Kenneth W. Dunn, PhD

Our laboratory studies endocytic membrane transport in polarized epithelial cells. The function of an epithelium depends upon the polar distribution of transporters and receptors on either the lumenal or basal side of the monolayer. We have shown that endocytosis is critically important to this polarity as the apical and basolateral membranes of epithelial cells are continuously and thoroughly intermixed in endosomes where proteins are sorted for return to the proper domain of the plasma membrane. We have developed powerful methods of quantitative confocal microscopy to address membrane transport in cells labeled with fluorescent conjugates of endocytic ligands and expressing GFP chimeras of endocytic regulatory proteins. The functions of regulatory proteins are evaluated through the expression of mutant proteins, and through novel methods of 3-dimensional, multi-parameter microscopy of living cells. This approach allows us to correlate defined steps of endocytic transport with the dynamic bahaviors of specific regulators. More recently our laboratory has begun using biochemical analyses of endosomes isolated by subcellular fractionation to identify proteins that regulate polar sorting in endosomes. Our laboratory has open access to the latest equipment for 3-dimensional microscopy, including four confocal microscopes, two multi-photon microscopes and an iterative deconvolution microscope system. We are also actively involved in developing advanced methods of microscopy and image processing. The equipment and research activities of our imaging facility are described at: Indiana Center for Biological Microscopy