Functional analysis of plant RNaseIII enzymes
Nahid Shamandi
Abstract:
Small RNAs, including miRNA and siRNA, play essential regulatory roles in genome stability, development and stress responses in most eukaryotes. Plants encode four DICER-LIKE (DCL) RNaseIII enzymes. DCL1 produces miRNAs, while DCL2, DCL3 and DCL4 produce diverse size classes of siRNA. Plants also encode RNASE THREE-LIKE (RTL) enzymes that lack DCL-specific domains and whose function is largely unknown. Arabidopsis plants over-expressing RTL1 exhibit morphological defects and lack all types of small RNAs produced by DCL2, DCL3 and DCL4, indicating that RTL1 is a general suppressor of plant siRNA pathways. RTL1 activity requires a functional RNaseIII domain. RTL1 is naturally expressed only weakly in roots, but virus infection strongly induces its expression in leaves, suggesting that RTL1 induction is a general strategy used by viruses to counteract the siRNA-based plant antiviral defense. Accordingly, transgenic plants over-expressing RTL1 are more sensitive to TYMV infection than wild-type plants, likely because RTL1 prevents the production of antiviral siRNAs. However, TCV, TVCV and CMV, which encode stronger suppressors of RNA silencing (VSR) than TYMV, are insensitive to RTL1 over-expression. Indeed, TCV VSR inhibits RTL1 activity, suggesting that inducing RTL1 expression and dampening RTL1 activity is a dual strategy used by viruses to establish a successful infection. Plants over-expressing RTL2 and rtl2 mutants do not exhibit morphological defects and do not show major changes in the endogenous small RNA repertoire. However, RTL2 over-expression enhances the accumulation of exogenous siRNAs in transient assays, and this activity requires a functional RNaseIII domain. Therefore, it is possible that plant RTL2 processes certain substrates to facilitate the action of DCL enzymes.
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Thesis of the University Paris Sud
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