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Targeted protein analysis of WT and erf34 mutant Arabidopsis using PRM (Parallel Reaction Monitoring) method

ABSTRACT: After seven days of cultivation under continuous light on 1/2 MS medium, Arabidopsis thaliana seedlings were subjected to a 3-hour dark treatment in liquid 1/2 MS medium. Subsequently, they were transferred to 1/2 MS liquid medium supplemented with 100 μM ABA for 3h

ORGANISM(S): Arabidopsis Thaliana

SUBMITTER: Zecheng Zuo

PROVIDER: PXD048436 | iProX | Tue Jan 09 00:00:00 GMT 2024

REPOSITORIES: iProX

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Project description:In Arabidopsis thaliana, four different DICER-LIKE (DCL) proteins have distinct, but partially overlapping functions in the biogenesis of microRNAs (miRNAs) and small interfering RNAs (siRNAs) from longer, non-coding precursor RNAs. To analyze the impact of different components of the small RNA (sRNA) biogenesis machinery on the transcriptome, we subjected dcl and other mutants impaired in sRNA biogenesis to whole-genome tiling array analysis. We compared both protein-coding genes and noncoding transcripts, including most pri-miRNAs, in two tissues and several stress conditions. We discovered distinct effects of dcl1, hyl1 and se mutations on the transcriptome, as well as a number of common genes affected in dcl1 and dcl2 dcl3 dcl4 triple mutants. Our results furthermore suggest that the DCL1 is not only involved in miRNA action, but can also contribute to silencing of certain transposons, apparently through an effect on DNA methylation. Together, our findings contribute to the knowledge of both specialization and overlap between different RNA silencing pathways. Wild type and dcl1-100, se-3, hyl1-2, rdr6-15 ans dcl2,3,4 mutants were grown on MS or soil at 23M-BM-0C and LL. 7 day old seedlings (grown on MS plates) and inflorescences from 28 (>40) day old plants (dcl1) were used for RNA extraction. For stress treatments, seedlings had been grown for 10 days on solid MS medium at 21M-BM-0C, they were transferred to liquid MS medium containing no additives (mock control), 200 mM NaCl (salt stress), 300 mM mannitol (osmotic stress) or 100 M-NM-<M ABA. For cold and heat stress, seedlings were transferred to pre-chilled or pre-warmed liquid MS medium and incubated at 8M-BM-0C and 30M-BM-0C, respectively. Samples were taken after 1 h and 12 h of continuous stress treatment. RNA was extracted from whole seedlings. All RNA samples were converted into double stranded DNA targets that were hybridized to whole genome tiling arrays (Affymetrix Arabidopis Tiling1.0RM-BM-.). All hybridizations were performed with 3 biological replicates.

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Targeted protein analysis of WT and erf34 mutant Arabidopsis using PRM (Parallel Reaction Monitoring) method

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