TY  - JOUR
A1  - Pollitt, Alice Y.
A1  - Poulter, Natalie S.
A1  - Gitz, Eelo
A1  - Navarro-Nuñez, Leyre
A1  - Wang, Ying-Jie
A1  - Hughes, Craig E.
A1  - Thomas, Steven G.
A1  - Nieswandt, Bernhard
A1  - Douglas, Michael R.
A1  - Owen, Dylan M.
A1  - Jackson, David G.
A1  - Dustin, Michael L.
A1  - Watson, Steve P.
T1  - Syk and Src Family Kinases Regulate C-type Lectin Receptor 2 (CLEC-2)-mediated Clustering of Podoplanin and Platelet Adhesion to Lymphatic Endothelial Cells*
JF  - The Journal of Biological Chemistry
N2  - The interaction of CLEC-2 on platelets with Podoplanin on lymphatic endothelial cells initiates platelet signalling events that are necessary for prevention of blood-lymph mixing during development. In the present study, we show that CLEC-2 signalling via Src family and Syk tyrosine kinases promotes platelet adhesion to primary mouse lymphatic endothelial cells at low shear. Using supported lipid bilayers containing mobile Podoplanin, we further show that activation of Src and Syk in platelets promotes clustering of CLEC-2 and Podoplanin. Clusters of CLEC-2-bound Podoplanin migrate rapidly to the centre of the platelet to form a single structure. Fluorescence life-time imaging demonstrates that molecules within these clusters are within 10 nm of one another and that the clusters are disrupted by inhibition of Src and Syk family kinases. CLEC-2 clusters are also seen in platelets adhered to immobilised Podoplanin using direct stochastic optical reconstruction microscopy (dSTORM). These findings provide mechanistic insight by which CLEC-2 signalling promotes adhesion to Podoplanin and regulation of Podoplanin signalling thereby contributing to lymphatic vasculature development.
KW  - endothelial cell
KW  - lipid bilayer
KW  - platelet receptor
KW  - tyrosine-protein kinase
KW  - CLEC-2 ITAM
KW  - podoplanin
KW  - Src family
KW  - kinase Syk
Y1  - 2014
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-120770
VL  - 289
IS  - 52
ER  - 
TY  - JOUR
A1  - Owen, Dylan M.
A1  - Sauer, Markus
A1  - Gaus, Katharina
T1  - Fluorescence localization  microscopy
JF  - Communicative & Integrative Biology
N2  - Localization microscopy techniques are super-resolution fluorescence imaging methods based on the detection of individual molecules. Despite the relative simplicity of the microscope setups and the availability of commercial instruments, localization microscopy faces unique challenges. While achieving super-resolution is now routine, issues concerning data analysis and interpretation mean that revealing novel biological insights is not. Here, we outline why data analysis and the design of robust test samples may hold the key to harness the full potential of localization microscopy.
Y1  - 2012
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-124416
VL  - 5
IS  - 4
ER  -