A mutant cotton fatty acid desaturase 2-1d allele causes protein mistargeting and altered seed oil composition

Authors: Shockey, Jay; Gilbert, Matthew; Thyssen, Gregory

Submitted to: BMC Plant Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/9/2023
Publication Date: 3/17/2023
Citation: Shockey, J.M., Gilbert, M.K., Thyssen, G.N. 2023. A mutant cotton fatty acid desaturase 2-1d allele causes protein mistargeting and altered seed oil composition. BMC Plant Biology. 23. Article 147. https://doi.org/10.1186/s12870-023-04160-8.
DOI: https://doi.org/10.1186/s12870-023-04160-8

Interpretive Summary: In this study we sought to gain a better understanding of the cause of the high oleic acid trait in a small group of pima cotton varieties that we found previously. A gene that acts a gatekeeper to production of polyunsaturated fatty acids from monounsaturates was affected. And rather than a relatively small DNA insertion as first thought, we found that the mutation was caused by movement of a poorly understood self-copying DNA element called a retrotransposon. The mutation alters the protein coding sequence of the gatekeeper enzyme, called a desaturase, leading to changes in the protein sequence and biochemical properties of this enzyme. Most or all of the negative change come from the loss of a section of the protein deleted by the transposon mutation, thus confirming the importance of maintaining strict accuracy of DNA sequence throughout the entire gene. These results also reaffirm the importance of the gatekeeper desaturases as determinants of seed oil fatty acid composition, including cottonseed oil.

Technical Abstract: Cotton (Gossypium sp.) has been cultivated for centuries for its spinnable fibers, but its seed oil also possesses untapped economic potential if, improvements could be made to it oleic acid content. Previous studies, including those from our laboratory, identified pima accessions containing approximately doubled levels of seed oil oleic acid, compared to standard upland cottonseed oil. Here, the molecular properties of a fatty acid desaturase encoded by a mutant allele identified by genome sequencing in an earlier analysis were analyzed. The mutant sequence is predicted to encode a C-terminally truncated protein lacking nine residues, including a predicted endoplasmic reticulum membrane retrieval motif. We determined that the mutation was caused by a relatively recent movement of a Ty1/copia type retrotransposon that is not found associated with this desaturase gene in other sequenced cotton genomes. The mutant desaturase, along with its repaired isozyme and the wild-type A-subgenome homoeologous protein were expressed in transgenic yeast and stably transformed Arabidopsis plants. All full-length enzymes had high levels of activity, and efficiently converted oleic acid to linoleic acid. The mutant desaturase protein displayed only trace amounts of activity and only when strongly overexpressed in yeast cells, indicating that the missing C-terminal amino acid residues are not strictly required for enzyme activity, yet are necessary for proper subcellular targeting to the endoplasmic reticulum membrane. These results provide the biochemical underpinning that links a genetic lesion present in a limited group of South American pima cotton accessions and their rare seed oil oleic acid traits. Markers developed to the mutant desaturase allele are currently being used in breeding programs designed to introduce this trait into agronomic upland cotton varieties.