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Analyses of diverse low alkaloid tobacco germplasm identify naturally occurring nucleotide variability contributing to reduced leaf nicotine accumulation.

ABSTRACT: Recent suggestions for mandated lowering of nicotine content in cigarettes have prompted tobacco breeders to search for N. tabacum germplasm with allelic variability contributing to low alkaloid accumulation. In this research, we phenotyped a series of 81 selected diverse tobacco introductions (TIs) to identify a sub-group with authentic low alkaloid phenotypes. We also genotyped these materials for sequences associated with the Nic1 and Nic2 loci previously reported to influence tobacco alkaloid biosynthesis. Only five low alkaloid TIs possessed previously described deletions of Ethylene Response Factor (ERF) genes at the Nic2 locus that contribute to lower alkaloid accumulation. Eleven TIs possessed an apparent deletion of ERF199, a gene recently reported to underlie the effect at the Nic1 locus. Quantitative trait locus (QTL) mapping was performed using populations derived from three selected low alkaloid TIs to possibly identify new genomic regions affecting alkaloid accumulation. A major QTL was identified on linkage group 7 in all three populations that aligned with the Nic1 locus. A newly discovered 5 bp deletion in the gene MYC2a on linkage group 5 was found to likely partially underlie the ultra-low alkaloid phenotype of TI 313. This new information is useful for tobacco breeders attempting to assemble novel genetic combinations with the potential for meeting future levels of tolerance for nicotine concentration in cigarette tobacco.

Supplementary information

The online version contains supplementary material available at 10.1007/s11032-021-01274-5.

SUBMITTER: Burner N

PROVIDER: S-EPMC10248598 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

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Analyses of diverse low alkaloid tobacco germplasm identify naturally occurring nucleotide variability contributing to reduced leaf nicotine accumulation.

Burner Nathaniel N McCauley Abigail A Pramod Sreepriya S Frederick Jesse J Steede Tyler T Kernodle Sheri P SP Lewis Ramsey S RS

Molecular breeding : new strategies in plant improvement 20220111 1

Recent suggestions for mandated lowering of nicotine content in cigarettes have prompted tobacco breeders to search for N. tabacum germplasm with allelic variability contributing to low alkaloid accumulation. In this research, we phenotyped a series of 81 selected diverse tobacco introductions (TIs) to identify a sub-group with authentic low alkaloid phenotypes. We also genotyped these materials for sequences associated with the Nic1 and Nic2 loci previously reported to infl ...[more]

PMID: 37309485

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Project description:Traditional breeding and molecular approaches have been used to develop tobacco varieties with reduced nicotine and secondary alkaloid levels. However, available low-alkaloid tobacco varieties have impaired leaf quality likely due to the metabolic consequences of nicotine biosynthesis downregulation. Recently, we found evidence that the unbalanced crosstalk between nicotine and polyamine pathways is involved in impaired leaf ripening of a low-alkaloid (LA) Burley 21 line having a mutation at the Nic1 and Nic2 loci, key biosynthetic regulators of nicotine biosynthesis. Since the Nic1 and Nic2 loci are comprised of several genes, all phenotypic changes seen in LA Burley 21 could be due to a mixture of genetics-based responses. Here, we investigated the commercial burley variety TN90 LC and its transgenic versions with only one downregulated gene, either putrescine methyl transferase (PMT-RNAi) or PR50-protein (PR50-RNAi). Nicotine levels of cured lamina of TN90 LC, TN90 PMT-RNAi and TN90 PR50-RNAi, were 70.5 ± 3.8, 2.4 ± 0.5, and 6.0 ± 1.1 mg/g dry weight, respectively. Low-alkaloid transgenic lines showed delayed leaf maturation and impaired leaf quality. We analyzed polyamine contents and ripening markers in wild-type TN90 control plants (WT) and the two transgenic lines. The ripening markers revealed that the PMT-RNAi line showed the most pronounced impaired leaf maturation phenotype at harvest, characterized by higher chlorophyll (19%) and glucose (173%) contents and more leaf mesophyll cells per area (25%), while the ripening markers revealed that maturation of PR50-RNAi plants was intermediate between PMT-RNAi and WT lines. Comparative polyamine analyses showed an increase in free and conjugated polyamines in roots of both transgenic lines, this being most pronounced in the PMT-RNAi plants. For PMT-RNAi plants, there were further perturbations of polyamine content in the leaves, which mirrored the general phenotype, as PR50-RNAi transgenic plants looked more similar to the WT than PMT-RNAi transgenic plants. Activity of ornithine decarboxylase, the enzyme that catalyzes the committing step of polyamine biosynthesis, was significantly higher in roots and mature leaves of PMT-RNAi plants in comparison to WT, while there was no increase observed for arginine decarboxylase. Treatment of both transgenic lines with polyamine biosynthesis inhibitors decreased the polyamine content and ameliorated the phenotype, confirming the intricate interplay of polyamine and nicotine biosynthesis in tobacco and the influence of this interplay on leaf ripening.

Dataset Information

Analyses of diverse low alkaloid tobacco germplasm identify naturally occurring nucleotide variability contributing to reduced leaf nicotine accumulation.

Supplementary information

Publications

Analyses of diverse low alkaloid tobacco germplasm identify naturally occurring nucleotide variability contributing to reduced leaf nicotine accumulation.

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