Phosphorylation of the H ATPase is induced by auxin without involvement of the transcriptional system that contains the ubiquitin ligase complex SCFTIR1 AFB. It should be noted, however, that PEO IAA and MG132 slightly suppressed IAA induced hypocotyl elongation . Calyculin A and Okadaic Acid Inhibit Auxin Induced H ATPase Phosphorylation Because calyculin A and okadaic acid sensitive protein phosphatases are most likely involved in the blue light induced activation of H ATPase in stomatal guard cells , we examined the effects of CA and OA, inhibitors of type 1 2A protein phosphatases, to determine the signaling pathway for auxin induced H ATPase phosphorylation. Interestingly, CA and OA completely inhibited the auxin induced phosphorylation of the H ATPase without affecting the quantity of the enzyme . Moreover, CA and OA inhibited the auxininduced hypocotyl elongation . Thus, OA and CA sensitive protein phosphatases are very probably positive regulators in the signaling pathway between auxin perception and H ATPase phosphorylation.
These results suggest that phosphorylation of the penultimate Thr of the H ATPase is required for auxininduced hypocotyl elongation. DISCUSSION Auxin Activates the Plasma Membrane H ATPase via Phosphorylation and Subsequently Induces Hypocotyl Elongation Auxin induced elongation of plant organs such as the stem, hypocotyl, and coleoptile is generally explained by the acid growth theory, in which the plasma membrane H ATPase plays a central role . However, the signaling pathway from Olaparib selleck auxin perception to H ATPase activation has not been fully explored to date. In this study, we showed that IAA enhanced the phosphorylation level of the penultimate Thr of the plasma membrane H ATPase in Arabidopsis hypocotyls within 10 min without altering the amount of the H ATPase . Then, a 14 3 3 protein bound to the phosphorylated H ATPase, resulting in an elevation of the catalytic activity of the H ATPase . IAA induced phosphorylation of the H ATPase approximately 5 min prior to hypocotyl elongation , and the P type ATPase inhibitor vanadate, suppressed the hypocotyl elongation.
These observations indicate that auxin induces elongation via activation of the H ATPase by phosphorylation of the penultimate Thr. To our knowledge, this is the first experimental evidence to clarify the activation mechanism of the H ATPase during early phase auxininduced clomifene elongation. A global quantitative analysis of the Arabidopsis phosphoproteome showed that the phosphorylation level of the penultimate Thr of AHA1 was elevated at 1, 3, and 6 h after application of 100 mM IAA in Arabidopsis suspension cells , indicating that the auxin induced H ATPase phosphorylation might also occur in tissues other than the etiolated hypocotyls and that the phosphorylation is maintained for much longer than 60 min.
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