FAD2 and FAD3 Desaturases Form Hetero-dimers That Facilitate Metabolic Channeling in vivo [Lipids]

May 8th, 2014 by Lou, Y., Schwender, J., Shanklin, J.

Plant desaturases comprise two independently evolved classes, a structurally well-characterized soluble class responsible for the production of monoenes in the plastids of higher plants and the poorly structurally characterized integral membrane class that has members in the plastid and endoplasmic reticulum that are responsible for producing mono- and polyunsaturated fatty acids. Both require Fe and oxygen for activity and are inhibited by azide and cyanide underscoring their common chemical imperatives. We previously showed that the Δ9 acyl-CoA integral membrane desaturase Ole1p from Saccharomyces cereviciae, exhibits dimeric organization, like the soluble plastidial acyl-ACP desaturases. Here we use two independent bimolecular complementation assays i.e., yeast two-hybrid analysis and Arabidopsis leaf protoplast split-luciferase assay to demonstrate that members of the plant integral membrane fatty acid desaturase (FAD) family, FAD2, FAD3, FAD6, FAD7 and FAD8 self-associate. Further, the endoplasmic reticulum-localized desaturase FAD2 can associate with FAD3 as can the plastid-localized FAD6 desaturase with either FAD7 or FAD8. These pairings appear to be specific because pairs, such as FAD3 and FAD7 (or FAD8), and FAD2 and FAD6 do not interact despite their high amino acid similarity. These results are consistent also with their known endoplasmic reticulum and plastid subcellular localizations. Chemical cross-linking experiments confirm FAD2 and FAD3, can form dimers, like the yeast Ole1p, and when co-expressed, can form FAD2-FAD3 heterodimers. Metabolic flux analysis of yeast co-expressing FAD2 and FAD3 indicates that heterodimers can form a metabolic channel in which 18:1-PC is converted to 18:3-PC without releasing a free 18:2-PC intermediate.