Delineating genetic regulation of cannabinoid biosynthesis during female flower development in Cannabis sativa

Abstract

Cannabinoids are predominantly produced in the glandular trichomes on cannabis female flowers. There is little known on how cannabinoid biosynthesis is regulated during female flower development. We aim to understand the rate-limiting step(s) in the cannabinoid biosynthetic pathway. We investigated the transcript levels of cannabinoid biosynthetic genes together with cannabinoid contents during 7 weeks of female flower development. We demonstrated that the enzymatic steps for producing cannabigerol (CBG), which involve genes GPPS, PT, TKS, and OAC, could rate limit cannabinoid biosynthesis. Our findings further suggest that upregulation of cannabinoid synthases, CBDAS and THCAS in a commercial hemp and medical marijuana variety, respectively, is not critical for cannabinoid biosynthesis. The cannabinoid biosynthetic genes are generally upregulated during flower maturation; increased expression occurs coincident with glandular trichome development and cannabinoid production in the maturing flower. The results also suggest that different cannabis varieties may experience discrete transcriptional regulation of cannabinoid biosynthetic genes. In addition, we showed that methyl jasmonate (MeJA) can potentially increase cannabinoid production. We propose that biweekly applications of 100 μM MeJA starting from flower initiation would be efficacious for promoting cannabinoid biosynthesis. Our findings provide important genetic information for cannabis breeding to generate new varieties with favorable traits.

Analysis of hormonal regulation of promoter activities ofCannabis sativaprenyltransferase 1 and 4 and salicylic acid mediated regulation of cannabinoid biosynthesis

“…MeJA is a major hormone involved in plant defense responses to stress stimuli (Cheong and Choi 2003;Truman et al 2007;Wasternack and Hause 2013). Our previous study also showed that MeJA foliar spray increased cannabinoid production (Apicella et al 2022).…”

“…ABA plays essential roles in the regulation of floral development and plant responses to abiotic stress (Martignago et al 2020;Hsu et al 2021). Our previous study showed that both CsPT1 and CsPT4 were upregulated during female flower maturation and associated with glandular trichome development (Apicella et al 2022). Previous studies demonstrated that ABA alone did not affect flower sex determination, however, ABA inhibited IAA induced female flower formation and impaired GA induced male flower development (Ram and Jaiswal 1972;Galoch 1978).…”

“…Cannabinoids are known to be toxic to cells and must be exported to the secretory cavity (Sirikantaramas et al 2005;Morimoto et al 2007), where they may act as a defense mechanism against herbivory and in some cases, fungi and bacteria (Tanney et al 2021). Our previous work showed that MeJA can increase cannabinoid production (Apicella et al 2022). However, no JA responsive element was identified in the CsPT genes (Table 1).…”

“…Very few studies have been published regarding how hormone signaling pathways affect cannabis plants. The published research takes time to emphasize the important role of hormones in cannabinoid production (Mansouri et al 2009;Mansouri et al 2013;Jalali et al 2019;Apicella et al 2022). For example, in (Apicella et al 2022), it is shown that jasmonic acid signaling pathways can increase cannabinoid content in flowering plants.…”

“…However, conclusions from more recent studies have determined that the correlation between THCAS gene expression and THCA content is poor, and attention should be paid to other enzymes in the cannabinoid biosynthetic pathway (Cascini et al, 2013(Cascini et al, , 2019. In recent work from this lab, this point was clearly established and further, the production of CBGA was identified as rate-limiting end point cannabinoid synthesis (Apicella et al 2022).…”