Pan-genome analysis reveals structural variation- associated expression and evolutionary diversity of the ZmCYP450...

The plant cytochrome P450 (CYP450) superfamily plays a key role in metabolic diversity and environmental adaptation; however, systematic analyses of its intraspecific structural variation, copy number dynamics, and evolutionary mechanisms remain limited. Using 27 high-quality maize reference genomes, we performed a pan-genomic analysis of the ZmCYP450 family, identifying 282 orthogroups (OGs) and 7,282 genes. The family exhibits a pattern of predominantly conserved genes with localized expansions and open pan-genome properties. PAV and CNV analyses revealed extensive gene deletions and copy number fluctuations outside core OGs, reflecting substantial intraspecific structural diversity. Analysis of duplication types and local colinearity suggested that proximal and dispersed duplications are the primary contributors to drive family expansion. Ka/Ks analysis indicated that most OGs are under purifying selection, while a subset shows evidence of positive selection. Further integration of structural variation and transcriptomic data suggested that SVs may affect gene function through mechanisms such as gene deletion, protein truncation, and remodeling of regulatory elements, suggesting a dual role of potential loss of function and expression modulation. Despite a relatively stable overall copy number, structural variant categories ('Typical', 'Atypical', and 'Missing') are widespread, and expression levels do not always correlate with copy number, suggesting a complex regulatory patterns. Tissue-specificity analysis revealed a large number of highly specific ZmCYP450 genes involved in secondary metabolism and environmental responses, with distinct expression profiles across different genetic backgrounds. This study provides a comprehensive pan-genomic view of structural variation, evolutionary patterns, and expression regulation in the ZmCYP450 family, offering a foundation for future functional studies and maize trait improvement.