## 3 Discussion  
Short-chain prenyl diphosphate synthases are enzymes of the isoprenoid pathway that use IPP and DMAPP to produce central intermediates in the isoprenoid metabolism. Short-chain prenyl diphosphate synthases are represented by three enzymes: GPPS, FPPS, and GGPPS [26]. Isolation and functional characterization of GPPSs from different plant species have confirmed the existence of homodimeric and heterodimeric forms, which are comprised of two similar subunits and one GPPS.SSU with one GPPS.LSU, respectively [15]. From the wintersweet transcriptome database [23], we isolated five transcripts that were annotated as GPPS/GGPPS-like genes: one homodimeric GPPS (_CpGPPS_), two GPPS small subunits (_CpGPPS.SSU1_, _CpGPPS.SSU2_), and two GGPPS (_CpGGPPS1_, and _CpGGPPS2_) sequences (Table 1). Subsequent sequence analysis indicated that homodimeric _Cp_GPPS and both _Cp_GGPPSs have two conserved aspartate-rich motifs, containing FARM (DD(X)2-4D) and SARM (DDXXD) (Figure S1C,D; Table 1), which are essential for catalysis and substrate binding [27]. However, _Cp_GPPS.SSUs lack one or both of these motifs (Figure S1A,B; Table 1). Besides, the CzxxC motif is needed for physical interaction between both subunits of heterodimeric GPPS [13; 14; 15]. In the present study, two CzxxC motifs were found in _Cp_GPPS.SSUs and one motif in _Cp_GGPPSs (Figure S1A,B,D; Table 1), which is consistent with the previous findings from other plant species. The scent of the wintersweet flower is composed of different volatile compounds, of which \(\alpha\)-linaloid is the most abundant monoterene volatile compound, accounting for about 36% of the wintersweet floral scent [28]. Moreover, the formation of monoterenes possesses tissue specificity and is often related to the maturity of a given tissue [29; 30]. The wintersweet flower develops a strong and specific fragrance during flower opening [31]. Further, wintersweet flowers contain the maximum number of scent-emitting nectaries at the fully opened flower stage. Dominantly, the maximum number  
Figure 5: Interaction of _Cp_GPPS.SSUs with _Cp_GGPPSs and _At_GGPPS11. **(A)** Key: 1, BD-_Cp_/SSU1 + AD-empty; 2, BD-_Cp_/SSU1 + AD-_Cp_GGPPS1; 3, BD-_Cp_/SSU1 + AD-_Cp_GGPPS2; 4, BD-_Cp_/SSU1 + AD-_At_GGPPS11; 5, BD-Lam + AD-Rec-T (negative control); 6, BD-53 + AD-Rec-T (positive control); 7, BD-_Cp_/SSU2 + AD-_Cp_GGPPS1; 8, BD-_Cp_/SSU2 + AD-_Cp_GGPPS2; 9, BD-_Cp_/SSU2 + AD-_At_GGPPS11; 10, BD-_Cp_/SSU2 + AD-empty. **(B)** The _Cp_GPPS.SSUs were fused with the DNA binding domain (BD) of PGBKT7 and GGPPSs were fused with the activation domain (AD) of the PGADT vector. Yeast strain AH109 co-transformed pairwise with both binding and activation vectors and spotted on the minimal synthetic dropout (SD) medium without leucine and tryptophan. **(C)** Yeast strains relative to those in the left panel were spotted on the SD medium lacking adenine, histidine, leucine, and tryptophan supplied with X-\(\alpha\)-Gal. of scent-emitting nectaries resides on petals during flower development [32]. In the present study, _Cp_GPPS/GGPPS genes were expressed in all tested tissues, with varying levels (Figure 2). Namely, the _CpGPPS.SSU1_ and _CpGGPPS1_ were highly expressed in the petal and flower, suggesting that these genes could have an important role in scent-producing tissues. Temporal expression analysis revealed that all _Cp_GPPS/GGPPS genes, except _CpGGPPS2_, showed the maximum expression at fully opened flower stage, followed by reduction at the senescence stage (Figure 3). These expression profiles are consistent with the finding of [PERSON] et al. [21], who reported that the _C. praccox_ L. 'H29' flowers had the maximum monoterpene contents at the partially opened (58.06%) and fully opened stage flowers (57.89%), followed by a sudden decline at senescence (9.44%). Overall, these results suggest that the spatiotemporal expression of _CpGPPS.SSU1_ and _CpGGPPS1_ is strictly confined to the scent-emitting tissues with a good association to monoterpene contents. Moreover, [PERSON] et al.