Effects of Inhibitors of Protein Synthesis on Transmembrane Auxin Transport in Cucurbita pepo L. Hypocotyl Segments

Pretreatment of 2?0 mm segments of etiolated zucchini (Cucurbitapepo L.) hypocotyl with cycloheximide (CH) or 2-(4-methyl-2,6-dinitroanilino)-N-methylpropionamide(MDMP) eliminated the stimulation by N-1-naphthylphthalamicacid (NPA) of net uptake of 1-¹⁴C]indol-3yl-acetic acid (1-¹⁴C]IAA),but had relatively little effect on the net uptake of IAA inthe absence of NPA. The efflux of 1-¹⁴C]IAA from preloadedsegments was not substantially affected by inhibitor pretreatmentin the absence of NPA, but CH pretreatment significantly inhibitedthe reduction of efflux caused by NPA. Pretreatment with CHor MDMP did not affect net uptake by segments of the pH probe2-¹⁴C]5,5-dimethyl-oxazolidine-2,4-dione (2-¹⁴C]DMO), or thenet uptake of ¹⁴C]-labelled 3-O-methylglucose (¹⁴C]3-0-MeGlu),suggesting that neither inhibitor affected intracellular pHor the general function of proton symporters in the plasma membrane.Both compounds reduced the incorporation of label from ³⁵S]methionineinto trichloroacetic acid (TCA)-insoluble fractions of zucchinitissue, confirming their inhibitory effect on protein synthesis. The steady-state association of ³H]IAA with microsomal vesiclesprepared from zucchini hypocotyl tissue was enhanced by theinclusion of NPA in the uptake medium. The stimulation by NPAof ³H]IAA association with microsomes was substantially reducedwhen the tissue was pretreated with CH. However, CH pretreatmentdid not affect the level of high affinity NPA binding to themembranes indicating that treatments did not result in lossof NPA receptors. It is suggested that the auxin transport site on the effluxcarrier system and the receptor site for NPA may reside on separateproteins linked by a third, rapidly turned-over, transducingprotein. Key words: Auxin carriers, auxin efflux, Cucurbita pepo, phytotropin receptors