Small RNAs, including microRNAs (miRNAs) and small interfering RNAs (siRNAs), are geared to respond to the complexity of developmental and physiological processes and negatively regulated gene expression by guiding chromatin modification mRNA degradation, and translational repression of their targets.
Increasing evidence indicates that host small RNAs play a crucial role during the plant-pathogen interactions. In the first chapter of this thesis, we reviewed pathogen-responsive endogenous miRNAs and siRNAs in plants. In addition, we discussed the important components involved in the canonical RNA silencing pathways, including dsRNA-specific endoribonuclease (RNase) Dicer-like proteins (DCLs), RNA-dependent RNA polymerases (RDRs) and Argonaute (AGO) Proteins.
MiRNAs are short noncoding RNA molecules with 21-24 nucleotides and are found in most eukaryotic cells. In the second chapter of this thesis, we globally profiled plant miRNAs in response to infection of bacterial pathogen Pseudomonas syringae pv. tomato (Pst) in Arabidopsis and found 19 miRNA precursors that can yield multiple distinct miRNA-like RNAs in addition to miRNAs and miRNA*s. These miRNA precursor-derived miRNA-like RNAs are often arranged in phase and form duplexes with approximately 2 nucleotides 3'-end overhang. These miRNA-like RNAs are potentially functional at least under certain conditions.
In the third chapter of this thesis, we identified 15, 27 and 20 miRNA families being differentially regulated after Pst DC3000 hrcC, Pst DC3000 EV and Pst DC3000 avrRpt2 infections, respectively, through the deep sequencing results from 13 small-RNA libraries. Some bacteria-regulated miRNAs targets have been annotated and believed to be involved in plant hormone biosynthesis and signaling pathways, including auxin, abscisic acid, and jasmonic acid pathways, which suggests the regulation of plant innate immunity through fine-tuning of multiple plant hormone pathways. Moreover, we discussed the procedural differences between sequencing-based profiling and small RNA Northern blot and stated the possibilities that cause the inconsistency.
AtlsiRNA-1, as the first example of the long siRNAs identified in Arabidopsis, contributes to resistance against Pseudomonas syringae infection by degrading the negative regulator of disease resistance, AtRAP. How does AtRAP regulate defense response became the major focus of my study to elucidate the mechanisms of plant innate immunity. In the fourth chapter of this thesis, I further investigated the molecular mechanism of AtRAP by characterizing the subcellular localization of AtRAP and determining the function of C-terminal RAP domain in disease resistance.