A lipid code-dependent phosphoswitch directs PIN-mediated auxin efflux in Arabidopsis development

Abstract

18 19 Directional intercellular transport of the phytohormone auxin mediated by PIN FORMED (PIN) 20 efflux carriers plays essential roles in both coordinating patterning processes and integrating 21 multiple external cues by rapidly redirecting auxin fluxes. Multilevel regulations of PIN activity 22 under internal and external cues are complicated; however, the underlying molecular mechanism 23 remains elusive. Here we demonstrate that 3'-Phosphoinositide-Dependent Protein Kinase1 24 (PDK1), which is conserved in plants and mammals, functions as a molecular hub integrating 25 the upstream lipid signalling and the downstream substrate activity through phosphorylation. 26 Genetic analysis uncovers that loss-of-function Arabidopsis mutant pdk1.1 pdk1.2 exhibits a 27 plethora of abnormalities in organogenesis and growth, due to the defective PIN-dependent 28 auxin transport. Further cellular and biochemical analyses reveal that PDK1 phosphorylates D6 29 Protein Kinase to facilitate its activity towards PIN proteins. Our studies establish a lipid-30 dependent phosphorylation cascade connecting membrane composition-based cellular signalling 31 with plant growth and patterning by regulating morphogenetic auxin fluxes. 32 33 34 Due to a sessile lifestyle, plants have evolved strict developmental programming as well as 35 adaptive plasticity in response to diverse environmental stimuli, which is ensured by a 36 framework of multiple signalling pathways. The phytohormone auxin (indole-3-acetic acid, IAA) 37 plays a pivotal role in shaping plant growth and development, in a concentration-dependent 38 manner. The establishment and maintenance of local auxin minima, maxima and gradients rely 39 on directional intercellular transport, which is mediated by a subset of auxin influx carriers 40 AUXIN1 (AUX1)/LIKE AUX1s (LAXs) 1 , PIN (PIN-FORMED) efflux carriers 2,3 and ABCB-41 type transporters 4 . It has been well documented that the activity and polar localization of PIN 42 proteins are regulated by intracellular trafficking as well as post-translational modifications, 43 including reversible phosphorylation 5 . 44 PINs are phosphorylated at multiple serine/threonine residues by various protein kinases, 45 including PINOID (PID)/WAVY ROOT GROWTHs (WAGs) 6-8 , D6 PROTEIN KINASEs 46 (D6PK/D6PKL1~3, hereafter as D6PKs) 9,10 , PROTEIN KINASE ASSOCIATED WITH BRX 47 (PAX) 11 , MITOGEN PROTEIN KINASE3/6 (MPK3/6) 12 and CDPK-RELATED KINASE5 48 3 (CRK5) 13 , and dephosphorylated by Protein Phosphatase 2A (PP2A) 8,14 , Protein Phosphatase 6 49 (PP6) 15 and PP1 16 . PID-dependent phosphorylation and PP2A-mediated dephosphorylation 50 antagonize with each other 6,14 , acting as a binary switch determining the apical-basal targeting 51 of PIN proteins. More PID kinase activity results in more apical localization of PINs; conversely, 52 less PIN phosphorylation by PID gives rise to the reduced apical (or more basal) localization. In 53 contrast to PID, D6PKs and PAX phosphorylate basal-localized PINs to regulate their activity 17 . 54To dat...

PDK1 regulates auxin transport and Arabidopsis vascular development through AGC1 kinase PAX

“…5g,h) suggest that phospholipids are part of the auxin transport-controlling regulatory pathway. Most likely they are necessary for the PM association of PAX and of other AGC1 clade kinases 34 downstream of PDK1. Moreover, since the PDK1S0 isoform is clearly more active than full-length PDK1 in auto-and transphosphorylation in vitro (Extended Data Fig.…”

“…8m-o). In a parallel study, Tan and co-workers showed that reduced lateral root initiation is caused by lack of activation of the PDK1 substrates D6PK/ D6PKLs 34 . This indicates that PAX has a very specific role in vascular development.…”