## 3 Discussion  
The TPS families in the plant kingdom are highly diversified [20]. In this study, we performed the first large-scale assessment of the number of TPS-subgroup members in 115 angiosperms (Tables S1 and S2). Despite the key roles that whole-genome duplication (WGD) and whole-genome triplication (WGT) events often have in increasing gene family size [47], we did not find evidence for such effects on TPS families (Figure 1). A previous study suggested that the large-scale expansion of the TPS family occurred after the divergence of dicot and monocot plants [48]. Our data support this conclusion. Moreover, we determined that the expansion of the TPS family was mainly due to the expansion of the TPS-a and TPS-b subgroups. The expansion of TPS-a suggests a correlation between the rate of evolutionary change--reflected by longer branch lengths--and the diversification of the TPS-a genes. These findings imply that species undergoing rapid evolutionary changes, as indicated by their placement on long branches of the phylogenetic tree, may have experienced selective pressures favoring the expansion of their TPS-a gene repertoire. Such expansions could be adaptive, potentially enhancing the species' ability to synthesize a diverse array of terpenoids, which are critical for various ecological functions including defense mechanisms and pollinator attraction. However, there were several exceptions. For instance, _L. chinense_, a magnoliid plant, contains 58 TPSs, including 24 TPS-a subgroup members. An expansion of the TPS-a subgroup is thought to have occurred after the split of monocots and dicots. The TPS-a subgroup mainly contains sesquiterpene synthases and is highly divergent in all seed plants [20]. We determined that the expansion of TPS-a occurred in monocots, super-rosids, and super-asterids after their split. The expansion of TPS-a in _L. chinense_ appears to be unique compared to the other species, which is worth exploring. The significant expansion of the TPS-a subgroup likely reflects multiple biological mechanisms. First, ecological pressures such as co-evolution with specialized herbivores may drive selection for sesquiterpene diversity, as evidenced by TPS-a copy number variation across maize populations adapting to distinct pest pressures [49]. Second, genomic analyses in Arabidopsis reveal TPS-a clusters flanked by transposable elements, suggesting repeated-mediated duplication as a key expansion driver [50]. Finally, subfunctionalization through promoter divergence could enable tissue-specific expression partitioning of defense-related terpenoids [51]. These mechanisms collectively suggest that TPS-a expansions represent adaptive genomic responses to biotic challenges. The variable number of genes among taxa points to the independent loss or duplication of genes in different genomes. The number of TPSs in the TPS-b subgroup appears to be irregular and this subgroup has been lost in many Poaceae species (Figure 1). The lost functionalities due to the loss of TPS-b, such as monosynthe activities, might be replaced by those of other subgroups. Notably, the absence of TPS-b raises questions about how Poaceae compensate for the loss of these volatile compounds. One possibility is that grasses have evolved alternative biosynthetic routes: TPS-g members might acquire monoterpene synthase activity through structural convergence, or non-TPS enzymes such as prenyltransferase could generate monoterpene precursors. Additionally, ecological shifts may reduce dependency on monoterpene; the prevalence of wind pollination diminishes the need for floral volatiles, while silica-based physical defenses in grasses might offset the loss of monoterpene-mediated chemical protection. Synteny could reflect important relationships between the genomic contexts of genes in terms of both function and regulation [52; 53]. Therefore, analyzing the syntenic relationships of genes across a wide range of species provides important information about the evolution of gene families involved in plant growth and development.