Abstract
Small RNAs derived from transfer RNAs were recently assigned as potential gene regulatory candidates for various stress responses in eukaryotes. In this study, we report on the cloning and identification of tRNA derived small RNAs from black pepper plants in response to the infection of the quick wilt pathogen, Phytophthora capsici. 5′tRFs cloned from black pepper were validated as highly expressed during P. capsici infection. A high-throughput systematic analysis of the small RNAome (sRNAome) revealed the predominance of 5′tRFs in the infected leaf and root. The abundance of 5′tRFs in the sRNAome and the defense responsive genes as their potential targets indicated their regulatory role during stress response in black pepper. The 5′AlaCGC tRF mediated cleavage was experimentally mapped at the tRF binding sites on the mRNA targets of Non-expresser of pathogenesis related protein (NPR1), which was down-regulated during pathogen infection. Comparative sRNAome further demonstrated sequence conservation of 5′Ala tRFs across the angiosperm plant groups, and many important genes in the defense response were identified in silico as their potential targets. Our findings uncovered the diversity, differential expression and stress responsive functional role of tRNA-derived small RNAs during Phytophthora infection in black pepper.
Highlights
Eukaryotic organisms respond to environmental stress with a specific gene expression programme at the messenger RNAs (mRNAs) and protein level
By further analysis from genomic transfer RNA (tRNA) database and tRNAdb, two small RNA (sRNA) of 23 and 19 nucleotide length were identified as the 5′tRNA derived small RNA fragments (tRF) of Ala-tRNACGC and Arg-tRNATCG, respectively (Figures 2A,B), while one sRNA candidate was identified as 3′tRF cleaved from the 3′end of Gly-tRNATCC with a typical 3′CCA end (Table 1) (Figure 2C)
Our results demonstrated the existence of small RNAs derived from tRNAs and their potential gene regulatory role during the P. capsici infection of black pepper
Summary
Eukaryotic organisms respond to environmental stress with a specific gene expression programme at the mRNA and protein level. Small RNAs (sRNAs) are produced as modulators of gene expression during stress responses. They are short, non-coding RNAs of 16–35 nt long that regulate diverse biological functions by guiding sequence-specific gene silencing at the transcriptional or post-transcriptional level (Bartel, 2004; Vaucheret, 2006; Padmanabhan et al, 2009). TRNA Derived sRNAs in Plant Defence sequencing technologies had greatly accelerated the discovery of new types of small non-coding RNAs of diverse origin such as tRNAs (Cole et al, 2009; Loss-Morais et al, 2013; Hackenberg et al, 2015), snoRNAs (Chen and Wu, 2009; Taft et al, 2009; Li et al, 2012), and rRNAs (Wei et al, 2013). Most of the sRNA types differed from one another, they were suggested to be interconnected in their mode of action (Morris and Mattick, 2014) and regulatory role in multitude of cellular processes in development, stress responses, metabolism, and maintenance of genome integrity of organisms (Ghildiyal and Zamore, 2009; Wan et al, 2014)
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