Discovery and Characterization of the 3-Hydroxyacyl-ACP Dehydratase Component of the Plant Mitochondrial Fatty Acid Synthase System

Plant Physiol. 2017 Apr;173(4):2010-2028. doi: 10.1104/pp.16.01732. Epub 2017 Feb 15.

Abstract

We report the characterization of the Arabidopsis (Arabidopsis thaliana) 3-hydroxyacyl-acyl carrier protein dehydratase (mtHD) component of the mitochondrial fatty acid synthase (mtFAS) system, encoded by AT5G60335. The mitochondrial localization and catalytic capability of mtHD were demonstrated with a green fluorescent protein transgenesis experiment and by in vivo complementation and in vitro enzymatic assays. RNA interference (RNAi) knockdown lines with reduced mtHD expression exhibit traits typically associated with mtFAS mutants, namely a miniaturized morphological appearance, reduced lipoylation of lipoylated proteins, and altered metabolomes consistent with the reduced catalytic activity of lipoylated enzymes. These alterations are reversed when mthd-rnai mutant plants are grown in a 1% CO2 atmosphere, indicating the link between mtFAS and photorespiratory deficiency due to the reduced lipoylation of glycine decarboxylase. In vivo biochemical feeding experiments illustrate that sucrose and glycolate are the metabolic modulators that mediate the alterations in morphology and lipid accumulation. In addition, both mthd-rnai and mtkas mutants exhibit reduced accumulation of 3-hydroxytetradecanoic acid (i.e. a hallmark of lipid A-like molecules) and abnormal chloroplastic starch granules; these changes are not reversible by the 1% CO2 atmosphere, demonstrating two novel mtFAS functions that are independent of photorespiration. Finally, RNA sequencing analysis revealed that mthd-rnai and mtkas mutants are nearly equivalent to each other in altering the transcriptome, and these analyses further identified genes whose expression is affected by a functional mtFAS system but independent of photorespiratory deficiency. These data demonstrate the nonredundant nature of the mtFAS system, which contributes unique lipid components needed to support plant cell structure and metabolism.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Arabidopsis / enzymology*
  • Arabidopsis / genetics
  • Arabidopsis Proteins / genetics
  • Arabidopsis Proteins / metabolism*
  • Blotting, Western
  • Carbon Dioxide / metabolism
  • Fatty Acid Synthase, Type II / genetics
  • Fatty Acid Synthase, Type II / metabolism*
  • Fatty Acid Synthases / genetics
  • Fatty Acid Synthases / metabolism*
  • Gene Expression Regulation, Plant
  • Glycolates / metabolism
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Hydro-Lyases / genetics
  • Hydro-Lyases / metabolism*
  • Metabolomics / methods
  • Microscopy, Confocal
  • Microscopy, Electron, Transmission
  • Mitochondria / enzymology*
  • Mitochondria / ultrastructure
  • Mutation
  • Myristic Acids / metabolism
  • Plants, Genetically Modified
  • RNA Interference
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Analysis, RNA / methods
  • Sequence Homology, Amino Acid
  • Sucrose / metabolism

Substances

  • Arabidopsis Proteins
  • Glycolates
  • Myristic Acids
  • glycolic acid
  • Carbon Dioxide
  • Green Fluorescent Proteins
  • beta-hydroxymyristic acid
  • Sucrose
  • Fatty Acid Synthases
  • Hydro-Lyases
  • 3-hydroxyacyl-(acyl-carrier-protein) dehydratase
  • Fatty Acid Synthase, Type II