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    The landscape of fusion transcripts in plants: a new insight into genome complexity
    (BioMed Central Ltd, 2024) Chitkara, Pragya; Singh, Ajeet; Gangwar, Rashmi; Bhardwaj, Rohan; Zahra, Shafaque; Arora, Simran; Hamid, Fiza; Arya, Ajay; Sahu, Namrata; Chakraborty, Srija; Ramesh, Madhulika; Kumar, Shailesh
    Background Fusion transcripts (FTs), generated by the fusion of genes at the DNA level or RNA-level splicing events significantly contribute to transcriptome diversity. FTs are usually considered unique features of neoplasia and serve as biomarkers and therapeutic targets for multiple cancers. The latest findings show the presence of FTs in normal human physiology. Several discrete reports mentioned the presence of fusion transcripts in planta, has important roles in stress responses, morphological alterations, or traits (e.g. seed size, etc.). Results In this study, we identified 169,197 fusion transcripts in 2795 transcriptome datasets of Arabidopsis thaliana, Cicer arietinum, and Oryza sativa by using a combination of tools, and confirmed the translational activity of 150 fusion transcripts through proteomic datasets. Analysis of the FT junction sequences and their association with epigenetic factors, as revealed by ChIP-Seq datasets, demonstrated an organised process of fusion formation at the DNA level. We investigated the possible impact of three-dimensional chromatin conformation on intra-chromosomal fusion events by leveraging the Hi-C datasets with the incidence of fusion transcripts. We further utilised the longread RNA-Seq datasets to validate the most reoccurring fusion transcripts in each plant species followed by further authentication through RT-PCR and Sanger sequencing. Conclusions Our findings suggest that a significant portion of fusion events may be attributed to alternative splicing during transcription, accounting for numerous fusion events without a proportional increase in the number of RNA pairs. Even non-nuclear DNA transcripts from mitochondria and chloroplasts can participate in intra- and inter-chromosomal fusion formation. Genes in close spatial proximity are more prone to undergoing fusion formation, especially in intra-chromosomal FTs. Most of the fusion transcripts may not undergo translation and serve as long non-coding RNAs. The low validation rate of FTs in plants indicated that the fusion transcripts are expressed at very low levels, like in the case of humans. FTs often originate from parental genes involved in essential biological processes, suggesting their relevance across diverse tissues and stress conditions. This study presents a comprehensive repository of fusion transcripts, offering valuable insights into their roles in vital physiological processes and stress responses.
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    Variety-specific transcript accumulation during reproductive stage in drought- stressed rice
    (John Wiley & Sons, 2022) Gour, Pratibha; Kansal, Shivani; Agarwal, Priyanka; Mishra, Bhuwaneshwar Sharan; Sharma, Deepika; Mathur, Saloni; Raghuvanshi, Saurabh
    The divergence of natural stress tolerance mechanisms between species is an intriguing phenomenon. To study it in rice, a comparative transcriptome analysis was carried out in ‘heading’ stage tissue (flag leaf, panicles and roots) of Nagina 22 (N22; drought-tolerant) and IR64 (drought-sensitive) plants subjected to field drought. Interestingly, N22 showed almost double the number of differentially expressed genes (DEGs) than IR64. Many DEGs colocalized within drought-related QTLs responsible for grain yield and drought tolerance and also associated with drought tolerance and critical drought-related plant traits such as leaf rolling, trehalose content, sucrose and cellulose content. Besides, co-expression analysis of the DEGs revealed several ‘hub’ genes known to actively regulate drought stress response. Strikingly, 1366 DEGs, including 21 ‘hub’ genes, showed a distinct opposite regulation in the two rice varieties under similar drought conditions. Annotation of these variety-specific DEGs (VS-DEGs) revealed that they are distributed in various biological pathways. Furthermore, 103 VS-DEGs were found to physically interact with over 1300 genes, including 32 that physically interact with other VS-DEGs as well. The promoter region of these genes have sequence variations among the two rice varieties, which might be in part responsible for their unique expression pattern.
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    The landscape and implications of chimeric RNAs in cervical cancer
    (Elsevier B.V., 2018) Wu, Peng; Yang, Shuo; Singh, Sandeep; Qin, Fujun; Kumar, Shailesh; Wang, Ling; Ma, Ding; Li, Hui
    Background: Gene fusions and fusion products have been proven to be ideal biomarkers and drug targets for cancer. Even though a comprehensive study of cervical cancer has been conducted as part of the Cancer Genome Atlas (TCGA) project, few recurrent gene fusions have been found, and none above 3% of frequency. Methods: We believe that chimeric fusion RNAs generated by intergenic splicing represent a new repertoire of biomarkers and/or therapeutic targets. However, they would be missed when only genome sequences and fusions at DNA level are considered. We performed extensive data mining for chimeric RNAs using both our and TCGA cervical cancer RNA-Seq datasets. Multiple criteria were applied. We analyzed the landscape of chimeric RNAs at various levels, and from different angles. Findings: The chimeric RNA landscape changed as different filters were applied. 15 highly frequent (N10%) chimeric RNAs were identified. LHX6-NDUFA8 was detected exclusively in cervical cancer tissues and Pap smears, but not in normal controls. Mechanistically, it is not due to interstitial deletion, but a product of cis-splicing between adjacent genes. Silencing of another recurrent chimera, SLC2A11-MIF, resulted in cell cycle arrest and reduced cellular proliferation. This effect is unique to the chimera, and not shared by the two parental genes. Interpretation: Highly frequent chimeric RNAs are present in cervical cancers. They can be formed by intergenic splicing. Some have clear implications as potential biomarkers, or for shedding new light on the biology of the disease. Fund: Stand Up To Cancer and the National Science Foundation of China
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    De novo transcriptome analysis of medicinally important Plantago ovata using RNA-seq
    (PLOS, 2016) Kotwal, Shivanjali; Kaul, Sanjana; Sharma, Pooja; Gupta, Mehak; Shankar, Rama; Jain, Mukesh; Dhar, Manoj K.
    Plantago ovata is an economically and medicinally important plant of the family Plantaginaceae. It is used extensively for the production of seed husk for its application in pharmaceutical, food and cosmetic industries. In the present study, the transcriptome of P. ovata ovary was sequenced using Illumina Genome Analyzer platform to characterize the mucilage biosynthesis pathway in the plant. De novo assembly was carried out using Oases followed by velvet. A total of 46,955 non-redundant transcripts (≥100 bp) using ~29 million high-quality paired end reads were generated. Functional categorization of these transcripts revealed the presence of several genes involved in various biological processes like metabolic pathways, mucilage biosynthesis, biosynthesis of secondary metabolites and antioxidants. In addition, simple sequence-repeat motifs, non-coding RNAs and transcription factors were also identified. Expression profiling of some genes involved in mucilage biosynthetic pathway was performed in different tissues of P. ovata using Real time PCR analysis. The study has resulted in a valuable resource for further studies on gene expression, genomics and functional genomics in P. ovata.
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    Comprehensive analysis of small RNA-seq data reveals that combination of miRNA with its isomiRs increase the accuracy of target prediction in Arabidopsis thaliana
    (Taylor & Francis Group, 2014) Ahmed, Firoz; Senthil-Kumar, Muthappa; Lee, Seonghee; Dai, Xinbin; Mysore, Kirankumar S; Zhao, Patrick Xuechun
    Along with the canonical miRNA, distinct miRNA-like sequences called sibling miRNAs (sib-miRs) are generated from the same pre-miRNA. Among them, isomeric sequences featuring slight variations at the terminals, relative to the canonical miRNA, constitute a pool of isomeric sibling miRNAs (isomiRs). Despite the high prevalence of isomiRs in eukaryotes, their features and relevance remain elusive. In this study, we performed a comprehensive analysis of mature precursor miRNA (pre-miRNA) sequences from Arabidopsis to understand their features and regulatory targets. The influence of isomiR terminal heterogeneity in target binding was examined comprehensively. Our comprehensive analyses suggested a novel computational strategy that utilizes miRNA and its isomiRs to enhance the accuracy of their regulatory target prediction in Arabidopsis. A few targets are shared by several members of isomiRs; however, this phenomenon was not typical. Gene Ontology (GO) enrichment analysis showed that commonly targeted mRNAs were enriched for certain GO terms. Moreover, comparison of these commonly targeted genes with validated targets from published data demonstrated that the validated targets are bound by most isomiRs and not only the canonical miRNA. Furthermore, the biological role of isomiRs in target cleavage was supported by degradome data. Incorporating this finding, we predicted potential target genes of several miRNAs and confirmed them by experimental assays. This study proposes a novel strategy to improve the accuracy of predicting miRNA targets through combined use of miRNA with its isomiRs.