An integrated transcriptomics and metabolomics analysis of the Cucurbita pepo nectary implicates key modules of primary metabolism involved in nectar synthesis and secretion

Abstract

Nectar is the main reward that flowers offer to pollinators to entice repeated visitation. Cucurbita pepo (squash) is an excellent model for studying nectar biology, as it has large nectaries that produce large volumes of nectar relative to most other species. Squash is also monoecious, having both female and male flowers on the same plant, which allows comparative analyses of nectary function in one individual. Here, we report the nectary transcriptomes from both female and male nectaries at four stages of floral maturation. Analysis of these transcriptomes and subsequent confirmatory experiments revealed a metabolic progression in nectaries leading from starch synthesis to starch degradation and to sucrose biosynthesis. These results are consistent with previously published models of nectar secretion and also suggest how a sucrose‐rich nectar can be synthesized and secreted in the absence of active transport across the plasma membrane. Nontargeted metabolomic analyses of nectars also confidently identified 40 metabolites in both female and male nectars, with some displaying preferential accumulation in nectar of either male or female flowers. Cumulatively, this study identified gene targets for reverse genetics approaches to study nectary function, as well as previously unreported nectar metabolites that may function in plant‐biotic interactions.

Systems analyses of key metabolic modules of floral and extrafloral nectaries of cotton

“…Expression profiles identified via RNA-seq analysis were validated by quantitative real time PCR (qRT-PCR) analysis using RNA isolated from floral and bracteal nectaries. These validation genes were chosen based on their known or suspected functionality in nectary development (Kram et al, 2008; Lin et al, 2014; Ruhlmann et al, 2010; Solhaug et al, 2019). Some of the selected genes display distinctive differential expression during nectary development, while others show a more stable expression pattern (e.g., NiR1 ).…”

“…Because nectar production is heavily dependent on sugar metabolism (Ren et al, 2007; Solhaug et al, 2019) and sugar transport, the RNA-seq data were annotated with respect to starch and sucrose metabolic pathways and transmembrane transporters, using MapMan (Thimm et al, 2004) and gene ontology terms. The resulting gene list was further filtered, selecting those genes that are upregulated in the nectary transcriptomes relative to the adjoining control non-nectary transcriptomes.…”

“…These studies provide evidence to support an eccrine-based model of nectar synthesis and secretion, which utilizes pores and transporters for movement of pre-nectar metabolites through the plasma membrane of nectariferous parenchyma tissues [reviewed by (Roy et al, 2017)]. In this model, prior to nectar secretion the ‘pre-nectar’ sugar metabolites are delivered through the vasculature and stored in the nectary parenchyma, primarily as starch (Chatt et al, 2018; Lin et al, 2014; Peng et al, 2004; Ren et al, 2007; Solhaug et al, 2019). At the time of nectar secretion, the stored starch is rapidly degraded, and the products are used to synthesize sucrose through the enzymatic action of sucrose-phosphate synthases (SPS) and sucrose-phosphate phosphatases.…”

“…The molecular underpinnings of nectar synthesis and secretion are beginning to be elucidated through advancements in “omics” technologies primarily using the floral nectaries of Arabidopsis, Cucurbita pepo and Nicotiana spp. (Kram et al, 2009; Lin et al, 2014; Ren et al, 2007; Solhaug et al, 2019). These studies provide evidence to support an eccrine-based model of nectar synthesis and secretion, which utilizes pores and transporters for movement of pre-nectar metabolites through the plasma membrane of nectariferous parenchyma tissues [reviewed by (Roy et al, 2017)].…”

“…The sucrose is exported into the apoplasm in a concentration dependent manner via the uniporter SWEET9, and subsequently hydrolyzed by cell wall invertase (CWINV4), to the hexose components glucose and fructose, thereby maintaining the sucrose concentration gradient (Lin et al, 2014; Ruhlmann et al, 2010). The last step of sucrose hydrolysis is critical to the production of hexose-rich nectars (Ruhlmann et al, 2010), but may play a minimal role in production of sucrose-rich nectars (Chatt et al, 2018; Solhaug et al, 2019).…”