The AGC kinase, PINOID, blocks interactive ABCB/PIN auxin transport

Abstract

Protein phosphorylation plays a key role in the regulation of polar auxin transport. 1 The AGC protein kinase, PINOID (PID), 2 has been shown to phosphorylate the hydrophilic loop of PIN proteins in vivo leading to apical PIN targeting.3 It appears that PID together with protein phosphatase PP2A/RCN1 act as a binary switch to control the balance of phosphorylation and dephosphorylation determining PIN polarity and controlling auxin flows. 4,5 Employing different phospho-proteomics approaches, plant ABCB proteins have recently shown to be phosphorylated in a so-called regulatory linker domain in analogy to their mammalian orthologs.6-9 Subsequently, the photoreceptor kinase, PHOTROPIN1 (phot1), was shown to phosphorylate ABCB19 and inhibit its auxin efflux activity. 10 Recently, PID was identified as a valid interaction partner with TWD1 by co-immunoprecipitation and shotgun LC-MS/MS analysis.11 In-vitro and yeast expression data indicated that PID specifically modulates ABCB1-mediated auxin efflux, which is dependent on its kinase activity. ABCB1/PID co-transfection in N. benthamiana revealed that PID phosphorylates at S634 of the linker in the absence of TWD1 enhancing ABCB1-mediated auxin efflux. PID had a negative impact on ABCB1 in triple ABCB1/PID/TWD1 co-transfection. This suggests that PID determines ABCB1 activity by means of protein phosphoryation in an action that is controlled by TWD1-PID interaction.However, it is an open question if altered PIN polarity is indeed directly caused by PIN phosphorylation or not simply the Plant growth and development is determined by intracellular and intercellular auxin gradients that are controlled at first hand by auxin efflux catalysts of the aBCB/PGP and Pin families. aBCB transport activity was shown to be counter-actively regulated by protein phosphorylation by the aGC protein kinase, PinoiD (PiD), that is coordinated by interaction with the immunophilin-like FKBP42, tWiStED DWarF1 (tWD1). in contrast, PiD was shown to determine Pin polarity, however, the direct impact of PiD on Pin activity has yet not been tested. Co-expression in yeast indicates that PiD had no effect on Pin1,2 alone but specifically inhibits interactive aBCB1-Pin1/Pin2 auxin efflux in an action that is dependent on its kinase activity. Pin1-PiD co-transfection in n. benthamiana revealed that PiD blocks Pin1-mediated auxin efflux without changing Pin1 location. in summary, these data provide evidence that PiD phosphorylation does not only determine Pin polarity but also has a direct impact on transport activity of the activity of the binary Pin-aBCB1 complex.

PINOID AGC Kinases Are Necessary for Phytochrome-Mediated Enhancement of Hypocotyl Phototropism in Arabidopsis

“…8). Because it has been reported that PID functions as a negative regulator for auxin transport activity of PIN proteins interacting with ABCB1 proteins (Wang et al, 2012), it seems that such a regulatory mechanism also participates in phototropic enhancement. Additional studies are necessary to clarify the molecular mechanisms underlying phytochrome-mediated phototropic enhancement.…”

Expression of TWISTED DWARF1 lacking its in‐plane membrane anchor leads to increased cell elongation and hypermorphic growth

“…This structural and functional analogy suggests that the principal role of TWD1 might be close to its FKBP38 ortholog in mediating ABCB folding at the cytoplasmic ER surface (Wu et al ., ) and that its interaction with ABCBs at the plasma membrane recently described in Wang et al . () might be secondary. However, in contrast with TWD1, absence of the FKBP38 membrane anchor leads to FKBP38 mis‐targeting and subsequent apoptosis (Shirane and Nakayama, ), while heterologous expression of a truncated version of FKBP38 abolishes its inhibition of CFTR synthesis but does influence its role in post‐translational folding.…”

“…Using a wide range of methods and constructs, TWD1 has been localized independently to several membrane locations, including the PM, the tonoplast and the ER (Kamphausen et al ., ; Geisler et al ., ; Henrichs et al ., ; Wang et al ., , ). Remarkably, in the root epidermis TWD1 immunodecoration was most pronounced at lateral PM domains with stronger labeling at the outward‐facing side, in line with their proposed function in preventing lateral apoplastic reflux (Wang et al ., ).…”

A leaf shape mutant provides insight into PINOID Serine/Threonine Kinase function in cucumber (Cucumis sativus L.)

“…Furthermore, CsPID is localized to all parts of the onion cell (Figure B), consistent with the pattern of auxin content. However, PID protein interacts with PIN protein on the plasma membrane in Arabidopsis (Wang et al ).…”