An Auxin Binding Protein is Localized in the Symbiosome Membrane of Soybean Nodules

Abstract

Binding of tritiated indole-3-acetic acid ([3H]IAA) to symbiosome membranes of soybean nodules occurred in a protein-dependent manner and was competitively inhibited by unlabeled indole-3-acetic acid (IAA), 1-naphthaleneacetic acid (1-NAA) and dithiothreitol (DTT), but not by tryptophan and benzoic acid. The symbiosome membranes bound IAA with a KD of 1 × 10-6 m. Photoaffinity labeling identified an auxin-binding protein (ABP) in the symbiosome membrane with an apparent molecular mass of 23 kDa. This 23 kDa protein was labeled either with 5-azido-[7-3H]indole-3-acetic acid ([3H]N3IAA) or with 5′-azido-[3,6-3H2]-1-naphthylphthalamic acid ([3H2]N3NPA). Labeling of the 23 kDa protein with [3H]N3IAA was competitively inhibited by unlabeled IAA and 1-NAA. NPA and quercetin, inhibitors of polar auxin transport, as well as rutin, a glycosylated derivative of quercetin, competed with IAA for binding. Conversely, [3H2]N3NPA labeling was inhibited by unlabeled IAA and NPA. The 23 kDa symbiosome membrane protein was partially solubilized with Triton X-100 and nearly completely using Triton X-114. The observation that auxin transport inhibitors compete with IAA for binding suggests that the symbiosome membrane ABP could be part of an auxin efflux carrier system required to control the auxin concentration in infected soybean nodule cells.