Genome-wide analysis of plant-specific calmodulin-binding IQD proteins in tobacco identifies key regulators of drought stress responses

Abstract Background Plants rely on complex signaling networks to perceive and respond to environmental stresses. Calcium ions (Ca 2+ ), acting as central second messengers, have their signals decoded by sensors such as calmodulin (CaM) and relayed to downstream target proteins. IQ67-Domain (IQD) proteins are a class of plant-specific CaM-binding proteins that function as molecular scaffolds integrating calcium signals with diverse cellular processes, playing crucial roles in growth, development, and stress responses. However, the IQD gene family has not yet been systematically investigated in tobacco ( Nicotiana tabacum ), an important economic crop and model plant species. Results In this study, a total of 77 NtIQD genes were identified in the tobacco genome through genome-wide analysis. Phylogenetic analysis classified them into four subgroups, with Group IV showing species-specific expansion within Solanaceae. Analyses of gene structure and conserved motifs revealed that while maintaining the core IQ domain, the family exhibits diversity in auxiliary motifs and domains. Cis-acting element analysis of promoters indicated that all NtIQD genes are enriched with elements responsive to light, phytohormones (e.g., jasmonic acid, abscisic acid), and abiotic stresses. Expression profiling demonstrated that NtIQD genes exhibit significant tissue specificity and developmental-stage specificity, and display differentiated response patterns to various stresses such as drought, cold, and pathogen infection, suggesting functional specialization. Protein–protein interaction network predictions implicated their potential role as signaling hubs. Further experimental validation showed that NtIQD1 , NtIQD13 , and NtIQD42 localize to distinct cellular compartments. Virus-induced gene silencing (VIGS) of these three genes significantly increased drought sensitivity in tobacco plants, confirming their positive regulatory roles in drought resistance. Conclusion This study presents the first comprehensive genome-wide identification and analysis of the IQD gene family in tobacco. It reveals the diversity of this family in evolution, structure, and expression, and validates the key functions of NtIQD1 , NtIQD13 , and NtIQD42 in drought stress response. These findings provide new insights into the decoding and transduction mechanisms of calcium signaling in Solanaceae plants and offer valuable candidate gene resources for crop genetic improvement aimed at enhancing stress resistance. Graphical abstract