Regulation of auxin transport by aminopeptidases and endogenous flavonoids

The 1-N-naphthylphthalamic acid (NPA)-binding protein is a putative negativ e regulator of polar auxin transport that has been shown to block auxin eff lux from both whole plant tissues and microsomal membrane vesicles. We prev iously showed that NPA is hydrolyzed by plasma-membrane amidohydrolases tha t co-localize with tyrosine, proline, and tryptophan-specific aminopeptidas es (APs) in the cotyledonary node, hypocotyl-root transition zone and root distal elongation zone of Arabidopsis thaliana (L.) Heynh. seedlings. Moreo ver, amino acyl-beta-naphthylamide (aa-NA) conjugates resembling NPA in str ucture have NPA-like inhibitory activity on growth, suggesting a possible r ole of APs in NPA action. Here we report that the same aa-NA conjugates and the AP inhibitor bestatin also block auxin efflux from seedling tissue. Be statin and, to a lesser extent, some aa-NA conjugates were more effective i nhibitors of low-affinity specific [H-3]NPA-binding than were the flavonoid s quercetin and kaempferol but had no effect on high-affinity binding. Sinc e the APs are inhibited by flavonoids, we compared the localization of endo genous flavonoids and APs in seedling tissue. A correlation between AP and flavonoid localization was found in 5- to 6-d-old seedlings. Evidence that these flavonoids regulate auxin accumulation in vivo was obtained using the flavonoid-deficient mutant, tt4. In whole-seedling [C-14]indole-3-acetic a cid transport studies, the pattern of auxin distribution in the tt4 mutant was shown to be altered. The defect appeared to be in auxin accumulation, a s a considerable amount of auxin escaped from the roots. Treatment of the t t4 mutant with the missing intermediate naringenin restored normal auxin di stribution and accumulation by the root. These results implicate APs and en dogenous flavonoids in the regulation of auxin efflux.