Auxin Oxidase and Growth Control in Tissue Cultures of Ephedra

Abstract

Tissue cultures derived from an unknown species of Ephedra require exogenous auxin in order to grow. However, indoleacteic acid (IAA) is ineffective, but 2,4-dichlorophenozyacetic acid (2,4-D) and alpha-naphthalene acetic acid (NAA) are. Investigation revealed that the tissue released an indoleacetic acid oxidase into the nutrient medium and also when simply soaked in water. The enzyme obtained in this manner requires Mn++ and 2,4-dichlorophenol (DCP) for activity. Homogenates of the tissue are inactive unless they have been dialyzed. Such preparations also require Mn++ and DCP for activity. The intact tissue destroys IAA within 14 hours in the absence of added cofactors (2g tissue destroyed 1400 [mu]g IAA). Concentrated enzyme preparations obtained either from dialyzed homogenates or "leakage" into water can also destroy IAA without cofactors, although Mn++ and DCP accelerate the activity of the enzyme. Chromatography of diethyl ether extracts of the tissues; the reaction medium in which tissue has been incubated with IAA; and reaction media containing no tissue but enzyme with IAA and cofactors revealed 8 Ehrlich-positive spots of which one was a trace of IAA. This indicates that the disappearance of IAA from solutions incubated with tissue is not due to uptake, but rather to destruction of the IAA. The evidence indicates that auxin oxidase, in these tissue cultures, at least, functions in vivo to destroy exogenous IAA. This explains why the tissue cannot grow on media in which the sole auxin supplied is IAA.