Expression Profile Of MiRNA Of Populus Cathayana Rehd And Salix Matsudana Koidz Under Salt Stress

Soil salinization brings increasing environmental and ecological problems worldwide. Approximately 10% of global land has been salinized in different extent, covering over 10 million hectares in more than 100 countries. microRNA (miRNA) might be regulated by abiotic stresses, such as salinity. miRNA is highly conserved, endogenous, non-coding small RNA of about 18-24 nucleotides in length. As negative regulators, miRNA inhibit gene expressions at the post-transcriptional level by guiding cleavage or attenuating the translation of target genes, which affects organismal morphogenesis, development and adaptability to changes occurring in the environment.In this study, we have identified novel miRNA from P. cathayana Rehd by constructing a library of small (18–24 nt) RNA. After sequencing 691 small RNAs from P. cathayana Rehd, alignment to the Populus sequence database and secondary structure prediction were performed. These bioinformatic analyses identified 5 candidate miRNA that were designated ptc-miR801, ptc-miR74, ptc-miR102, ptc-miR367 and ptc-miR213. Expression of these miRNA was validated using Real-time quantitative PCR (qRT-PCR) analysis and northern blotting, and four of them displayed distinct expression patterns in leaves, stems and roots. Our results show that four of the miRNA are likely specific to leaves. Putative targets for these miRNA play important roles in development and response to abiotic stress. One such miRNAs (ptc-miR801) is predicted to target members of the NAC or Scarecrow-like (SCL) protein family which play an important role in salt stress. A candidate ptc-miR213 target is MYB4 (v-myb avian myeloblastosis oncogene ortholog), which may play an important role in plant stress tolerance. We analyzed the expression of the ptc-miR801 and ptc-miR213 genes in plants treated for 72 h with salt (100 mM NaCl) as well as untreated plants. Both genes were induced in response to salt stress.Plants under such stresses may express some specific miRNA with novel functions. To analyze the expression profile of miRNA, we took two different plants, P. cathayana Rehd (salt-sensitive plants) and S. matsudana Koidz (high salt-tolerant plants). miRNA microarray hybridization revealed that the expression of 161 miRNA changed in treated P. cathayana Rehd, and 32 miRNA changed in treated S. matsudana Koidz. These results suggest that the same miRNA has different expression patterns in salt-sensitive plants and salt-tolerant plants under salt stress. Furthermore, one miRNA may have different purposes in salt-sensitive plants and salt-tolerant plants. In our research, the expression of both ptc-miR474c and ptc-miR398b in P. cathayana Rehd was down-regulated under salt stress with log values of -1.71 and -0.56, respectively. Contrastingly, the expression of these same miRNAs was up-regulated in S. matsudana Koidz, with log values of 4.3 and 2.79, respectively. However, not all miRNAs had different expression patterns between salt-sensitive plants and salt-tolerant plants. To examine this, we used qRT-PCR to select five Populus cathayana Rehd and four Salix matsudana Koidz miRNAs and their family target genes that are involved in the responses of plants to abiotic salt stress, respectively. Specifically, 6 month-old in vitro cultured P. cathayana Rehd and S. matsudana Koidz plantlets were subjected to 0,3,6,9,12,48 and 72 h salt-stress. In summary, our results showed that some miRNAs were different expressed in P.cathayana Rehd and S.matsudana Koidz under salt stress. For example, the miR398 and miR474c family were down-regulated in P. cathayana Rehd, but up-regulated in S. matsudana Koidz in response to 72h salt stress. The results can be utilized in future research to study post-transcriptional gene regulation in P.cathayana Rehd and S.matsudana Koidz under salt stress.