Reference : Overexpression of the partially activated alpha(IIb)beta3D723H integrin salt bridge m...
Scientific journals : Article
Life sciences : Biochemistry, biophysics & molecular biology
http://hdl.handle.net/10993/14968
Overexpression of the partially activated alpha(IIb)beta3D723H integrin salt bridge mutant downregulates RhoA activity and induces microtubule-dependent proplatelet-like extensions in Chinese hamster ovary cells
English
Schaffner-Reckinger, Elisabeth mailto [University of Luxembourg > Faculty of Science, Technology and Communication (FSTC) > Life Science Research Unit >]
Salsmann, Alexandre mailto [University of Luxembourg > Faculty of Science, Technology and Communication (FSTC) > Life Science Research Unit >]
Debili, Najet [INSERM U790, Université Paris XI, Villejuif]
Bellis, Julien [ESBS, Institut Gilbert Laustriat (CNRS/UMR-7175), ULP, Illkirch, France]
De Mey, Jan [ESBS, Institut Gilbert Laustriat (CNRS/UMR-7175), ULP, Illkirch, France]
Vainchenker, William [INSERM U790, Université Paris XI, Villejuif]
Ouwehand, Willem H. [University of Cambridge & National Health Service Blood and Transplant, Cambridge, UK]
Kieffer, Nelly [University of Luxembourg > Faculty of Science, Technology and Communication (FSTC) > Life Science Research Unit >]
2009
Journal of Thrombosis and Haemostasis [=JTH]
Blackwell Publishing
7
7
1207-1217
Yes (verified by ORBilu)
1538-7933
1538-7836
Oxford
United Kingdom
[en] BACKGROUND: We have recently reported a novel mutation in the beta3 subunit of the platelet fibrinogen receptor (alpha(IIb)beta3D723H) identified in a patient with dominantly inherited macrothrombocytopenia, and we have shown that this mutation promotes a new phenotype in Chinese hamster ovary (CHO) cells, characterized by fibrinogen-dependent, microtubule-driven proplatelet-like cell extensions. RESULTS: Here we demonstrate that the partially activated alpha(IIb)beta3D723H or alpha(IIb)beta3D723A salt bridge mutants, but not fully activated alpha(IIb)beta3 mutants, cause this phenotype. Time-lapse videomicroscopy clearly differentiated these stable microtubule-driven and nocodazole-sensitive extensions from common dynamic actin-driven pseudopodia. In addition, overexpression of a mitochondrial marker confirmed their functional role in organelle transport. Comparative immunofluorescence analysis of the subcellular localization of alpha(IIb)beta3, the focal adhesion proteins talin or vinculin and actin revealed a similar membrane labeling of CHO cell extensions and CD34+-derived megakaryocyte proplatelets. Mutant alpha(IIb)beta3D723H signaling was independent of Src, protein kinase C or phosphoinositide 3-kinase, but correlated with decreased RhoA activity as compared with wild-type alpha(IIb)beta3 signaling, reminiscent of integrin signaling during neurite outgrowth. Accordingly, overexpression of constitutively active RhoA in CHO alpha(IIb)beta3D723H cells prevented protrusion formation on fibrinogen. Most interestingly, RhoA/ROCK inhibition was necessary, but not sufficient, and integrin activity was additionally required to induce CHO cell extension formation. CONCLUSIONS: CHO alpha(IIb)beta3D723H cell protrusions and megakaryocyte proplatelets, like neuronal cell neurites, result from a common integrin-dependent signaling pathway, promoting strongly decreased RhoA activity and leading to microtubule-driven formation of cytoplasmic extensions.
http://hdl.handle.net/10993/14968
10.1111/j.1538-7836.2009.03494.x
Elisabeth Schaffner-Reckinger and Alexandre Salsmann contributed equally to this work

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