallel strands. Expanding the known topologies of DNA i-motifs, such atypical geometries of i-motifs may have implications in their recognition by proteins.

Molecular mapping of the grain iron and zinc concentration, protein content and thousand kernel weight in wheat (Triticum aestivum L.)

Genomic regions responsible for accumulation of grain iron concentration (Fe), grain zinc concentration (Zn), grain protein content (PC) and thousand kernel weight (TKW) were investigated in 286 recombinant inbred lines (RILs) derived from a cross between an old Indian wheat variety WH542 and a synthetic derivative (Triticum dicoccon PI94624/Aegilops squarrosa [409]//BCN). RILs were grown in six environments and evaluated for Fe, Zn, PC, and TKW. The population showed the continuous distribution for all the four traits, that for pooled Fe and PC was near normal, whereas, for pooled Zn, RILs exhibited positively skewed distribution. A genetic map spanning 2155.3cM was constructed using microsatellite markers covering the 21 chromosomes and used for QTL analysis. 16 quantitative trait loci (QTL) were identified in this study. Four QTLs (QGFe.iari-2A, QGFe.iari-5A, QGFe.iari-7A and QGFe.iari-7B) for Fe, five QTLs (QGZn.iari-2A, QGZn.iari-4A, QGZn.iari-5A, QGZn.iari-7A and QGZn.iari-7B) for Zn, two QTLs (QGpc.iari-2A and QGpc.iari-3A) for PC, and five QTLs (QTkw.iari-1A, QTkw.iari-2A, QTkw.iari-2B, QTkw.iari-5B and QTkw.iari-7A) for TKW were identified. The QTLs together explained 20.0%, 32.0%, 24.1% and 32.3% phenotypic variation, respectively, for Fe, Zn, PC and TKW. QGpc.iari-2A was consistently expressed in all the six environments, whereas, QGFe.iari-7B and QGZn.iari-2A were identified in two environments each apart from pooled mean. QTkw.iari-2A and QTkw.iari-7A, respectively, were identified in four and three environments apart from pooled mean. A common region in the interval of Xgwm359-Xwmc407 on chromosome 2A was associated with Fe, Zn, and PC. One more QTL for TKW was identified on chromosome 2A but in a different chromosomal region (Xgwm382-Xgwm359). Two more regions on 5A (Xgwm126-Xgwm595) and 7A (Xbarc49-Xwmc525) were found to be associated with both Fe and Zn. A QTL for TKW was identified (Xwmc525-Xbarc222) in a different chromosomal region on the same chromosome (7A). This reflects at least a partly common genetic basis for the four traits. It is concluded that fine mapping of the regions of the three chromosomes of A genome involved in determining the accumulation of Fe, Zn, PC, and TKW in this mapping population may be rewarding.

A triple stranded G-quadruplex formation in the promoter region of human myosin β(Myh7) gene

Regulatory regions in human genome, enriched in guanine-rich DNA sequences have the propensity to fold into G-quadruplex structures. On exploring the genome for search of G-tracts, it was interesting to find that promoter of Human Myosin Gene (MYH7) contains a conserved 23-mer G-rich sequence (HM-23). Mutations in this gene are associated with familial cardiomyopathy. Enrichment of MYH7 gene in G-rich sequences could possibly play a critical role in its regulation. We used polyacrylamide gel electrophoresis (PAGE), UV-Thermal denaturation (UV-Tm) and Circular Dichroism (CD), to demonstrate the formation of a G-quadruplex by 23-mer G-rich sequence HM23 in promoter location of MYH7 gene. We observed that the wild G-rich sequence HM23 containing consecutive G5 stretch in two stacks adopt G-quadruplexes of diverse molecularity by involvement of four-strand, three-strand and two-strands with same parallel topology. Interestingly, the mutated sequence in the absence of continuous G5 stretch obstructs the formation of three-stranded G-quadruplex. We demonstrated that continuous G5 stretch is mandatory for the formation of a unique three-stranded G-quadruplex. Presence of various transcription factors (TF) in vicinity of the sequence HM23 leave fair possibility of recognition by TF binding sites, and so modulate gene expression. These findings may add on our understanding about the effect of base change in the formation of varied structural species in similar solution condition. This study may give insight about structural polymorphism arising due to recognition of non-Watson-Crick G-quadruplex structures by cellular proteins and designing structure specific molecules.

Luminescence, circular dichroism and in silico studies of binding interaction of synthesized naphthylchalcone derivatives with bovine serum albumin

Chalcones possess various biological properties, for example, antimicrobial, anti-inflammatory, analgesic, antimalarial, anticancer, antiprotozoal and antitubercular activity. In this study, naphthylchalcone derivatives were synthesized and characterized using 1 H NMR 13 C NMR, Fourier transform infrared and mass techniques. Yields for all derivatives were found to be >90%. Protein-drug interactions influence the absorption, distribution, metabolism and excretion (ADME) properties of a drug. Therefore, to establish whether the synthesized naphthylchalcone derivatives can be used as drugs, their binding interaction toward a serum protein (bovine serum albumin) was investigated using fluorescence, circular dichroism and molecular docking techniques under physiological conditions. Fluorescence quenching of the protein in the presence of naphthylchalcone derivatives, and other derived parameters such as association constants, number of binding sites and static quenching involving confirmed non-covalent binding interactions in the protein-ligand complex were observed. Circular dichroism clearly showed changes in the secondary structure of the protein in the presence of naphthylchalcones, indicating binding between the derivatives and the serum protein. Molecular modelling further confirmed the binding mode of naphthylchalcone derivatives in bovine serum albumin. A site-specific molecular docking study of naphthylchalcone derivatives with serum albumin showed that binding took place primarily in the aromatic low helix and then in subdomain II. The dominance of hydrophobic, hydrophilic and hydrogen bonding was clearly visible and was responsible for stabilization of the complex.

Structure-Specific Ligand Recognition of Multistranded DNA Structures.

Structural polymorphism is an extremely significant phenomenon of nucleic acids, in which DNA and RNA oligonucleotide sequences are able to adapt various canonical, alternative and multistranded structures. These alternative forms of DNA and RNA have an enormous potential of participating in various cellular processes by recognizing ligands such as proteins, drugs and metal ions in a sequence and structure-specific manner. Such DNA-ligand interactions prove to be highly beneficial when exploited for therapeutic purposes. Many of these DNA/ RNA structures recognizing drugs have already proved their potential as anticancer, antibacterial, anthelmintic and antiviral properties. Over the last 2-3 decades, many mechanisms of DNA-drug interactions have been documented, but still many other new mechanisms are being explored. Designing new drugs with improved efficacy and specificity is of prime concern for all researchers which not only deals with the experiments related to synthesizing drugs, but also takes care of searching novel routes or agents for administration or delivery of these therapeutic agents by increasing their nuclear and cellular uptake. This review aims at explaining the structural polymorphs/ multistranded DNA structures and their interactions with pharmaceutical drugs in a structure-specific manner, along with their modes of interactions and biological relevance. This detailed overview of multistranded DNA structures and interacting drugs might further facilitate our understanding about molecular targets and drug development in a more precise manner for the larger benefit of mankind.

Tourism and Solid Waste Management

Himalaya provides breath-refreshing pure air with peaceful environment and a pleasant climate, making it an ideal site for tourism and recreational activities. The carrying capacity of this region is low however, due to its fragile environment which gets unstable with miniscule changes. The town of Dharamsala lying in the Kangra district of Himachal Pradesh is a preferred destination for both domestic and international tourists due to of variety of reasons. Domestic tourism has increased by 14 % in 2006–2011 and international tourism by 3 % during the same period. The increased tourism in town leads to air pollution and sewerage problems, along with huge increased quantity of solid waste. The total solid waste generated in town is about 14,000 kg/day, and the town does not have enough infrastructures to dispose of it all. Only one landfill site is available and it is not properly managed. The growing garbage heaps and lack of proper management lead to various environmental and health problems to the various stakeholders, also having negative implications on tourism industry. Integrated Solid Waste Management is a key to mitigating the problem.

Climate Change Variability in Coastal Karnataka, India

Climate change is one of the biggest environmental threats to food production, water availability, forest biodiversity and livelihoods. It is widely believed that developing countries such as India will be impacted more severely than developed countries. Global warming and Climate Change is projected to increase the number of extreme temperature and rainfall events, and hence climate variability is expected to show an upward trend. It is very important to understand the past trends and variability in rainfall, minimum and maximum temperature in Karnataka since the knowledge on the past could provide guidance for the future. The Arabian Sea and the North Kanara coast belong to the “Indo-Australian Marine Bio-geographic Region” considered to be the richest in the world for biodiversity. The coastal backwaters, estuaries, river-mouths are well known for their productivity. Some of them like the Aghanashini, Kali and Sharavati river backwaters, are even today so. The rivers from the Western Ghats carry great quantity of forest organic matter and deposit the same in the coastal waters including the sea. The current climate variability in Karnataka has been analyzed using the IMD daily rainfall data for the period of 1971–2005 and CRU data for the temperature for the period 1901–2002. The Costal Karnataka districts have an average rainfall of >25 mm/day. Decrease in precipitation trend has been observed in Coastal Karnataka. Rainfall has decreased by 17.69, 1.87 and 22.38 mm/day/100 year in Mangalore, North Kanara and Udupi respectively. The Increase in minimum temperature is 0.141, 0.146 and 0.111°C/100 year, whereas increase in maximum temperature is observed as 0.148, 0.146 and 0.113°C/100 year in districts of Mangalore, North Kanara and Udupi respectively.

Assessment of genetic diversity based on quality and morphological characters in spring wheat (Triticum aestivum L em Thell)

Wheat (Triticum aestivum L Em Thell) is the leading staple food crop globally which contribute about 30 % of global cereal production. It is being grown in 218.5 million hectare area with an average productivity of 3.26 tonnes/ ha (FAO, 2014). Wheat has a unique property of having a storage protein called ‘Gluten’, which has viscoelastic nature due to which it can be processed into the large number of end products. Currently, only 15 to 18 per cent of total wheat produced in India is used by roll flour mills. Of the remaining harvested wheat less than 7 per cent goes as seed and for grain export. The remaining 76+ percent is used by the village based unorganized stone roller grinding mills to produce whole grain Atta that is used for Chapati (Nagarajan 2013). When wheat grains are grounded without the aleuron layer and seed embryo the resultant white flour is called Maida. Whole wheat is rich in gluten, a protein conglomerate that increases the elasticity of the dough. From Maida, pan bread, noodles, cakes, biscuit and pizza can be made. For every good quality product specific and adapted wheat flour is required (Nagarajan 2013).

Homoduplex to i-motif structural switch exhibited by a cytosine rich strand of the MYH7 heavy chain β gene promoter at physiological pH

Genomic locations such as promoter, exon, intron, telomeric and non-telomeric regions are rich in GC-rich sequences with the potential to form G- and C-tetraplexes on both strands independently. Herein, we employed biophysical and biochemical methods to study a 34-mer C-rich DNA sequence of the myosin heavy chain β gene (MYH7β) promoter, namely HM34C for humans and the rabbit counterpart, RM34C, which differs from HM34C at three positions (three bases). Circular dichroism (CD), UV-thermal denaturation and native gel electrophoresis studies demonstrated that both the C-rich promoter segments form C-tetraplex (i-motif) structures. The CD studies revealed that HM34C forms the i-motif structure at acidic pH (5.2) in the presence of 0.1 M NaCl but remains unstructured at physiological pH. Interestingly, RM34C can form the stable i-motif structure in acidic as well as physiological pH. A shift in the positive peak from 280 nm to 275 nm with the increase in temperature from 4 °C to 30 °C was observed in temperature-dependent CD studies. UV-melting studies showed a biphasic transition for RM34C, indicating the existence of two structural species at neutral pH. In view of these findings we suggest that at physiological pH, the RM34C sequence exists in equilibrium between two structural motifs, i.e. the i-motif and homoduplex structure. This study may add to the understanding of the i-motif/homoduplex in equilibrium in physiological environments.