Abstract
Small RNA-mediated gene silencing pathways play important roles in the regulation of development, genome stability and various stress responses in many eukaryotes. Recently, a new type of small interfering RNAs (qiRNAs) approximately 20–21 nucleotides long in Neurospora crassa have been shown to mediate gene silencing in the DNA damage response (DDR) pathway. However, the mechanism for RNA silencing in the DDR pathway is largely unknown in plants. Here, we report that a class of small RNAs (qiRNAs) derived from rDNA was markedly induced after treatment by DNA-damaging agents [ethyl methanesulphonate (EMS and UV-C)], and that aberrant RNAs (aRNAs) as precursors were also highly induced after the DNA damage treatment in rice. However, these RNAs were completely abolished in OsRecQ1 (RecQ DNA helicase homologue) and OsRDR1 (RNA-dependent RNA polymerase homologue) mutant lines where either gene was disrupted by the insertion of rice retrotransposon Tos17 after the same treatment. DNA damage resulted in a more significant increase in cell death and a more severe inhibition of root growth in both mutant lines than in the WT. Together, these results strongly suggest that both OsRecQ1 and OsRDR1 play a pivotal role in the aRNA and qiRNA biogenesis required for the DDR and repair pathway in rice, and it may be a novel mechanism of regulation to the DDR through the production of qiRNA in plants.
Highlights
Chromosomal DNA damage in most organisms is caused by two major sources from exogenous factors such as ultraviolet light (UV), ionizing radiation and chemical exposure [1,2], as well as through endogenous cellular processes such as cellular metabolism and replication errors [3,4]
Northern blot analysis with an RNA probe specific for the antisense 25S rDNA region showed that a class of small RNAs about 20–21 nucleotides in length was significantly induced after UV or EMS treatment, but it was at an undetectable level under normal conditions in WT (Figure 1A), and a similar result was obtained using an RNA probe specific for the sense 25S rDNA region, suggesting that these small RNA are double stranded
These results suggest that OsRecQ1 is required for qiRNA biogenesis in the DNA damage response (DDR) pathway
Summary
Chromosomal DNA damage in most organisms is caused by two major sources from exogenous factors such as ultraviolet light (UV), ionizing radiation and chemical exposure [1,2], as well as through endogenous cellular processes such as cellular metabolism and replication errors [3,4]. Many proteins acting as sensors, transducers or effectors are required for cell cycle checkpoint regulation, DNA repair and apoptosis in different DDR pathways [9]. The anthrax toxin receptor (ATR) and ataxia telangiectasia mutated (ATM) protein kinases have known to be involved in a wide variety of responses to DNA damage in plants [12], both ATM and ATR play central roles in the cellular response to DSBs by regulating DNA repair, cell-cycle arrest and apoptosis [13], and suppressor of gamma response 1 (SOG1) participates in pathways governed by both ATR and ATM sensor kinases in plants [9]. A novel protein in mammals, RHINO (Rad, Rad, Hus interacting nuclear orphan) is shown to be required for ATR (ataxia telangiectasia and the Rad3-related) signaling and cell cycle checkpoint activation in the DDR pathway [14], and WolfHirschhorn syndrome candidate 1 (WHSC1) gene in human cells recruited to sites of DNA damage in the DDR [15]
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