Browsing by Author "Khurana, Jitendra P."

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Analysis of differentially expressed genes in leaf rust infected bread wheat involving seeding resistance gene Lr28
(CSIRO Publishing, 2011) Dhariwal, Raman; Vyas, Shailendra; Bhaganagare, Govindraj R.; Jha, Shailendra K.; Khurana, Jitendra P.; Tyagi, Akhilesh K.; Prabhu, Kumble V.; Balyan, Harindra S.; Gupta, Pushpendra K.
Genome-wide transcriptome analysis of seedling resistance to leaf rust conferred by Lr28 gene in wheat (Triticum aestivum L.) was conducted to identify differentially expressed genes during incompatible interaction. A virulent leaf rust race 77–5 was used for inoculation of resistant (HD2329 + Lr28) and susceptible (HD2329 – Lr28) wheat NILs and cDNA-AFLP analyses was carried out. As many as 223 differential transcripts appeared following leaf rust inoculation; these included 122 transcripts that appeared exclusively in resistant NIL, whereas 39 transcripts appeared both in resistant and susceptible NILs. Sequence analyses of 37 transcripts, which appeared in the resistant NIL revealed that 15 transcripts had homology with genes involved in protein synthesis, signal transduction, transport, disease resistance and metabolism. The functions of remaining 22 transcripts could not be determined; these included six novel genes reported for the first time in wheat. Specific primers could be designed for 18 of the 37 transcripts, which included genes with putative and unknown functions. Quantitative real time PCR analysis was conducted using these 18 pairs of primers. A majority (13) of these transcripts appeared within 48 h reaching a peak value at 96 h in resistant NIL signifying their role in providing leaf rust resistance.
Arabidopsis RSS1 mediates cross-talk between glucose and light signaling during hypocotyl elongation growth
(Nature Publishing Group, 2017) Singh, Manjul; Gupta, Aditi; Singh, Dhriti; Khurana, Jitendra P.; Laxmi, Ashverya
Plants possess exuberant plasticity that facilitates its ability to adapt and survive under challenging environmental conditions. The developmental plasticity largely depends upon cellular elongation which is governed by a complex network of environmental and phytohormonal signals. Here, we report role of glucose (Glc) and Glc-regulated factors in controlling elongation growth and shade response in Arabidopsis. Glc controls shade induced hypocotyl elongation in a dose dependent manner. We have identified a Glc repressed factor REGULATED BY SUGAR AND SHADE1 (RSS1) encoding for an atypical basic helix-loop-helix (bHLH) protein of unknown biological function that is required for normal Glc actions. Phenotype analysis of mutant and overexpression lines suggested RSS1 to be a negative regulator of elongation growth. RSS1 affects overall auxin homeostasis. RSS1 interacts with the elongation growth-promoting proteins HOMOLOG OF BEE2 INTERACTING WITH IBH 1 (HBI1) and BR ENHANCED EXPRESSION2 (BEE2) and negatively affects the transcription of their downstream targets such as YUCs, INDOLE-3-ACETIC ACID INDUCIBLE (IAAs), LONG HYPOCOTYL IN FAR-RED1 (HFR1), HOMEOBOX PROTEIN 2 (ATHB2), XYLOGLUCAN ENDOTRANSGLUCOSYLASE/HYDROLASES (XTHs) and EXPANSINS. We propose, Glc signals might maintain optimal hypocotyl elongation under multiple signals such as light, shade and phytohormones through the central growth regulatory bHLH/HLH module.
Characterization of rice homeobox genes, OsHOX22 and OsHOX24, and over-expression of OsHOX24 in transgenic Arabidopsis suggest their role in abiotic stress response
(Frontiers Media S.A., 2016) Bhattacharjee, Annapurna; Khurana, Jitendra P.; Jain, Mukesh
Homeobox transcription factors are well known regulators of plant growth and development. In this study, we carried out functional analysis of two candidate stress-responsive HD-ZIP I class homeobox genes from rice, OsHOX22, and OsHOX24. These genes were highly up-regulated under various abiotic stress conditions at different stages of rice development, including seedling, mature and reproductive stages. The transcript levels of these genes were enhanced significantly in the presence of plant hormones, including abscisic acid (ABA), auxin, salicylic acid, and gibberellic acid. The recombinant full-length and truncated homeobox proteins were found to be localized in the nucleus. Electrophoretic mobility shift assay established the binding of these homeobox proteins with specific DNA sequences, AH1 (CAAT(A/T)ATTG) and AH2 (CAAT(C/G)ATTG). Transactivation assays in yeast revealed the transcriptional activation potential of full-length OsHOX22 and OsHOX24 proteins. Homo- and hetero-dimerization capabilities of these proteins have also been demonstrated. Further, we identified putative novel interacting proteins of OsHOX22 and OsHOX24 via yeast-two hybrid analysis. Over-expression of OsHOX24 imparted higher sensitivity to stress hormone, ABA, and abiotic stresses in the transgenic Arabidopsis plants as revealed by various physiological and phenotypic assays. Microarray analysis revealed differential expression of several stress-responsive genes in transgenic lines as compared to wild-type. Many of these genes were found to be involved in transcriptional regulation and various metabolic pathways. Altogether, our results suggest the possible role of OsHOX22/OsHOX24 homeobox proteins as negative regulators in abiotic stress responses.
Development of SSR markers and construction of a linkage map in jute
(Indian Academy of Sciences, 2012) Das, Moumita; Banerjee, Sumana; Dhariwal, Raman; Vyas, Shailendra; Mir, Reyazul R.; Topdar, Niladri; Kundu, Avijit; Khurana, Jitendra P.; Tyagi, Akhilesh K.; Sarkar, Debabrata; Sinha, Mohit K.; Balyan, Harindra S.; Gupta, PUSHPENDRA K.
Jute is an important natural fibre crop, which is only second to cotton in its importance at the global level. It is mostly grown in Indian subcontinent and has been recently used for the development of genomics resources. We recently initiated a programme to develop simple sequence repeat markers and reported a set of 2469 SSR that were developed using four SSR-enriched libraries (Mir et al. 2009). In this communication, we report an additional set of 607 novel SSR in 393 SSR containing sequences. However, primers could be designed for only 417 potentially useful SSR. Polymorphism survey was carried out for 374 primer pairs using two parental genotypes (JRO 524 and PPO4) of a mapping population developed for fibre fineness; only 66 SSR were polymorphic. Owing to a low level of polymorphism between the parental genotypes and a high degree of segregation distortion in recombinant inbred lines, genotypic data of only 53 polymorphic SSR on the mapping population consisting of 120 RIL could be used for the construction of a linkage map; 36 SSR loci were mapped on six linkage groups that covered a total genetic distance of 784.3 cM. Hopefully, this map will be enriched with more SSR loci in future and will prove useful for identification of quantitative trait loci/genes for molecular breeding involving improvement of fibre fineness and other related traits in jute.
Emerging roles of auxin in abiotic stress responses
(Springer, 2015) Sharma, Eshan; Sharma, Raghvendra; Borah, Pratikshya; Jain, Mukesh; Khurana, Jitendra P.
Auxin is among the key growth regulators that play diverse roles in virtually all aspects of plant growth and development. Intensive investigations during the past two decades have helped in elucidation of auxin perception and signal transduction mechanisms operative in plants. In addition to its primary role in regulating plant development, several studies in recent years have provided unflinching evidence for the involvement of auxin in abiotic stress responses. Functional genomics studies and genome-wide expression analysis have revealed altered expression of auxin-responsive genes, such as Aux/IAA, GH3, SAURs, and ARFs, under abiotic stress conditions. Variations in endogenous levels of auxin at global and local levels under various abiotic stress conditions have been associated with phenotypic changes and provided intriguing evidences regarding its role in response to environmental changes. Modulation of reactive oxygen species (ROS) levels in response to exogenous auxin as well as to drought, salinity, and ABA have indicated towards a complex relationship network between auxin, ROS, and abiotic stresses in plants. The advent of recent functional genomics technologies has led to identification of several candidate genes that may modulate crosstalk between auxin and abiotic stresses. This chapter discusses auxin homeostasis, signal transduction mechanisms, and how these processes are modulated under abiotic stresses, thus emphasizing on the emerging roles of auxin as a key integrator of abiotic stress pathways and plant development.
Identification and characterization of high temperature stress responsive genes in bread wheat (Triticum aestivum L.) and their regulation at various stages of development
(Springer Science, 2011) Chauhan, Harsh; Khurana, Neetika; Tyagi, Akhilesh K.; Khurana, Jitendra P.; Khurana, Paramjit
To elucidate the effect of high temperature, wheat plants (Triticum aestivum cv. CPAN 1676) were given heat shock at 37 and 42°C for 2 h, and responsive genes were identified through PCR-Select Subtraction technology. Four subtractive cDNA libraries, including three forward and one reverse subtraction, were constructed from three different developmental stages. A total of 5,500 ESTs were generated and 3,516 high quality ESTs submitted to Genbank. More than one-third of the ESTs generated fall in unknown/no hit category upon homology search through BLAST analysis. Differential expression was confirmed by cDNA macroarray and by northern/RT-PCR analysis. Expression analysis of wheat plants subjected to high temperature stress, after 1 and 4 days of recovery, showed fast recovery in seedling tissue. However, even after 4 days, recovery was negligible in the developing seed tissue after 2 h of heat stress. Ten selected genes were analyzed in further detail including one unknown protein and a new heat shock factor, by quantitative real-time PCR in an array of 35 different wheat tissues representing major developmental stages as well as different abiotic stresses. Tissue specificity was examined along with cross talk with other abiotic stresses and putative signalling molecules.
MediatorWeb: a protein-protein interaction network database for the RNA polymerase II Mediator complex
(John Wiley & Sons, 2024) Maji, Sourobh; Waseem, Mohd; Sharma, Manish Kumar; Singh, Maninder; Singh, Anamika; Dwivedi, Nidhi; Thakur, Pallabi; Cooper, David G.; Bisht, Naveen C.; Fassler, Jan S.; Subbarao, Naidu; Khurana, Jitendra P.; Bhavesh, Neel Sarovar; Thakur, Jitendra K.
The protein-protein interaction (PPI) network of the Mediator complex is very tightly regulated and depends on different developmental and environmental cues. Here, we present an interactive platform for comparative analysis of the Mediator subunits from humans, baker's yeast Saccharomyces cerevisiae, and model plant Arabidopsis thaliana in a user-friendly web-interface database called MediatorWeb. MediatorWeb provides an interface to visualize and analyze the PPI network of Mediator subunits. The database facilitates downloading the untargeted and unweighted network of Mediator complex, its submodules, and individual Mediator subunits to better visualize the importance of individual Mediator subunits or their submodules. Further, MediatorWeb offers network visualization of the Mediator complex and interacting proteins that are functionally annotated. This feature provides clues to understand functions of Mediator subunits in different processes. In an additional tab, MediatorWeb provides quick access to secondary and tertiary structures, as well as residue-level contact information for Mediator subunits in each of the three model organisms. Another useful feature of MediatorWeb is detection of interologs based on orthologous analyses, which can provide clues to understand the functions of Mediator complex in less explored kingdoms. Thus, MediatorWeb and its features can help the user to understand the role of Mediator complex and its subunits in the transcription regulation of gene expression.
Modulation of transcription factor and metabolic pathway genes in response to water-deficit stress in rice
(Springer Science, 2011) Ray, Swatismita; Dansana, Prasant K.; Giri, Jitender; Deveshwar, Priyanka; Arora, Rita; Agarwal, Pinky; Khurana, Jitendra P.; Kapoor, Sanjay; Tyagi, Akhilesh K.
Water-deficit stress is detrimental for rice growth, development, and yield. Transcriptome analysis of 1-week-old rice (Oryza sativa L. var. IR64) seedling under water-deficit stress condition using Affymetrix 57 K GeneChip® has revealed 1,563 and 1,746 genes to be up- and downregulated, respectively. In an effort to amalgamate data across laboratories, we identified 5,611 differentially expressing genes under varying extrinsic water-deficit stress conditions in six vegetative and one reproductive stage of development in rice. Transcription factors (TFs) involved in ABA-dependent and ABA-independent pathways have been found to be upregulated during water-deficit stress. Members of zinc-finger TFs namely, C₂H₂, C₂C₂, C₃H, LIM, PHD, WRKY, ZF-HD, and ZIM, along with TF families like GeBP, jumonji, MBF1 and ULT express differentially under water-deficit conditions. NAC (NAM, ATAF and CUC) TF family emerges to be a potential key regulator of multiple abiotic stresses. Among the 12 TF genes that are co-upregulated under water-deficit, salt and cold stress conditions, five belong to the NAC TF family. We identified water-deficit stress-responsive genes encoding key enzymes involved in biosynthesis of osmoprotectants like polyols and sugars; amino acid and quaternary ammonium compounds; cell wall loosening and structural components; cholesterol and very long chain fatty acid; cytokinin and secondary metabolites. Comparison of genes responsive to water-deficit stress conditions with genes preferentially expressed during panicle and seed development revealed a significant overlap of transcriptome alteration and pathways.
A multi-step phosphorelay two-component system impacts on tolerance against dehydration stress in common wheat
(Springer, 2014) Gahlaut, Vijay; Mathur, Saloni; Dhariwal, Raman; Khurana, Jitendra P.; Tyagi, Akhilesh K.; Balyan, Harindra S.; Gupta, Pushpendra K.
Wheat is an important staple crop, and its productivity is severely constrained by drought stress (DS). An understanding of the molecular basis of drought tolerance is necessary for genetic improvement of wheat for tolerance to DS. The two-component system (TCS) serves as a common sensor-regulator coupling mechanism implicated in the regulation of diverse biological processes (including response to DS) not only in prokaryotes, but also in higher plants. In the latter, TCS generally consists of two signalling elements, a histidine kinase (HK) and a response regulator (RR) associated with an intermediate element called histidine phosphotransferase (HPT). Keeping in view the possible utility of TCS in developing water use efficient (WUE) wheat cultivars, we identified and characterized 62 wheat genes encoding TCS elements in a silico study; these included 7 HKs, 45 RRs along with 10 HPTs. Twelve of the 62 genes showed relatively higher alterations in the expression under drought. The quantitative RT-PCR (qRT-PCR)-based expression analysis of these 12 TCS genes was carried out in wheat seedlings of a drought sensitive (HD2967) and a tolerant (Dharwar Dry) cultivar subjected to either dehydration stress or cytokinin treatment. The expression of these 12 genes under dehydration stress differed in sensitive and tolerant genotypes, even though for individual genes, both showed either up-regulation or down-regulation. In response to the treatment of cytokinin, the expression of type-A RR genes was higher in the tolerant genotype, relative to that in the sensitive genotype, the situation being reverse for the type-B RRs. These results have been discussed in the context of the role of TCS elements in drought tolerance in wheat.
Polyphasic taxonomic analysis establishes Mycobacterium indicus pranii as a distinct species
(PLOS, 2009) Saini, Vikram; Raghuvanshi, Saurabh; Talwar, Gursaran P.; Ahmed, Niyaz; Khurana, Jitendra P.; Hasnain, Seyed E.; Tyagi, Akhilesh K.; Tyagi, Anil K.
Background: Mycobacterium indicus pranii (MIP), popularly known as Mw, is a cultivable, non-pathogenic organism, which, based on its growth and metabolic properties, is classified in Runyon Group IV along with M. fortuitum, M. smegmatis and M. vaccae. The novelty of this bacterium was accredited to its immunological ability to undergo antigen driven blast transformation of leukocytes and delayed hypersensitivity skin test in leprosy patients, a disease endemic in the Indian sub- continent. Consequently, MIP has been extensively evaluated for its biochemical and immunological properties leading to its usage as an immunomodulator in leprosy and tuberculosis patients. However, owing to advances in sequencing and culture techniques, the citing of new strains with almost 100% similarity in the sequences of marker genes like 16S rRNA, has compromised the identity of MIP as a novel species. Hence, to define its precise taxonomic position, we have carried out polyphasic taxonomic studies on MIP that integrate its phenotypic, chemotaxonomic and molecular phylogenetic attributes. Methodology/Principal Findings: The comparative analysis of 16S rRNA sequence of MIP by using BLAST algorithm at NCBI (nr database) revealed a similarity of $99% with M. intracellulare, M. arosiense, M. chimaera, M. seoulense, M. avium subsp. hominissuis, M. avium subsp. paratuberculosis and M. bohemicum. Further analysis with other widely used markers like rpoB and hsp65 could resolve the phylogenetic relationship between MIP and other closely related mycobacteria apart from M. intracellulare and M. chimaera, which shares $99% similarity with corresponding MIP orthologues. Molecular phylogenetic analysis, based on the concatenation of candidate orthologues of 16S rRNA, hsp65 and rpoB, also substantiated its distinctiveness from all the related organisms used in the analysis excluding M. intracellulare and M. chimaera with which it exhibited a close proximity. This necessitated further analysis of MIP with more sensitive and segregating parameters to ascertain its precise taxonomic position as a new species. The analysis of MIP and its comparison with other mycobacterial reference strains based on cellular and biochemical features, growth characteristics and chemotaxonomic studies like FAME profiling confirmed that MIP is uniquely endowed with diverse metabolic attributes that effectively distinguishes it from all the closely related mycobacteria including M. intracellulare and M. chimaera. Conclusion: The results presented in this study coupled with the non-pathogenic nature and different biochemical and immunomodulatory properties of MIP affirm it as a distinct species belonging to M. avium complex (MAC). It is further proposed to use an earlier suggested name Mycobacterium indicus pranii for this newly established mycobacterial species. This study also exemplifies the growing need for a uniform, consensus based broader polyphasic frame work for the purpose of taxonomy and speciation, particularly in the genus Mycobacterium.
Rice improvement through genome-based functional analysis and molecular breeding in India
(Springer, 2016) Agarwal, Pinky; Parida, Swarup K.; Raghuvanshi, Saurabh; Kapoor, Sanjay; Khurana, Paramjit; Khurana, Jitendra P.; Tyagi, Akhilesh K.
Rice is one of the main pillars of food security in India. Its improvement for higher yield in sustainable agriculture system is also vital to provide energy and nutritional needs of growing world population, expected to reach more than 9 billion by 2050. The high quality genome sequence of rice has provided a rich resource to mine information about diversity of genes and alleles which can contribute to improvement of useful agronomic traits. Defining the function of each gene and regulatory element of rice remains a challenge for the rice community in the coming years. Subsequent to participation in IRGSP, India has continued to contribute in the areas of diversity analysis, transcriptomics, functional genomics, marker development, QTL mapping and molecular breeding, through national and multi-national research programs. These efforts have helped generate resources for rice improvement, some of which have already been deployed to mitigate loss due to environmental stress and pathogens. With renewed efforts, Indian researchers are making new strides, along with the international scientific community, in both basic research and realization of its translational impact.
Small RNA regulation of rice homeobox genes
(Landes Bioscience, 2008) Jain, Mukesh; Khurana, Jitendra P.
Recently, we reported the genome-wide identification of 107 homeobox genes in rice and classified them into ten distinct subfamilies based upon their domain composition and phylogenetic analysis. Microarray analysis revealed the tissue-specific and overlapping expression profiles of these genes during various stages of floral transition, panicle development and seed set. Several homeobox genes were also found to be differentially expressed under abiotic stress conditions. Based on massively parallel signature sequencing (MPSS) data analysis, we report here that a large number of small RNA signatures are associated with rice homeobox genes, which may be involved in their tissue-specific/developmental regulation and stress responses. The association of a very large number of small RNA signatures suggested an unusually high degree of regulation of homeobox genes by small RNAs during inflorescence development.
Transcript profiling reveals diverse roles of auxin-responsive genes during reproductive development and abiotic stress in rice
(Wiley-Blackwell, 2009) Jain, Mukesh; Khurana, Jitendra P.
Auxin influences growth and development in plants by altering gene expression. Many auxin-responsive genes have been characterized in Arabidopsis in detail, but not in crop plants. Earlier, we reported the identification and characterization of the members of the GH3, Aux/IAA and SAUR gene families in rice. In this study, whole genome microarray analysis of auxin-responsive genes in rice was performed, with the aim of gaining some insight into the mechanism of auxin action. A comparison of expression profiles of untreated and auxin-treated rice seedlings identified 315 probe sets representing 298 (225 upregulated and 73 downregulated) unique genes as auxin-responsive. Functional categorization revealed that genes involved in various biological processes, including metabolism, transcription, signal transduction, and transport, are regulated by auxin. The expression profiles of auxin-responsive genes identified in this study and those of the members of the GH3, Aux/IAA, SAUR and ARF gene families were analyzed during various stages of vegetative and reproductive (panicle and seed) development by employing microarray analysis. Many of these genes are, indeed, expressed in a tissue-specific or developmental stage-specific manner, and the expression profiles of some of the representative genes were confirmed by real-time PCR. The differential expression of auxin-responsive genes during various stages of panicle and seed development implies their involvement in diverse developmental processes. Moreover, several auxin-responsive genes were differentially expressed under various abiotic stress conditions, indicating crosstalk between auxin and abiotic stress signaling.
The water-deficit stress and red-rot-related genes in sugarcane
(Springer, 2010) Gupta, Vikrant; Raghuvanshi, Saurabh; Gupta, Ambika; Saini, Navin; Gaur, Anupama; Khan, M.S.; Gupta, R.S.; Singh, J.; Duttamajumder, S.K.; Srivastava, S.; Suman, A.; Khurana, Jitendra P.; Kapur, Raman; Tyagi, Akhilesh K.
Sugarcane is an important international commodity as a valuable agricultural crop especially in developing countries. Sequencing was carried out to generate >35,000 expressed sequence tags (ESTs) from healthy as well as red-rot-infected tissue of Indian subtropical variety of sugarcane. Subsequent clustering with existing sugarcane ESTs in public databases identified 4,087 clusters, including 85 clusters that preferentially express upon Colletotrichum falcatum (red-rot) infection, which were previously unreported. Real-time reverse transcription-PCR profiling of selected EST clusters identified several sugarcane clusters that show differential expression in response to biotic and abiotic stress conditions. Twenty-five stress-related clusters showed >2-fold relative expression during water-deficit stress in sugarcane. Similarly, EST clusters could be identified, which exhibit association with red-rot disease when assessed in red-rot-susceptible and red-rot-resistant varieties of sugarcane. Such EST clusters are good candidates for in-depth analysis to elucidate stress-responsive pathways in sugarcane and facilitate genetic manipulation to tailor this crop for tolerance to various stresses.