The PID-directed shift in PIN polarity has been broadly accepted as one of the essential mechanisms for the regulation of auxin transport polarity. We verified that both PID functionality and its... Show moreThe PID-directed shift in PIN polarity has been broadly accepted as one of the essential mechanisms for the regulation of auxin transport polarity. We verified that both PID functionality and its subcellular localization do not depend on PDK1 function. However, by detailed analysis on these phenotypes and the expression of the auxin response reporter, we found the pdk1 pdk2 double mutant to be impaired in auxin transport in vascular tissues. Together with other mutant phenotypes, we suspect that PDK1 may be the master regulator of AGC1 kinases. The pdk1 pdk2 short root phenotype caused by phloem differentiation defects phenocopied the pax mutant. Complementation results of wild type and phosphomimic PAX in the pdk1 pdk2 background suggest that PDK1-dependent PAX phosphorylation and activation are essential for its full biological function. We also explain the molecular basis of PDK1 basal localization and the unnecessity of this polarity for vascular development. In addition, we investigated downstream action after PID phosphorylation. Several conserved tyrosine residues close to serine phosphorylation sites in the PIN1 and PIN2 HLs are mutated. Two of these tyrosines redundantly affected PIN polarity. However, PID mediated phosphorylation and tyrosine-based PIN trafficking are independent processes. Show less
Dudok, J.J.; Murtaza, M.; Alves, C.H.; Rashbass, P.; Wijnholds, J. 2016
Chondrocytes interact with their neighbours through their cartilaginous extracellular matrix (ECM). Chondrocyte__matrix interactions compensate the lack of cell__cell contact and are modulated by... Show moreChondrocytes interact with their neighbours through their cartilaginous extracellular matrix (ECM). Chondrocyte__matrix interactions compensate the lack of cell__cell contact and are modulated by proteoglycans and other molecules. The epiphyseal growth plate is a highly organized tissue responsible for long bone elongation. The growth plate is regulated by gradients of morphogens that are established by proteoglycans. Morphogens diffuse across the ECM, creating short- and long-range signalling that lead to the formation of a polarized tissue. Mutations affecting genes that modulate cell__matrix interactions are linked to several human disorders. Homozygous mutations of EXT1/EXT2 result in reduced synthesis and shortened heparan sulphate chains on both cell surface and matrix proteoglycans. This disrupts the diffusion gradients of morphogens and signal transduction in the epiphyseal growth plate, contributing to loss of cell polarity and osteochondroma formation. Osteochondromas are cartilage-capped bony projections arising from the metaphyses of endochondral bones adjacent to the growth plate. The osteochondroma cap is formed by cells with homozygous mutation of EXT1/EXT2 and committed stem cells/wild type chondrocytes. Osteochondroma serves as a niche (a permissive environment), which facilitates the committed stem cells/wild-type chondrocytes to acquire secondary genetic changes to form a secondary peripheral chondrosarcoma. In such a scenario, the microenvironment is the site of the initiating processes that ultimately lead to cancer. Show less
Polar cell-to-cell transport of plant hormone auxin mediated by plasma membrane (PM)-localized PIN-FORMED (PIN) auxin efflux carriers generates auxin gradients that provide positional information... Show morePolar cell-to-cell transport of plant hormone auxin mediated by plasma membrane (PM)-localized PIN-FORMED (PIN) auxin efflux carriers generates auxin gradients that provide positional information for various plant developmental processes. The apical-basal polar localization of the PIN proteins that determines the direction of auxin flow is directed by reversible phosphorylation of the PIN hydrophilic loop (PINHL). Here, we identified three evolutionarily conserved TPRXS(N/S) motifs within the PINHL, and proved that the central serine residues located in three motifs were phosphorylated by the PINOID (PID) serine/threonine kinase protein and its closely related AGC3 kinases WAG1 and WAG2. Loss-of-phosphorylation PIN1 or PIN2 protein (serine to alanine mutation) induced auxin-related plant defects in inflorescence development or root gravity-response, respectively, correlating with their apical-to-basal polarity changing in their expressing plant tissues. Furthermore, phosphorylation at the conserved serine residues is important for PIN protein PM localization, as loss-of-phosphorylation PIN1 protein exhibited internalized signals in plant embryos and enhanced vacuolar accumulation in dark-incubated Arabidopsis protoplast cells. Our data indicate that phosphorylation of our identified conserved serine residues in the PIN1HL by AGC3 kinases is required for proper PIN polar localization, and is thus essential for generating the differential auxin distribution that directs plant development. Show less
