Joseph E. Mazurkiewicz
Professor and Director of AMC Imaging Core Facility
University of Colorado Health Science Center
1. Cellular biology of the Follicle Stimulating Hormone Receptor (FSHR), the G-protein Coupled Receptor (GPCR) for the anterior pituitary hormone, follicle stimulating hormone (FSH).
2. Development and application of various imaging modalities using single and multi-photon confocal microscopy such as fluorescence resonance energy transfer (FRET), fluorescence spectral imaging and linear unmixing, and the application of fluorescence biosensors.
The G-protein coupled receptor (GPCR) superfamily is the largest protein family known and its members are involved in all types of stimulus-response pathways, from intercellular communication to physiological senses. In particular, it is well accepted that the FSHR, the pituitary glycoprotein hormone receptor for follicle stimulating hormone (FSH), can transduce signals not only through the protein kinase A (PKA) pathways but also through mitogen activated protein kinase (MAPK), protein kinase B (PKB) and calcium signaling pathways. The macromolecular cytoplasmic interfacial membrane complexes that provide a framework for discrimination between these pathways or coordination of these pathways is only now being revealed.
The Mazurkiewicz lab has developed and applied biophysical (e.g., FRET) and imaging modalities (e.g., confocal microscopy) to examine the molecular interaction of GPCRs on the cell surface of live cells, and to visually demonstrate several signaling pathways in live cells in real time that include stimulus induced PKC-GFP isoform translocations and the sites of PI3K activity using fluorescent monitors of signaling pathways. These modalities are being used to investigate the interaction of FSH with its receptor and the consequences of this interaction on intracellular signaling.
We hypothesize that the FSH/FSHR complex intersects with different signaling pathways that are located in several macromolecular complexes or scaffolds that are present at the cytoplasmic face of the plasma membrane of target cells that are either preformed or that form subsequent to FSH binding to the FSHR. Two posibilities are envisioned: (a) the [FSH/FSHR] complex associates with arrestin in endosomes during desensitization or (b) the [FSH/FSHR] complex participates in a microdomain on early endosomes that has been organized by Rab5 and a newly discovered adapter protein. APPL.
Three specific aims have been developed to test this hypothesis: (1) Determine the spatial distribution of FSHR in lipid rafts/caveolae in both the hormone bound and unbound state. (2) Define the intracellular trafficking pathway of activated-FSHR in association with arrestin to clathrin-coated vesicles in endosomes and with an early endosomal compartment bearing Rab5 and APPL. (3) Establish the signaling pathways with which the FSH/FSHR complexes associate upon activation. The recognition of signaling scaffolds in the membrane provides an important advancement and an opportunity to target specific microdomains so as to differentially influence receptor activity within them. The resultant information from these studies could be used to guide development of more potent antagonists or mimetics of follitropin action, which could be clinically useful. In addition, such information could aid in a better appreciation of how this family of hormones binds to and activates their receptors.
As director of the Albany Medical College Imaging Core Facility, Dr. Mazurkiewicz is involved in a variety of collaborations that utilize confocal microscopy and scanning electron microcopy in the analysis of mammalian cell signaling, bacterial biofilm technology and cellular microbiology of the interaction of the pathogenic bacterium, Francisella tularensis, with the macrophage.
- Thomas, R.M., Nechamen, C.A., Mazurkiewicz, J.E., Muda, M., Palmer, S. and Dias, J.A. - FSH Receptor forms and shows evidence of a C-terminal proteolytic processing. Endocrinology 148:1987-1995, 2007.
- Lynch, D.L., Fountain, T.L., Mazurkiewicz, J.E. and Banas, J.A. - Glucan-binding proteins are essential for shaping Streptococcus mutans biofilm architecture, FEMS Microbiol. Lett. 268:158-165, 2007.
- Banas, J.A., Fountain, T.L., Mazurkiewicz, J.E., Sun, K. and Vickerman, M.M., 2007, Streptococcus mutans Glucan-binding protein-A affects Streptococcus gordonii biofilm architecture. FEMS Microbiol Lett 267:80-88.
- Herrick-Davis, K., Weaver, B.A., Grinde, E. and Mazurkiewicz, J.E. - Serotonin 5-HT2C Receptor Homodimer Biogenesis in the Endoplasmic Reticulum: Real-time visualization with confocal fluorescence resonance energy transfer. J. Biol. Chem. 281:27109-27116, 2006
- Herrick-Davis, K., Grinde, E. Timothy J. Harrigan, T.J. and Mazurkiewicz, J.E., 2005, Inhibition of serotonin 5-HT2C receptor function through heterodimerization: Receptor dimers bind two molecules of ligand and one G-protein. J. Biol. Chem. 280:40144-40151.