FAX1, a Novel Membrane Protein Mediating Plastid Fatty Acid Export

Abstract

Fatty acid synthesis in plants occurs in plastids, and thus, export for subsequent acyl editing and lipid assembly in the cytosol and endoplasmatic reticulum is required. Yet, the transport mechanism for plastid fatty acids still remains enigmatic. We isolated FAX1 ( fatty acid export 1), a novel protein, which inserts into the chloroplast inner envelope by α-helical membrane-spanning domains. Detailed phenotypic and ultrastructural analyses of FAX1 mutants in Arabidopsis thaliana showed that FAX1 function is crucial for biomass production, male fertility and synthesis of fatty acid-derived compounds such as lipids, ketone waxes, or pollen cell wall material. Determination of lipid, fatty acid, and wax contents by mass spectrometry revealed that endoplasmatic reticulum (ER)-derived lipids decreased when FAX1 was missing, but levels of several plastid-produced species increased. FAX1 over-expressing lines showed the opposite behavior, including a pronounced increase of triacyglycerol oils in flowers and leaves. Furthermore, the cuticular layer of stems from fax1 knockout lines was specifically reduced in C29 ketone wax compounds. Differential gene expression in FAX1 mutants as determined by DNA microarray analysis confirmed phenotypes and metabolic imbalances. Since in yeast FAX1 could complement for fatty acid transport, we concluded that FAX1 mediates fatty acid export from plastids. In vertebrates, FAX1 relatives are structurally related, mitochondrial membrane proteins of so-far unknown function. Therefore, this protein family might represent a powerful tool not only to increase lipid/biofuel production in plants but also to explore novel transport systems involved in vertebrate fatty acid and lipid metabolism.

Abstract

The novel protein FAX1 mediates the export of free fatty acids across the inner membrane of chloroplasts so that they can be processed in other plant cell organelles to generate oils, waxes, and other lipids.

Author Summary

Fatty acid synthesis in plants occurs in chloroplasts—the organelle more commonly known for conducting photosynthesis. For subsequent lipid assembly to be possible in the endoplasmatic reticulum (ER), export of these fatty acids across the chloroplast envelope membranes is required. The mechanism of this transport until now has not been known. We isolated FAX1 ( fatty acid export 1), a novel membrane protein in chloroplast inner envelopes. FAX1 function is crucial for biomass production, male fertility, and the synthesis of fatty acid-derived compounds like lipids, waxes, or cell wall material of pollen grains. Whereas ER-derived lipids decreased when FAX1 was missing, levels of plastid-produced lipids increased. FAX1 over-expressing mutants showed the opposite behavior, including an increase of triacyglycerol oils. Because FAX1 could complement for fatty acid transport in yeast, we concluded that FAX1 mediates the export of free fatty acids from chloroplasts. In vertebrates, FAX1 relatives are structurally related proteins of so-far unknown function in mitochondria. This protein family may thus represent a powerful tool not only to increase lipid oil and biofuel production in plants but also to explore novel transport systems in animals.