• Kuang L, Yu J, Shen Q, Fu L, Wu L. (2021) Identification of microRNAs Responding to Aluminium, Cadmium and Salt Stresses in Barley Roots. Plants 10(12), 2754.

Plants are frequently exposed to various abiotic stresses, including aluminum, cadmium and salinity stress. Barley (Hordeum vulgare) displays wide genetic diversity in its tolerance to various abiotic stresses. In this study, small RNA and degradome libraries from the roots of a barley cultivar, Golden Promise, treated with aluminum, cadmium and salt or controls were constructed to understand the molecular mechanisms of microRNAs in regulating tolerance to these stresses. A total of 525 microRNAs including 198 known and 327 novel members were identified through high-throughput sequencing. Among these, 31 microRNAs in 17 families were responsive to these stresses, and Gene Ontology (GO) analysis revealed that their targeting genes were mostly highlighted as transcription factors. Furthermore, five (miR166a, miR166a-3p, miR167b-5p, miR172b-3p and miR390), four (MIR159a, miR160a, miR172b-5p and miR393) and three (miR156a, miR156d and miR171a-3p) microRNAs were specifically responsive to aluminum, cadmium and salt stress, respectively. Six miRNAs, i.e., miR156b, miR166a-5p, miR169a, miR171a-5p, miR394 and miR396e, were involved in the responses to the three stresses, with different expression patterns. A model of microRNAs responding to aluminum, cadmium and salt stresses was proposed, which may be helpful in comprehensively understanding the mechanisms of microRNAs in regulating stress tolerance in barley.

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• Su T, Fu L, Kuang L, Chen D, Zhang G, Shen Q, Wu D. (2022) Transcriptome-wide m6A methylation profile reveals regulatory networks in roots of barley under cadmium stress. Journal of Hazardous Materials 423, 127140.

Cadmium (Cd) pollutants restrict crop yield and food security in long-term agricultural activities. Crops have evolved adaptive strategies under Cd condition, however, the transcriptional regulatory mechanism of Cd-tolerant genes remains to be largely illustrated. In this study, barley roots were exposed to 5 µM CdCl2 for physiological response and transcriptome-wide m6A methylation profile. Cd stress inhibited root growth after 7 d Cd treatment, which is mainly associated with inhibited absorption of Mn. After Cd treatment, 8151 significantly modified m6A sites and 3920 differentially expressed genes were identified. Transcriptome-wide m6A hypermethylation was widely induced by Cd stress and enriched near the stop codon and 3′ UTR regions. Among 435 m6A modified DEGs, 319 hypermethylated genes were up-regulated and 84 hypomethylated genes were down-regulated, respectively, indicating a positive correlation of m6A methylation and expression. But well-known Cd transporter genes (HvNramp5, HvIRT1, HvHMA3, etc.) were not modified by m6A methylation, except for ABC transporters. We further found key Cd-responding regulatory genes were positively modulated with m6A methylation, including MAPK, WRKY and MYB members. This study proposed a transcriptional regulatory network of Cd stress response in barley roots, which may provide new insight into gene manipulation of controlling low Cd accumulation for crops.

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• Zeng J. (2019) Identification of microRNAs and their targets responding to low-potassium stress in two barley genotypes differing in low-K tolerance. Journal of Plant Physiology 234-235, 44-53.

MicroRNAs (miRNAs) have diverse and crucial roles in plant growth and development, including in the response to abiotic stresses. Although plant responses to K deficiency are well documented at the physiological and transcriptional levels, the miRNA-mediated post-transcriptional pathways are still not clearly elucidated. In this study, high-throughput sequencing and degradome analysis were performed using two barley genotypes differing in low-K tolerance (XZ149, tolerant and ZD9, sensitive), to determine the genotypic difference in miRNAs profiling. A total of 270 miRNAs were detected in the roots of XZ149 and ZD9 at 2 d and 10 d after low-K treatment, of which 195 were commonly found in both genotypes. Their targets were further investigated by bioinformatics prediction and degradome sequencing approach. The results showed that ata-miR1432-5p might act as a regulator participating in Ca2+ signaling pathways in response to low-K stress. The difference in the miR444/MADS-box model as well as pathways mediated by miR319/TCP4 and miR396/GRF could be attributed to high tolerance to low-K stress in XZ149. In addition, other conserved and novel miRNAs families associated with low-K tolerance were also detected. The current results provide molecular evidence for understanding the possible involvement of miRNAs in the regulation of low-K tolerance.

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• Kuang L. (2019) Identification of microRNAs responding to salt stress in barley by high-throughput sequencing and degradome analysis. Environmental and Experimental Botany 160, 59-70.

A deep understanding of the mechanisms underlying salt tolerance should be helpful for breeding of salt-tolerant crop cultivars. Our previous study identified some wild barley accessions with high salt tolerance. In this study, small RNA and degradome sequencing was performed to identify salt-responsive microRNAs (miRNAs) and their target genes in a Tibetan wild barley accession XZ16 and a cultivar Golden Promise (GP), differing in salt tolerance. Under salt stress, XZ16 showed higher salt tolerance than GP in terms of biomass and shoot Na+ accumulation. A total of 278 and 320 miRNAs were found in roots and shoots of both genotypes, respectively. Among them, 40 and 51 miRNAs were salt-responsive in roots and shoots, respectively. In roots, miR156d, miR164a, miR393a, miR319a and miR172b targeting UGTs, NAC079, HvAFB2/HvTIR1, TCP4 and HvAP2, respectively, are probably responsible for salt tolerance. In shoots, miR159a, miR169i, miR319a/miR396e module and miR172b regulating MYB33, NHX1/LEA7, TCP4, GRFs and HvAP2, respectively, might contribute to salt tolerance. Compared with GP, XZ16 showed differential expression of miR156d, miR164a, miR169i, miR172b, miR319a, miR393a and a novel miRNA PC-miR124. We proposed that these differentially expressed miRNAs could account for the difference in salt tolerance of the two genotypes and drawn up a putative regulatory network of miRNAs to reveal molecular mechanisms of salt tolerance in barley.

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• Yu J. (2019) Genotypic difference of cadmium tolerance and the associated microRNAs in wild and cultivated barley. Plant Growth Regulation 87(3), 389-401.

Little study was performed to know how microRNAs (miRNAs) are responsive to cadmium (Cd) stress in barley (Hordeum vulgare). In this study, 16 small RNA libraries of shoot and root tissues from a wild barley accession (WB-1) and cultivated barley (Golden Promise) with contrasting Cd tolerance were constructed and sequenced. Moreover, a degradome library was constructed and analyzed to identify target genes of the miRNAs. Based on high-throughput sequencing, 216 conserved miRNAs (in 59 miRNA families) and 87 novel miRNAs were identified. A total of 238 target genes for 149 miRNAs (113 conserved and 36 novel miRNAs) were detected by the degradome analysis. Among these miRNAs, 45 miRNAs (40 conserved and 5 novel miRNAs) and 43 miRNAs (40 conserved and 3 novel miRNAs) showed differential expression in roots and shoots of two genotypes under Cd conditions. Compared with cultivar Golden Promise, the wild genotype WB-1 had genotype-dependent responses of miR156, miR159, miR166, miR167, miR171 and miR393, which regulate target genes including SPL, MYB, HD-Zip, ARF, GRAS and TIR. Correspondingly, WB-1 had lower Cd concentration and stronger Cd tolerance than Golden Promise. It indicates that miRNAs may play critical roles underlying genotypic difference of Cd tolerance in barley.

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• Wu L. (2018) Identification of microRNAs in response to aluminum stress in the roots of Tibetan wild barley and cultivated barley. BMC Genomics 19(1), 560.

Barley is relatively sensitive to Aluminum (Al) toxicity among cereal crops, but shows a wide genotypic difference in Al tolerance. The well-known Al-tolerant mechanism in barley is related to Al exclusion mediated by a citrate transporter HvAACT1 (Al-activated citrate transporter 1). A 1-kb insertion in the promoter region of HvAACT1 gene results in a dramatic increase of its expression level, which only occurs in some Al-tolerant cultivars. However, Al-tolerant Tibetan wild barley accession XZ29 did not have the 1-kb insertion.

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