Genome-wide identification and expression analysis of HMAs gene family in tobacco under abiotic stress

The heavy metal ATPase (HMA) family, classified as P_1B-ATPases, plays a pivotal role in maintaining plant metal homeostasis by coupling ATP hydrolysis to the active transport of metal ions across biological membranes. However, comprehensive characterization of the HMA family in tobacco (Nicotiana tabacum) remains insufficient. In this study, we conducted a systematic genome-wide bioinformatic analysis and identified 27 HMA-encoding genes in tobacco, designated NtHMA1 through NtHMA27. Phylogenetic and structural analyses revealed that these genes fall into two major clades, further resolved into six evolutionarily conserved subfamilies. Chromosomal localization showed that NtHMA genes are predominantly distributed on chromosomes 1, 17, and 25, and all retain hallmark domains characteristic of the HMA family. Comparative genomic analysis identified ten segmentally duplicated NtHMA gene pairs, all exhibiting signatures of purifying selection (Ka/Ks < 1), suggesting functional constraint and evolutionary conservation. Cis-regulatory element analysis uncovered 14 types of stress-responsive cis-acting elements associated with responses to biotic and abiotic stresses. Integrated transcriptomic profiling across multiple abiotic stress conditions and heavy metal treatments indicated that NtHMA6, NtHMA14, and NtHMA23 are preferentially upregulated under heavy metal exposure, implicating their roles in metal transport and detoxification, while NtHMA2, NtHMA7, NtHMA10, NtHMA11, NtHMA13, NtHMA18, NtHMA22, and NtHMA25 show significant transcriptional responsiveness to abiotic stresses or hormone stimuli, suggesting involvement in stress adaptation. In summary, this study provides the first comprehensive genomic, structural, evolutionary, and expression-based characterization of the tobacco NtHMA family, establishing a foundation resource for functional genomics studies and offering valuable insights to guide molecular breeding strategies aimed at enhancing tobacco resilience and metal tolerance.