miRNA-CNGC/GLR circuits coordinate heat-stress Ca²⁺ signaling in wheat

Calcium (Ca²⁺) signatures are central to plant stress signaling, and GLR and CNGC channels are major Ca²⁺ entry routes. We integrated comparative phylogenomics, protein and promoter analyses, localization prediction, protein-interaction inference, miRNA-target prediction, and RT-qPCR validation to examine the regulation of wheat GLR/CNGCs under heat stress. Phylogenies separated GLR and CNGC lineages and revealed wheat-specific expansion consistent with polyploid retention. TaCNGCs were generally alkaline and more often predicted as unstable, whereas TaGLRs showed broader pI variation and greater predicted stability. Most proteins were predicted to localize to the plasma membrane, with a subset predicted to localize to chloroplasts. Promoters were enriched for light, ABA/MeJA/auxin, MYB drought, and anaerobic/energy-stress motifs. Network analyses highlighted tae-miR399, tae-miR167a, tae-miR156a, tae-miR164, and tae-miR171a as inferred regulatory hubs targeting selected TaGLR and TaCNGC transcripts. RT-qPCR across 0-72 h heat exposure showed early miRNA induction, reciprocal repression of many channel transcripts, and genotype-dependent recovery. The tolerant genotype displayed stronger transient miRNA activation and clearer channel transcript rebound by 48-72 h. These data support an inferred, reversible miRNA-channel regulatory model during heat acclimation and nominate candidates for direct target and functional validation.