2,4-Dichlorophenoxyacetic Acid Inhibits the Outer Membrane NADH Dehydrogenase of Plant Mitochondria

Abstract

The NADH dehydrogenase of potato (Solanum tuberosum) and mung bean (Phaseolus aureus) outer mitochondrial membranes is specifically inhibited by both 2,4-dchlorophenoxyacetic and 2,4,5-trichlorophenoxyacetic acids but not by the natural auxin indole-3-acetic acid.The chemical agent 2,4-D is widely used as a selective herbicide although its biochemical mode of action is not fully understood. We have found that 2,4-D inhibits the NADH dehydrogenase system associated with the outer membranes of potato (Solanum tuberosum) and mung bean (Phaseolus aureus) mitochondria. [ Inhibition by 2,4-D is detectable only at initial NADH levels below this value (Fig. 2), indicating that 2,4-D acts at the dehydrogenase. The mode of inhibition of the NADH dehydrogenase by 2,4-D is complex, appearing competitive with NADH at concentrations of this substrate just below 0.2 mm and uncompetitive at very low NADH concentrations (Fig. 2 and ref. 3). At an initial NADH level of 0.1 mm, concentrations of 2,4-D approaching its water solubility limit (about 2 mM) produce essentially complete inhibition of this outer mitochondrial membrane activity, while inhibiting the microsomal NADH-Cyt c oxidoreductase to a far lesser extent (Fig. 1). It was found that the related compound 2,4,5-trichlorophenoxyacetic acid is also a specific inhibitor of the outer mitochondrial membrane NADH dehydrogenase while IAA, the natural auxin whose hormonal action appears to be mimicked by these chlorophenoxy acids, is not.Since the function of the NADH dehydrogenase-Cyt bs,% system of the outer mitochondrial membrane is not known, the physiological relevance of its inhibition by these compounds is unclear. A possibility suggested by these results is that this electron transfer system is involved, in plants, in

The outer membrane of plant mitochondria contains a calcium‐dependent protein kinase and multiple phosphoproteins

“…Dominant polypeptides of 30, 31, 33, 34, 47, 48 and 74 kDa are strongly enriched in the outer membrane fraction. The presence of strong bands at 30-35 and 45-50 kDa is similar to what has been reported for the outer membrane of Neurospora crassa [23,24], yeast [25] and mung bean cotyledons [8].…”

“…This protein kinase(s) is Ca2'-dependent since the phosphorylation of histones is abolished in the presence of EGTA (results not shown). The kinase is most likely located on the outer surface of the outer membrane since histones are too large to pass the outer membrane which is impermeable to molecules larger than 5-10 kDa [8]. Consistent with this conclusion, total incorporation is most enhanced by histones in the outer membrane fraction (Table II).…”

“…1) are also phospho~la~d (Fig. 3); they are probably porin since they are close to the molecular mass of the subunit of porin [8] and since they cross-react with antibodies raised against porin isolated from the outer membrane of mitochondria from Dictyostelium discoides [31]. Mannella [S] speculates that the pores in the outer membrane are open in State 4 (high ATP, low ADP) and closed in State 3 (low ATP, high ADP) due to changes in the mitochondrial structure bringing the two membranes in closer contact in State 3.…”

“…The only known enzyme marker is the antimycin A-insensitive NADH-cytochrome c reductase (CCR). Other markers for the outer membrane characteristic for mammalian mitochondria, such as monoamine oxidase and kynurenine hydroxylase, appear to be missing [7,8]. The outer membrane is normally considered to be freely permeable to all molecules smaller than 5-10 kDa, probably due to the presence of pores [9], and for that reason it is often ignored in studies on isolated mitochondria.…”