Identification of the core member of the HAKs family and primary analysis on their functions in allotetraploid rapeseed (Brassica napus L.)

Objectives The HAKs (high-affinity K transporter) family genes, encode high-affinity potassium (K⁺) transporters, play key roles in the absorption and transport of K⁺. Identification of the core genes of the HAKs family in rapeseed, and the study of their expression responses when exposure to different nutrient stresses, will lay deeper the understanding of the functions of HAKs, and also provide valuable genetic resource references for the genetic improvement of HAKs-mediated nutrient resistance in rapeseed.

Methods The genome-wide identification and analysis of the rapeseed HAKs gene family in this study included phylogenetic relationships, gene structure, conserved motifs, chromosomal localization, cis-acting elements. Differential expression of HAKs family genes were analyzed using transcriptomic data under different nutrient stresses, and the co-expression network was constructed by Cytoscape. Low-k⁺ resistant line “H280” and the low-k⁺ sensitive line “L49” were subjected to K stress in a hydroponic experiment, The K⁺ contents in the roots of line “H280” and “L49” were analyzed using ICP-MS. The response and function of BnaA7.HAK5 under low-K stress were analyzed by RT-qPCR and subcellular localization.

Results A total of 40 HAKs family members were identified through genome-wide identification. Evolutionary relationship analysis of the HAKs family genes showed that their Ka/Ks values were all less than 1.0. The analysis of cis-acting elements showed that MYB and the hormone response cis-acting regulatory element ABRE were highly enriched in the promoter region of the HAKs family genes. Collinear analysis showed that most of the HAKs genes in Brassica napus were intact during the evolutionary process. Differential expression gene analysis revealed that the expression levels of most HAKs family genes were significantly induced by low potassium stress. The co-expression network analysis showed that BnaA7.HAK5 responded to low potassium stress in oilseed rape. The RT-qPCR results showed that the expression level of BnaA7.HAK5 in the roots of low-potassium-resistant line “H280” was significantly higher than that of low-potassium-sensitive line “L49”. The results of subcellular localization showed that BnaA7.HAK5 was localized on the plasma membrane of cells.

Conclusions There are 40 BnaHAKs genes in rapeseed, and the BnaHAKs genes respond to nutrient stresses such as low potassium or salt. BnaA7.HAK5, localized in the plasma membrane, is significantly up-regulated under low-potassium conditions, indicating that BnaA7.HAK5 played a major role in the differential expression in root between “H280”and “L49” oilseed rape lines, it could be concluded that BnaA7.HAK5 plays an important role in the high-affinity uptake and transport of potassium ions in rapeseed.