- G. Cascardo (1)
- C. Cassinelli (1)
- G. Ceccone (1)
- G.W. Esse van (2)
- M. Gallet (1)
- D. Gilliland (1)
- A. Haeger (1)
- A. Hoek van (1)
- J.P. Knox (1)
- A. Kruis (1)
- I. Liakos (1)
- J. Luchtenberg (1)
- M. Morra (1)
- M.D. Nagel (1)
- M. Rodrigues-Valverde (1)
- H.A. Schols (1)
- S. Siboni (1)
- R.P. SURENDRAN (1)
- M. Vayssade (1)
- M.B. Veen van (1)
- E. Velzenberger (1)
- R.P. Verhoef (1)
- P. Vigneron (1)
- C.D. Volpe (1)
- S.C. Vries de (2)
- A.H. Westphal (2)
Visualization of BRI1 and BAK1(SERK3) membrane receptor heterooligomers during brassinosteroid signaling.
Bücherl, C.A. ; Esse, G.W. van; Kruis, A. ; Luchtenberg, J. ; Westphal, A.H. ; Aker, J.C.M. ; Hoek, A. van; Albrecht, C. ; Borst, J.W. ; Vries, S.C. de - \ 2013
Plant Physiology 162 (2013)2. - ISSN 0032-0889 - p. 1911 - 1925.
agrobacterium-mediated transformation - growth-factor receptors - gsk3-like kinase bin2 - transcription factors - plant-growth - arabidopsis-thaliana - extracellular domain - chemokine receptors - plasma-membrane - gene-expression
The leucine-rich repeat receptor-like kinase BRASSINOSTEROID-INSENSITIVE1 (BRI1) is the main ligand-perceiving receptor for brassinosteroids (BRs) in Arabidopsis (Arabidopsis thaliana). Binding of BRs to the ectodomain of plasma membrane (PM)-located BRI1 receptors initiates an intracellular signal transduction cascade that influences various aspects of plant growth and development. Even though the major components of BR signaling have been revealed and the PM was identified as the main site of BRI1 signaling activity, the very first steps of signal transmission are still elusive. Recently, it was shown that the initiation of BR signal transduction requires the interaction of BRI1 with its SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE (SERK) coreceptors. In addition, the resolved structure of the BRI1 ectodomain suggested that BRI1-ASSOCIATED KINASE1 [BAK1](SERK3) may constitute a component of the ligand-perceiving receptor complex. Therefore, we investigated the spatial correlation between BRI1 and BAK1(SERK3) in the natural habitat of both leucine-rich repeat receptor-like kinases using comparative colocalization analysis and fluorescence lifetime imaging microscopy. We show that activation of BR signaling by exogenous ligand application resulted in both elevated colocalization between BRI1 and BAK1(SERK3) and an about 50% increase of receptor heterooligomerization in the PM of live Arabidopsis root epidermal cells. However, large populations of BRI1 and BAK1(SERK3) colocalized independently of BRs. Moreover, we could visualize that approximately 7% of the BRI1 PM pool constitutively heterooligomerizes with BAK1(SERK3) in live root cells. We propose that only small populations of PM-located BRI1 and BAK1(SERK3) receptors participate in active BR signaling and that the initiation of downstream signal transduction involves preassembled BRI1-BAK1(SERK3) heterooligomers.
Quantification of the BRI1 receptor in planta
Esse, G.W. van; Westphal, A.H. ; SURENDRAN, R.P. ; Albrecht, C. ; Veen, M.B. van; Borst, J.W. ; Vries, S.C. de - \ 2011
Plant Physiology 156 (2011)4. - ISSN 0032-0889 - p. 1691 - 1700.
green fluorescent protein - plasma-membrane receptor - growth-factor receptors - signal-transduction - arabidopsis-thaliana - kinase bri1 - gene-expression - erbb receptors - cell - root
In plants, green fluorescent protein (GFP) is routinely used to determine the subcellular location of fusion proteins. Here, we show that confocal imaging can be employed to approximate the number of GFP-labeled protein molecules present in living Arabidopsis (Arabidopsis thaliana) root cells. The technique involves calibration with soluble GFP to provide a usable protein concentration range within the confocal volume of the microscope. As a proof of principle, we quantified the Brassinosteroid Insensitive1 (BRI1) receptor fused to GFP, under control of its own promoter. The number of BRI1-GFP molecules per root epidermal cell ranges from 22,000 in the meristem and 130,000 in the elongation zone to 80,000 in the maturation zone, indicating that up to 6-fold differences in BRI1 receptor content exist. In contrast, when taking into account differences in cell size, BRI1-GFP receptor density in the plasma membrane is kept constant at 12 receptors µm-2 in all cells throughout the meristem and elongation zone. Only the quiescent center and columella cells deviate from this pattern and have 5 to 6 receptors µm-2. Remarkably, root cell sensitivity toward brassinosteroids appears to coincide with uniform meristem receptor density
Enzymatically-tailored pectins differentially influence the morphology, adhesion, cell cycle progression and survival of fibroblasts
Nagel, M.D. ; Verhoef, R.P. ; Schols, H.A. ; Morra, M. ; Knox, J.P. ; Ceccone, G. ; Volpe, C.D. ; Vigneron, P. ; Bussy, C. ; Gallet, M. ; Velzenberger, E. ; Vayssade, M. ; Cascardo, G. ; Cassinelli, C. ; Haeger, A. ; Gilliland, D. ; Liakos, I. ; Rodrigues-Valverde, M. ; Siboni, S. - \ 2008
Biochimica et Biophysica Acta. General subjects 1780 (2008)7-8. - ISSN 0304-4165 - p. 995 - 1003.
hairy ramified regions - growth-factor receptors - surface-chemistry - fibronectin conformation - arabinogalactan-proteins - monoclonal-antibodies - human keratinocytes - integrin binding - adsorbed fibronectin - rhamnogalacturonan-i
Improved biocompatibility and performance of biomedical devices can be achieved through the incorporation of bioactive molecules on device surfaces. Five structurally distinct pectic polysaccharides (modified hairy regions (MHRs)) were obtained by enzymatic liquefaction of apple (MHR-B, MHR-A and MHR-), carrot (MHR-C) and potato (MHR-P) cells. Polystyrene (PS) Petri dishes, aminated by a plasma deposition process, were surface modified by the covalent linking of the MHRs. Results clearly demonstrate that MHR-B induces cell adhesion, proliferation and survival, in contrast to the other MHRs. Moreover, MHR- causes cells to aggregate, decrease proliferation and enter into apoptosis. Cells cultured in standard conditions with 1% soluble MHR-B or MHR- show the opposite behaviour to the one observed on MHR-B and --grafted PS. Fibronectin was similarly adsorbed onto MHR-B and tissue culture polystyrene (TCPS) control, but poorly on MHR-. The Fn cell binding site (RGD sequence) was more accessible on MHR-B than on TCPS control, but poorly on MHR-. The disintegrin echistatin inhibited fibroblast adhesion and spreading on MHR-B-grafted PS, which suggests that MHRs control fibroblast behaviour via serum-adhesive proteins. This study provides a basis for the design of intelligently-tailored biomaterial coatings able to induce specific cell functions.