Developing Heat- and Drought-Tolerant Canola by Regulating Oil-Associated Enzyme: Phase Two

Term: 3 years
Status: Ongoing
Researcher(s): Gavin Chen, Habibur Rahman, U of A; Bianyun Yu, Irina Zaharia, NRC- Canada; Yang Xu, U of Guelph
SaskOilseeds Investment: $58,263
Total Project Cost: $450,281
Funding Partners: WGRF, ACPC, MCGA

Objective

In this phase-two project, we plan to (1) systemically test the canola lines developed in CARP 2022.17 under heat and drought stresses at the above-mentioned three developing stages, (2) generate canola lines with PDAT upregulation driven by heat-, drought-, and heat & drought-inducible promoters, (3) evaluate their performance to identify ideal canola lines with good performance under both normal condition and stress, and (4) assess genetically diverse canola lines to identify those with better heat and drought tolerance and to discover naturally occurring PDAT genes within the Brassica gene pool, which contribute to stress tolerance.

Project Description

Canola is the most important oilseed crop in Canada, but it is vulnerable to heat and drought stresses. These stresses may cause abnormal vegetative growth, flower abortion, abnormal siliques, and substantially reduced seed yield and quality in canola and other crops.

Based on climate projections for 2100, the average temperature could increase by 3.7°C, and more than 50% of field crops would suffer yield and quality decreases if no mitigation procedures are taken. Moreover, drought continues to be a major concern across western Canada in the past several years, which can affect canola from seedling to seed maturing stages. In addition, sometimes heat and drought may happen simultaneously, especially at the reproductive stage. Therefore, it is important to develop canola varieties with increased tolerance to heat and drought stress, without the penalty of seed yield and quality.

In terms of seed quality, oil (mainly triacylglycerol) is the most valuable component of canola seeds. Triacylglycerol biosynthesis in canola seeds takes place mainly during seed development in the embryo and involves various enzymes and functional proteins operating within the plastid and endoplasmic reticulum (ER). In brief, fatty acids are first synthesized in the plastid and then esterified to form acyl-CoAs outside the plastid. In the ER, triacylglycerol assembly follows the acyl-CoA-dependent pathway, where acyl-CoAs are sequentially added to glycerol-3-phosphate to form triacylglycerol. Triacylglycerol can also be produced via the acyl-CoA-independent pathway, where acyl chains are transferred from phospholipids to triacylglycerol, catalyzed by phospholipid:diacylglycerol acyltransferase (PDAT).

Recent studies have revealed that some triacylglycerol-associated genes play important roles in abiotic stress tolerance in plants. Regulating genes in triacylglycerol biosynthesis, such as PDAT, may thus represent a novel strategy for breeding oilseed crops with enhanced heat and drought tolerance and better seed quality. For example, in the model plant Arabidopsis, PDAT1 has a critical role in rapid plastid lipid metabolism driving TAG accumulation during heat stress, which in turn effectively increases the plant’s heat and cold resistance. Since seed development and oil biosynthesis in Arabidopsis and canola share very similar pathways, it would be of great value to determine whether canola PDAT1 overexpression can similarly increase canola’s heat and drought tolerance and maintain or even increase seed yield and quality.

Supported by the CARP project, CARP 2022.17, we have generated canola lines with the upregulation of canola PDAT1 driven by a constitutive 35S promoter (35SBnPDAT1OE), and evaluated the performance of the canola lines under heat and drought stress over the past two years. We are generating canola lines with BnPDAT1 downregulation driven by the constitutive 35S promoter (35SBnPDAT1KD). The proof-of-concept results indicated that the homozygous BnPDAT1OE canola lines performed significantly better under heat or the combination of heat and drought stresses at the flowering and seed developing stage. This was evidenced by the production of more siliques per plant (p<0.05), seeds per silique (p<0.05), yield per plant (p<0.05), and comparable oil content, indicating its promise for canola breeding. Moreover, we also preliminarily tested their performance at the seedling stage, and the results demonstrated that these canola lines exhibited higher survival rate under drought stress (66% with BnPDAT1 OE vs 47% in wild type), indicating the possible functions of PDAT in dealing with drought stress at seedling stage.

To fully evaluate the potential of using BnPDAT1 and the associated canola lines in breeding canola with heat and drought tolerance, it is necessary to comprehensively test the homozygous 35SBnPDAT1OE and 35SBnPDAT1KD canola lines at the seedling, vegetative and flowering/seed development stages under heat, drought and the combination of heat and drought stresses. Moreover, although constitutive promoters are frequently used in proof-of-concept studies, they may possibly have negative effects on crops when growing without severe stresses in some cases. While no obvious agronomic defects were observed in our Phase-One research, it is worthy to test the use of stress-inducible promoters to finely regulate PDAT for generating canola lines with good performance in both normal and non-ideal years. Meanwhile, based on the Phase One research results, it would be valuable to evaluate the PDAT gene expression in a genetically diverse canola population to identify those with improved heat and drought tolerance. The University of Alberta Canola Program has developed thousands of genetically diverse canola lines carrying genome contents of different Brassica gene pools, such as different variants of B. oleracea, B. rapa, rutabaga, and winter canola. This unique canola germplasm offers a promising genetic resource for discovering the naturally occurring genes in the Brassica gene pool that confer stress tolerance. These genes can be readily used in canola breeding to enhance abiotic stress tolerance.