Comparative transcriptome reveals molecular insights into the role of beneficial fungal Trichoderma sp. alleviating Cd toxicity in barley

Soil cadmium (Cd) pollution poses a serious threat to agricultural production and food safety, becoming a worldwide agricultural and environmental issue. We previously identified a beneficial fungal strain, Trichoderma nigricans T32781, which effectively alleviates Cd stress in plants. Here, we investigated gene expression profiles of barley in response to T32781 inoculation under both controlled and open-field conditions with Cd treatment. Comparative transcriptomic analysis revealed T32781-induced differential gene expression in response to Cd stress, identifying 443/91 and 561/623 up/down-regulated differentially expressed genes (DEGs) in barley leaves from the pot and field trials, respectively. Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that these DEGs are predominantly involved in plant-pathogen interaction and photosynthesis pathways. Therefore, we proposed an integrated schematic diagram according to the identified Cd tolerance-associated DEGs. Further comparative analysis of DEGs across different experimental conditions identified 9 shared key genes, including a novel aspartic proteinase PCS1-like (HvPCS1L) from a gene family of 20 members in barley genome. Bioinformatic characterization suggested a potential evolutionary origin of HvPCS1Ls from green algae with the conserved psi loop motif. Functional analysis using virus-induced gene silencing (VIGS) of HvPCS1L-11 showed significant increases of Cd concentration and accumulation in shoots of silenced lines, demonstrating its crucial role in Cd absorption in barley. Our findings provide novel insights into the molecular mechanisms and regulatory pathways through which the novel fungal strain T32781 alleviates Cd tolerance and accumulation in barley.