Laboratory of Plasma Membrane Receptors
Head: Andrzej SOBOTA
Staff: Kinga Borzęcka (PhD student), Maciej Czerkies (PhD student), Katarzyna Kwiatkowska, Anna Łukasik (PhD student), Ewelina Marszałek-Sadowska, Kazimiera Mrozińska, Agnieszka Płuciennikowska (PhD student), Agata Samonek (PhD student)
Research profile:
Our research is focused on understanding the mechanisms of signal generation by immunoreceptors of macrophages and monocytes that are involved in phagocytosis and inflammatory responses. We aim to elucidate signaling path- ways behind the FcγRIIA-mediated uptake of pathogen and TLR4-dependent responses to microbial cell wall compo- nents such as LPS. The essential approach is to examine how the activated receptors initiate a cascade of events leading to reorganization of membrane constituents, activation of tyrosine kinases and phosphoinositide kinases. Specifically, we investigate the involvement of plasma membrane rafts in immunoreceptor signaling, the role of the sphingomyelin cycle and ceramide generation in the transduction of signals by the receptor, the role of PIP2 in modulation of submembrane cytoskeleton and in internalization of membrane-associated particles.
Methods:
• cloning
• cell transfection and production of recombinant proteins
• silencing of gene expression
• confocal and immuno-electron microscopy
• surface plasmon resonance
• cell fractionation and raft isolation
• ELISA and FACS analysis
Current research activities:
• elucidating the mechanisms governing an association of activated Fcγ receptor IIA and Toll-like receptors with rafts of the plasma membrane – an engagement of acid sphingomyelinase and ceramide; our approaches range from gene silencing through immunoassays to enzymatic analysis
• plasma membrane rafts as centres of PIP5-kinase activation and PIP2 turnover; expression of PIP5-kinase and its fragments is performed to reveal how the kinase is recruited to the plasma membrane
• studies on the molecular composition of detergent-resistant membrane domains (DRM) and how it affects a raft involvement in the immunoreceptors signaling
• exploration of suitability of lysenin – sphingomyelin-binding protein, as a tool for studies of sphingomyelin-cycle during Fcγ receptor activation; the mechanism of inter- action of lysenin with sphingomyelin in membranes is studied by variety of biophysical approaches
• ultrastructural studies on participation of plasma membrane domains (rafts) in signal generation by Fcγ receptor IIA and Toll-like receptor 4
• studies of the effect of Fcγ receptors and TLR4 activation on organization of cortical actin cytoskeleton during the receptor-mediated phagocytosis and inflammatory responses
Selected publications:
Kwiatkowska K. (2010) One lipid, multiple functions: how various pools of PIP2 are created in the plasma membrane. Cellular and Molecular Life Sciences, 67: 3927-3946.
Józefowski S., Czerkies M., Łukasik A., Bielawska A., Bielawski J., Kwiatkowska K., Sobota A. (2010) Ceramide and ceramide 1-phosphate are negative regulators of TNF-a production induced by lipopolysaccharide. Journal of Immunology, 185: 6960-6973.
Kulma M., Hereć M., Grudziński W., Anderluh G., Gruszecki W.I., Kwiatkowska K., Sobota A. (2010) Sphingomyelin-rich domains are sites of lysenin oligomerization: implications for raft studies. Biochimica et Biophysica Acta, 1798: 471-481.
Szymańska E, Korzeniowski M, Raynal P, Sobota A, Kwiatkowska K. (2009) Contribution of PIP-5 kinase I alpha to raft-based FcγRIIA signaling. Experimental Cell Research, 315: 981-895.
Hereć M., Gagoś M., Kulma M., Kwiatkowska K., Sobota A., Gruszecki W. (2008) Secondary structure and orientation of the pore-forming toxin lysenin in a sphingomyelin-containing membrane. Biochimica et Biophysica Acta, 1778: 872-879.
Gładkowska E., Sobota A., Czuryło E., Kwiatkowska K. (2008) Expression of PtdIns(4,5)P2-binding proteins lowers PtdIns(4,5)P2 level and inhibits FcγRIIA-mediated cell spreading and phagocytosis. European Journal of Immunology, 38: 260-276.
