Dataset Information

Sex-dependent variation of pumpkin (Cucurbita maxima cv Big Max) nectar and nectaries as determined by proteomics and metabolomics

ABSTRACT: Cucurbita maxima c.v. Big Max is a monoecious plant with separate male and female flowers which utilizes nectar as a floral reward for the attraction of bees to the flowers. Maintenance of this mutualism with bees is essential for the reproductive success in this economically valuable crop. In this study, the sex-dependent variation in composition of male and female nectar and the nectaries were defined using a combination of GC-MS based metabolomics and LC-MS/MS based proteomics. Nectar proteins were detected non-quantitatively using LC-MS/MS coupled to Orbitrap. The male and female nectary proteomes were quantified using iTRAQ labeled peptides which were fractioned and concatenated prior to detection using LC-MS/MS coupled to Orbitrap. The nectar proteome consists of 46 proteins, of which 70% overlap between nectar types. Only two proteins are unique to female nectar, and 10 are specific to male nectar. Nectarins (nectar proteins) commonly alter nectar carbohydrates. In our datasets, significant differences in carbohydrate abundance, specifically galactose and sucrose, may be explained by the unique presence of galactinol--sucrose galactosyltransferase 2 (functioning in galactose metabolism) and invertase (catalyzes the hydrolysis of sucrose to glucose and fructose) in the nectar of female and male flowers respectively. The nectary proteome contained 339 identifiable proteins, 71% of which were descriptively annotatable by homology to Plantae. The abundance of 45 proteins differs significantly between male and female nectaries. GO slim analysis revealed a high abundance of proteins related to transport, protein metabolism, carbohydrate metabolism, response to stress, and amino acid metabolism. Comparisons of relative protein abundance revealed that 45 proteins displayed differential expression between male and female nectaries (p-value <0.05); 20 of these proteins were more abundant in male nectaries and 25 were more abundant in female nectaries. GO enrichment of nectary proteins with increased female abundances indicate that female-enriched GO terms are associated with proteins functioning as plasma membrane proton pumps and central metabolism. Proteins associated with male nectary-enriched GO terms were related to cinnamic acid biosynthesis and neutralization of superoxide radicals and hydrogen peroxide.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Cucurbita Maxima

TISSUE(S): Nectar, Nectary

SUBMITTER: Elizabeth Chatt

LAB HEAD: Dr. Basil J. Nikolau

PROVIDER: PXD009810 | Pride | 2018-07-03

REPOSITORIES: Pride

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Publications

Sex-Dependent Variation of Pumpkin (<i>Cucurbita maxima</i> cv. Big Max) Nectar and Nectaries as Determined by Proteomics and Metabolomics.

Chatt Elizabeth C EC von Aderkas Patrick P Carter Clay J CJ Smith Derek D Elliott Monica M Nikolau Basil J BJ

Frontiers in plant science 20180629

Nectar is a floral reward that sustains mutualisms with pollinators, which in turn, improves fruit set. While it is known that nectar is a chemically complex solution, extensive identification and quantification of this complexity has been lacking. <i>Cucurbita maxima</i> cv. Big Max, like many cucurbits, is monoecious with separate male and female flowers. Attraction of bees to the flowers through the reward of nectar is essential for reproductive success in this economically valuable crop. In ...[more]

PMID: 30008725

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Project description:Floral nectar is a rich secretion produced by nectaries offered as an attractive reward to pollinators. Floral nectaries secrete an aqueous solution largely composed of sugars and amino acids as well as an array of proteins called nectarins. These provide one of the main mechanisms of floral defense in Nicotiana nectar. Due to its composition, floral nectar provides an ideal source of nutrients for specialized microorganisms that can change the nectar composition. Understanding of floral defenses and the role of nectar proteins is essential to predict the impacts in microbial growth, nectar composition, and relationships between plants and pollinators. In order to deepen knowledge about Nicotiana spp. nectar proteins, we used LC-MS/MS-based comparative proteomic analysis to identify 22 proteins from Petunia hybrida, 35 proteins from Datura stramonium, and 144 proteins from 23 different species of Nicotiana. GO enrichment analysis and secretory signal peptide prediction demonstrated that defense/stress was the largest group of proteins in nectars with differential abundance and distribution throughout genus Nicotiana. The Nicotiana spp. proteome consisted of 105 exclusive proteins such as lipid transfer proteins (LTPs), Nectar Redox Cycle proteins, proteases inhibitors (PI), and PR-proteins. Analysis by taxonomic sections demonstrated that LTPs were the most abundant group of proteins in the nectar of Undulatae and Noctiflora sections while nectarins were more abundant proteins in Rusticae, Suaveolens, Polydicliae, and Alata sections. Peroxidases (Pox) and chitinases (Chit) were exclusive to P. hybrida nectar, while D. stramonium proteome had only seven unique proteins. Significant differences were identified between the proteome of taxonomic sections providing relevant insights into the group of proteins related to defense/stress associated with Nectar Redox Cycle, antimicrobial proteins and signaling pathways. The activity of floral nectar proteins is suggested impact the microbial growth. The knowledge about these proteomes provides significant insights into the diversity of proteins secreted in the nectars and the array of mechanisms used by Nicotiana spp. in its floral defense.

Project description:Many flowering plants attract pollinators by offering a reward of floral nectar. Remarkably, the molecular events involved in the development of nectaries, the organs that produce nectar, as well as the synthesis and secretion of nectar itself, are poorly understood. Indeed, to date, no genes have been shown to directly affect the de novo production or quality of floral nectar. To address this gap in knowledge, the ATH1 Affymetrix GeneChip array was used to systematically investigate the Arabidopsis nectary transcriptome to identify genes and pathways potentially involved in nectar production. In this study, we identified a large number of genes differentially expressed between secretory lateral nectaries and non-secretory median nectary tissues, as well as between mature lateral nectaries (post-anthessis) and immature lateral nectary tissue (pre-anthesis). To identify genes that are specifically upregulated in nectary tissues, and therefore may contribute to nectar production, we compared individual nectary samples (ILN, MLN & MMN) with 51 non-nectary reference tissues (downloaded .cel files from other studies). 270 genes were identified as being significantly upregulated in nectaries. The expression patterns for multiple genes were also confirmed via RT-PCR. The co-normalized RMA expression data (i.e., our own 8 nectary samples and 51 samples downloaded from GEO) are available as a supplementary file at the foot of this record. Three different types of RNA samples were prepared from Arabidopsis thaliana ecotype Columbia-0 nectaries: mature lateral nectaries (MLN; Stage 14-15 flowers), immature lateral nectaries (ILN, Stage 11-12 flowers), and mature median nectaries (MMN, Stage 14-15 flowers) [developmental stages defined by (Smyth et al., 1990)]. MLN are secretory tissues, whereas, ILN and MMN are pre-secretory and nonsecretory tissues, respectively. All nectary tissues were separately dissected by hand from the flowers of primary inflorescences of ca. 30-35 day-old plants. All plants were grown in soil with a 16h light/8h dark light regimen. Due to the small size of nectaries, dissections took place over several days from 4-8 hours after dawn (h.a.d.). The gene expression dataset obtained by our laboratory was enriched with third party expression data (i.e., 51 GEO Samples: GSM131510..GSM131660). The final gene expression dataset includes 59 hybridizations.

Dataset Information

Sex-dependent variation of pumpkin (Cucurbita maxima cv Big Max) nectar and nectaries as determined by proteomics and metabolomics

Publications

Sex-Dependent Variation of Pumpkin (<i>Cucurbita maxima</i> cv. Big Max) Nectar and Nectaries as Determined by Proteomics and Metabolomics.

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