Identification and expression analysis of miR396 and its target genes in Jerusalem artichoke under temperature stress.

The highly conserved miR396 plays a pivotal role in the growth, development, and responses to abiotic and biotic stresses in plants. However, research on miR396 and its targets in Jerusalem artichoke remains largely unexplored. In this study, we employed bioinformatics and experimental techniques, such as cloning and qRT-PCR, to investigate the regulatory role of miR396 on its targets, leveraging our lab's transcriptomic and degradomic data of Jerusalem artichoke. Specifically, we initially cloned and characterized the precursors (htu-MIR396a/b/c) and mature sequences (htu-miR396a/b/c) of three miR396 isoforms. Subsequently, we identified nine target genes, including seven Growth-Regulating Factors (GRFs) (HtGRF3/4/6/9/10/12/13), one WRKY transcription factor (HtWRKY40), and one Scarecrow-like (SCL) transcription factor (HtSCL33). Finally, we conducted an analysis of their expression patterns across various tissues and their responses to temperature stress. Notably, htu-MIR396s exhibited high expression in seedling stems, while htu-miR396s predominantly expressed in seedling leaves. Moreover, HtWRKY40 and HtSCL33 displayed higher expression levels than HtGRFs in most tissues, except leaves. Remarkably, HtGRF4/6/10/12/13 exhibited higher expression in leaves than in roots and stems during seedling growth. Furthermore, during tuber development, HtGRF4/6/10, HtWRKY40, and HtSCL33 were highly expressed, while HtGRF3/9/12/13 showed relatively lower expression levels. Under heat stress (42℃), htu-MIR396 expression was up-regulated, and htu-miR396 showed dynamic expression patterns in seedlings, resulting in the induction of HtGRF4/6/10/12/13 in leaves and HtSCL33 in roots, while HtWRKY40 in leaves was repressed. Conversely, under cold stress (4℃), htu-MIR396s showed fluctuating expression levels, and htu-miR396s were up-regulated in seedlings. Notably, HtGRF4/13 and HtSCL33 in seedlings were reduced, whereas HtGRF6 in roots and HtWRKY40 in leaves were enhanced. These findings offer valuable insights into the functional roles of miR396-target interactions under abiotic stress in Jerusalem artichoke.

Unveiling HSP40/60/70/90/100 gene families and abiotic stress response in Jerusalem artichoke.

Heat shock proteins (HSPs) are essential for plant growth, development, and stress adaptation. However, their roles in Jerusalem artichoke are largely unexplored. Using bioinformatics, we classified 143 HSP genes into distinct families: HSP40 (82 genes), HSP60 (22 genes), HSP70 (29 genes), HSP90 (6 genes), and HSP100 (4 genes). Our analysis covered their traits, evolution, and structures. Using RNA-seq data, we uncovered unique expression patterns of these HSP genes across growth stages and tissues. Notably, HSP40, HSP60, HSP70, HSP90, and HSP100 families each had specific roles. We also studied how these gene families responded to various stresses, from extreme temperatures to drought and salinity, revealing intricate expression dynamics. Remarkably, HSP40 showed remarkable flexibility, while HSP60, HSP70, HSP90, and HSP100 responded specifically to stress types. Moreover, our analysis unveiled significant correlations between gene pairs under stress, implying cooperative interactions. qRT-PCR validation underscored the significance of particular genes such as HtHSP60-7, HtHSP90-5, HtHSP100-2, and HtHSP100-3 in responding to stress. In summary, our study advances the understanding of how HSP gene families collectively manage stresses in Jerusalem artichoke. This provides insights into specific gene functions and broader plant stress responses.