Proteomics and molecular biology of abiotic stress response and tolerance
Control of gene expression during development or adaptation to an environment involves perception and integration of cellular and environmental signals, the regulators. Whereas the roles of proteins as gene regulatory factors are well established, the functions of regulatory RNA molecules in development are just beginning to emerge. Among these regulatory RNAs, MicroRNAs (miRNAs) have generated considerable excitement recently. A growing body of evidence suggests that these ∼21-nucleotide (nt) noncoding RNA molecules play crucial roles as regulators of gene expression in eukaryotes. The regulatory proteins, nevertheless, also regulate their expression.
As proteins mediate all the metabolic activities of living organism, it is likely that the concerned genes would have a distinctly different expression patter in the plant under contrasting environmental situation. Similarly, two cultivars contrast for stress tolerance or yield potential would have set of differentially expressing genes influencing the trait. Identification of the differentially expressed genes for a trait may throw information on the biochemical basis of phenotype displayed. Study of expression pattern of the differentially expressing genes for a environmental trait across plant species contrast for the trait would further shortlist the genes associated with the trait. The work is also being extended to crop cultivars sensitive and tolerant to salinity. Among the crop cultivars, rice provides extensive variability for both yield potential and abiotic stress tolerance. Besides, its panicle architecture is such that the crop yield is reflected in terms of spatial positioning of spikelets in the position. Hence, the plant provides opportunity to look into differential expression of genes for a trait, particularly yield or stress-induced loss of yield, in the same plant.
With regard to survival of plants or yield potential of crop plants under an environmental condition or cultivar-specific yield potential of crop plants, the following pertinent questions thus surface: 1) whether the trait is governed by one or more effectors or isoform-specific effector and how the effectors are regulated, 2) if isoform-specific effector(s), how the expression the individual isoforms is regulated, 3) what is the nature of the environmental signal that activates the regulatory network. These questions are being looked into through genomics and proteomics approaches under the following heads:
a) Identification and functional characterization of the key effectors and regulators making a plant salt tolerant and those leading to poor grain filling in rice
b) Understanding transcriptional regulation of effectors and regulators and their link to environmental signals.
c) QTL identification for yield and abiotic stress tolerance.
Membership of professional bodies, societies, academies, etc.
- Society of Toxicology (STOX), Izzatnagar
- Orissa Bigyan Academy (OBA), Bhubaneswar
- Indian Society for Plant Physiologists (ISPP), New Delhi
- Society for Plant Biochemistry and Biotechnology (SPBB), New Delhi
Proteomics and molecular biology of abiotic stress response and tolerance
i) Identification and functional characterization of the key effectors and regulators making a plant salt tolerant and those leading to poor grain filling in rice
Genomic approach:
It is very likely that only naturally salt tolerant plants, and not the non-tolerant ones, would give valuable salt-response information related to salt-tolerance, and hence NGS transcriptome data have been generated for two salt tolerant plants, S. maritima (a dicot) and Porteresia coarctata (a monocot) grown in presence and absence of NaCl. The data analysis has been planned to provide information on three aspects: 1) the genes expressing differentially in response to NaCl in the individual plants, 2) the commonality in differential expression in the two species, and 3) biochemical function of the differentially expressing gene products, and if not effector, their connectivity network to the effector. Similarly, trascriptome data for superior (apical) and inferior (basal) spikelets of compact panicle rice cultivar are being analyzed for differential expression of effectors and regulators.
Proteomic approach:
Since metabolic processes are mostly carried out by protein and the level of proteins and expression of the concerned genes may not match at a given time, proteomics become a more direct approach in attacking a problem. The hypothesis for the proteomic approach would be a straightforward: 1) For the study on salt tolerance, the differentially expressing proteins would be resolved by 2D-PAGE and sequenced on MALDI-TOF/TOF and the presence/absence of these differentially expressing proteins will be seen in other salt-tolerant species, which would be considered the key genes making a plant salt tolerant. 2) For the grain filling problem, the proteins expressing differentially in the apical and basal spikelets of a compact panicle rice showing poor grain filling in the basal spikelets will be identified by MALDI-TOF/TOF after resolving on 2D-PAGE and those found showing expression pattern similar in the basal spikelets of rice cultivar showing good grain filling in them would be considered the candidate genes for grain filling. A working model for both salt tolerance and grain filling would be developed after proper validation of the result considering the result of genomic studies and functional validation of the model will be tested taking transgenic approach on suitable rice cultivar.
Involvement of miRNAs:
It is now well established that miRNAs play important role in post-transcritional gene silencing in plant. Several salt-responsive conserved and novel miRNAs has been identified in S. maritima, and similar work is in progress for P. coarctata. Effector and regulator targets of these salt-responsive miRNAs would be identified and expression of these miRNAs and their targets would be validated in other test plants. A working model will be developed and tested on Arabidopsis thaliana and Brachypodium distachyon taking transgenic approach. Similar approach will also be taken to address the problem of poor grain filling in the basal spikelets of rice panicle. A working model will be developed and tested on suitable rice cultivar taking transgenic approach.
ii) Understanding transcriptional regulation of effectors and regulators and their link to environmental signals.
A recent experiment in the lab has shown poor filling of the grain in rice at sub-lethal concentration of NaCl application. The result also indicated differential expression of several genes. Besides, several differentially expressing genes, including isoforms, have been identified. To understand the reason, the trans-element interacting with ~2 Kb upstream of the transcription start site of the individual genes will be identified by MALDI-TOF/TOF. Studies pertaining to biochemical characterization of the proteins to understand their regulatory role in gene expression and their possible link to abiotic factors will be carried out as per the requirement of the results obtained.
iii) QTL identification for panicle compactness:
Ongoing experiments in rice have revealed association of panicle compactness with high expression of ethylene receptors. Hence, it is conceivable that the two phenotypes are linked, and QTL identification would reveal the genes associated with panicle compactness. This will be achieved by generating RIL mapping population of cross between rice cultivar Mahalxmi (compact panicle) and Upahar (lax-panicle) and using SSR marker in collaboration with CRRI, Cuttack.
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