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Plihal, O.

Publications and source records attributed to Plihal, O..

2 recordsLinked to original sources
bioRxiv2019

Cytokinin fluoroprobe and receptor CRE1/AHK4 localize to both plasma membrane and endoplasmic reticulum

The plant hormone cytokinin regulates various cell and developmental processes, including cell division and differentiation, embryogenesis, activity of shoot and root apical meristems, formation of shoot and root lateral organs and others 1. Cytokinins are perceived by a subfamily of sensor histidine kinases (HKs), which via a two-component phosphorelay cascade activate transcriptional responses in the nucleus. Based on the subcellular localization of cytokinin receptors in various transient expression systems, such as tobacco leaf epidermal cells, and membrane fractionation experiments of Arabidopsis and maize, the endoplasmic reticulum (ER) membrane has been proposed as a principal hormone perception site 2-4. Intriguingly, recent study of the cytokinin transporter PUP14 has pointed out that the plasma membrane (PM)-mediated signalling might play an important role in establishment of cytokinin response gradients in various plant organs 5. However, localization of cytokinin HK receptors to the PM, although initially suggested 6, remains ambiguous. Here, by monitoring subcellular localizations of the fluorescently labelled natural cytokinin probe iP-NBD 7 and the cytokinin receptor ARABIDOPSIS HISTIDINE KINASE 4 (CRE1/AHK4) fused to GFP reporter, we show that pools of the ER-located cytokinin fluoroprobes and receptors can enter the secretory pathway and reach the PM. We demonstrate that in cells of the root apical meristem, CRE1/AHK4 localizes to the PM and the cell plate of dividing meristematic cells. Brefeldin A (BFA) experiments revealed vesicular recycling of the receptor and its accumulation in BFA compartments. Our results provide a new perspective on cytokinin signalling and the possibility of multiple sites of perception at PM and ER, which may determine specific outputs of cytokinin signalling.

plant biology↗
bioRxiv2019

Cell-surface receptors enable perception of extracellular cytokinins

Cytokinins are mobile multifunctional plant hormones with roles in development and stress resilience 1,2. Although cytokinin receptors are substantially localised to the endoplasmic reticulum 3-5, the cellular sites of cytokinin perception continue to be debated 1,6,7. Several cytokinin types display bioactivity 8,9 and recently a cell-specific cytokinin gradient was reported in roots 10. Yet, the importance of spatially heterogeneous cytokinin distribution and the specific cytokinin(s) that account for the different responses remain unclear. Here we show that cytokinin perception by plasma membrane receptors is an effective path for cytokinin response in root cells. Readout from a Two Component Signalling cytokinin-specific reporter (TCSn::GFP;11) is closely matched to intracellular cytokinin content, yet a proportion of bioactive cytokinins are detected in the extracellular fluid. Using cytokinins covalently linked to beads that could not pass the plasma membrane, we demonstrate that strong TCSn activation still occurs and that this response is greatly diminished in cytokinin receptor mutants. Although intracellular receptors play significant roles, we argue for a revision of concepts of cytokinin perception to include the spatial dimensions. In particular, selective ligand-receptor affinities, cellular localisation and tissue distribution of bioactive cytokinins, their receptors, transporters and inactivation enzymes appear all to be components of the signalling regulatory mechanisms.

plant biology↗