Browsing by Author "Bhattacharjee, Oindrila"

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CRISPR-Cas9 system: A new-fangled dawn in gene editing
(Elsevier B.V., 2019) Gupta, Darshana; Bhattacharjee, Oindrila; Mandal, Drishti; Sen, Madhab Kumar; Dey, Dhritiman; Dasgupta, Adhiraj; Kazi, Tawsif Ahmed; Gupta, Rahul; Sinharoy, Senjuti; Acharya, Krishnendu; Chattopadhyay, Dhrubajyoti; Ravichandiran, V.; Roy, Syamal; Ghosh, Dipanjan
Till date, only three techniques namely Zinc Finger Nuclease (ZFN), Transcription-Activator Like Effector Nucleases (TALEN) and Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR-Associated 9 (CRISPR-Cas9) are available for targeted genome editing. CRISPR-Cas system is very efficient, fast, easy and cheap technique for achieving knock-out gene in the cell. CRISPR-Cas9 system refurbishes the targeted genome editing approach into a more expedient and competent way, thus facilitating proficient genome editing through embattled double-strand breaks in approximately any organism and cell type. The off-target effects of CRISPR Cas system has been circumnavigated by using paired nickases. Moreover, CRISPR-Cas9 has been used effectively for numerous purposes, like knock-out of a gene, regulation of endogenous gene expression, live-cell labelling of chromosomal loci, edition of single-stranded RNA and high-throughput gene screening. The execution of the CRISPR-Cas9 system has amplified the number of accessible scientific substitutes for studying gene function, thus enabling generation of CRISPR-based disease models. Even though many mechanistic questions are left behind to be answered and the system is not yet fool-proof i.e., a number of challenges are yet to be addressed, the employment of CRISPR-Cas9–based genome engineering technologies will increase our understanding to disease processes and their treatment in the near future. In this review we have discussed the history of CRISPR-Cas9, its mechanism for genome editing and its application in animal, plant and protozoan parasites. Additionally, the pros and cons of CRISPR-Cas9 and its potential in therapeutic application have also been detailed here.
Genome-wide identification of auxin response factors (ARFs) in three different species of Arachis
(Springer Nature Publishing AG, 2021) Raul, Bikash; Bhattacharjee, Oindrila; Tembhare, Kunal; Khanna, Tanyya; Shaheen, Tarannum; Sinharoy, Senjuti; Bandyopadhyay, Kaustav
The phytohormone auxin is involved in the regulation of plant growth, nutrient acquisition, and response to environmental stimuli. Auxin response factors (ARFs) are transcription factors containing B3 DNA binding domain. ARFs play central role in auxin response, using Aux/IAA proteins as partners. Arachis is a genus within the Dalbergioid clade of papilionoid legumes, which out-branched from other members of papilionoids. Cultivated peanut (Arachis hypogaea L.) is an allotetraploid formed by hybridization of two parental genotypes Arachis duranensis, and Arachis ipaensis merely 10,000 years ago. We have made a genome-wide inventory of all the ARFs present in tetraploid A. hypogaea, as well as in two diploid parental genotypes. Our data show that Arachis contains more ARFs per diploid genome (around 31), compared to other legumes (around 25). We further observed few ARF-like genes which are defective in important domains. Most of the ARFs in tetraploid Arachis are redundant, representing the A and B sub-genomes. Some of the ARFs show expression bias from either A or B sub-genome, while some of the pairs are expressed from both sub-genomes. Many ARFs do not express in any of the conditions for which we have expression data. Finally, few pairs show diferential spatio-temporal expression pattern from A and B sub-genomes, indicative of diversifcation of function. This is the frst efort to list all the ARFs from an allotetraploid legume. The list of ARFs in all three species of Arachis will help the scientifc community working to understand auxin regulation in crop legumes.
Microscopic and transcriptomic analyses of Dalbergoid legume peanut reveal a divergent evolution leading to Nod Factor dependent epidermal crack-entry and terminal bacteroid differentiation
(American Phytopathological Society, 2022) Raul, Bikash; Bhattacharjee, Oindrila; Ghosh, Amit; Upadhyay, Priya; Tembhare, Kunal; Singh, Ajeet; Shaheen, Tarannum; Ghosh, Asim Kumar; Torres-Jerez, Ivone; Krom, Nick; Clevenger, Josh; Udvardi, Michael; Scheffler, Brian E.; Ozias-Akins, Peggy; Sharma, Ravi Datta; Bandyopadhyay, Kaustav; Gaur, Vineet; Kumar, Shailesh; Sinharoy, Senjuti
Root nodule symbiosis (RNS) is the pillar behind sustainable agriculture and plays a pivotal role in the environmental nitrogen cycle. Most of the genetic, molecular, and cell-biological knowledge on RNS come from model legumes that exhibit a root-hair mode of bacterial infection in contrast to the Dalbergoid legumes exhibiting crack-entry of rhizobia. As a step towards understanding this important group of legumes, we have combined microscopic analysis and temporal transcriptome to obtain a dynamic view of plant gene expression during Arachis hypogaea (peanut) nodule development. We generated a comprehensive transcriptome data by mapping the reads to A. hypogaea, and two diploid progenitor genomes. Additionally, we performed BLAST searches to identify nodule-induced yet-to-be annotated peanut genes. Comparison between peanut, Medicago truncatula, Lotus japonicus, and Glycine max showed upregulation of 61 peanut orthologs among 111 tested known RNS-related genes, indicating conservation in mechanisms of nodule development among members of the Papilionoid family. Unlike model legumes, recruitment of class 1 phytoglobin derived symbiotic hemoglobin (SymH) in peanut indicates diversification of oxygen scavenging mechanisms in the Papilionoid family. Finally, absence of cysteine-rich motif-1 containing-NCRs, but the recruitment of defensin like NCRs suggest a diverse molecular mechanism of terminal bacteroid differentiation. In summary, our work describes genetic conservation and diversification in legume-rhizobial symbiosis in the Papilionoid family, as well as among members of the Dalbergoid legumes.
Nodule INception (NIN)-independent epidermal events lead to bacterial entry during nodule development in peanut (Arachis hypogaea)
(John Wiley & Sons, 2022) Bhattacharjee, Oindrila; Raul, Bikash; Ghosh, Amit; Bhardwaj, Akanksha; Bandyopadhyay, Kaustav; Sinharoy, Senjuti
Summary Legumes can host nitrogen-fixing rhizobia inside root nodules. In model legumes, rhizobia enter via infection threads (ITs) and develop nodules where infection-zone contains a mixture of infected and uninfected cells. Peanut (Arachis hypogaea) diversified from model legumes ~50-55 million years ago. Rhizobia enter through ‘cracks’ to form nodules in peanut roots where the cells of infection-zone are uniformly infected. Phylogenomic studies indicated symbiosis as a labile trait in peanut. These atypical features prompted us to investigate the molecular mechanism of peanut nodule development. Combining cell biology, genetics, and genomic tools, we visualized the status of hormonal signaling in peanut nodule primordia. Moreover, we dissected the signaling modules of Nodule INception (NIN), a master regulator of both epidermal infection and cortical organogenesis. Cytokinin signaling operates in a broad zone, from the epidermis to the pericycle inside nodule primordia, while auxin signaling is narrower and focused. NIN is involved in the nodule organogenesis, but not in the crack entry. Nodulation Pectate Lyase (NPL), which remodels cell walls during IT-formation, is not required. Whereas Nodule enhanced Glycosyl Hydrolases (AhNGHs) is recruited for cell wall modification during crack entry. While the hormonal regulation is conserved, the function of the NIN signaling modules is diversified in peanut.
RSD-mediated suppression of NIN and NLP2 transcription is crucial for symbiotic nitrogen fixation
(John Wiley & Sons, 2026) Bhardwaj, Akanksha; Gupta, Megha; Bhattacharjee, Oindrila; Raul, Bikash; Ghosh, Asim Kumar; Nagalla, L. V. Sairam; Yadav, Prashant; Bandyopadhyay, Kaustav; Ranjan, Aashish; Sinharoy, Senjuti
Indeterminate nodules establish a developmental gradient along their longitudinal axis, separating cell differentiation from symbiotic nitrogen fixation (SNF). The apex contains differentiating cells, while the differentiated leghemoglobin-rich cells of the nitrogen-fixation zone shield the rhizobial nitrogenase complex from oxygen to facilitate SNF. By combining biochemical, genetic, and genomic approaches, we demonstrate the pivotal role of the Regulator of Symbiosome Differentiation (RSD), a transcriptional repressor, in the transition from symbiosome development to SNF. Interacting Protein of DMI3 (IPD3) activates RSD expression in the invasion zone (ZII) and interzone (IZ). RSD interacts with Nodule Inception (NIN), and NIN-like protein 2 (NLP2) through a novel protein-protein interaction domain. RSD determines cell fate in ZII and the IZ by suppressing several targets of NIN and NLP2, including Leghemoglobins, Nodule-specific Cysteine-Rich genes, and Symbiotic Cysteine-rich Receptor-like Kinase. Our findings underscore the critical role of RSD-mediated suppression of transcription in facilitating the transition from bacteroid differentiation to SNF.