The Cellular Physiology Core provides a series of graded in vitro model systems and technologies for studying the function and regulation of transport and other membrane proteins in isolated expression systems and organized epithelia. The Core will interact with the other Cores in such a way that studies of transport proteins and interacting proteins developed either in model systems or isolated tubules may be exploited by direct measures of their function in in vitro systems that also lend themselves to further analysis by imaging. The Core technologies also are directly applicable to the study of plasma membrane proteins identified by novel model systems. To accomplish these goals, the Core will: a) provide a center for expression of transport proteins and regulatory/interacting proteins in isolated in vitro systems, which will permit direct assessment of the influence of these interactions on electrophysiologic characteristics both at the macroscopic and single channel level. These systems include Xenopus oocytes and naive cell expression systems (e.g., HEK-293 cells) and will permit direct measure of plasma membrane expression of channels and their electrophysiologic features;b) establish systems to expand the study of transport proteins and interacting proteins/pathways in organized epithelia either natively expressing the transport proteins or in model epithelia (e.g., MDCK and FRT cells) where transport protein mutations may be evaluated more fully than in single cell expression systems. These techniques will include standard voltage clamp for assessment of short-circuit current and transepithelial resistance and direct measures of tissue capacitance;c) provide methods for modulating gene expression in epithelia. Recombinant viruses will be generated to allow reconstitution of wild-type or mutated channel subunits and expression of other genes of interest. Silencing RNAs will be expressed using recombinant viruses and lipid-mediated transfer methods to permit downregulation of gene expression;and d) provide analysis of post-translational modifications of transport proteins and regulatory proteins, including phosphorylation, ubiquitination, glycosylation and palmitoylation using both biochemical and mass spectrometry approaches.
The Pittsburgh Center for Kidney Research Cellular Physiology Core provides mechanistic analyses of the functions of membrane transport and other associated proteins through a series of graded in vitro model systems. This Core interfaces with and complements the other Cores and has the overall goal of elucidating at a molecular and cellular level the function and regulation of key proteins involved in kidney diseases.
|Sheng, Shaohu; Chen, Jingxin; Mukherjee, Anindit et al. (2018) Thumb domains of the three epithelial Na+ channel subunits have distinct functions. J Biol Chem 293:17582-17592|
|Hughes, Andrew D; Lakkis, Fadi G; Oberbarnscheidt, Martin H (2018) Four-Dimensional Imaging of T Cells in Kidney Transplant Rejection. J Am Soc Nephrol 29:1596-1600|
|Theodoraki, M-N; Hoffmann, T K; Jackson, E K et al. (2018) Exosomes in HNSCC plasma as surrogate markers of tumour progression and immune competence. Clin Exp Immunol 194:67-78|
|Apodaca, Gerard (2018) Role of Polarity Proteins in the Generation and Organization of Apical Surface Protrusions. Cold Spring Harb Perspect Biol 10:|
|Jackson, Travis C; Kotermanski, Shawn E; Kochanek, Patrick M et al. (2018) Oxidative Stress Induces Release of 2'-AMP from Microglia. Brain Res :|
|Balchak, Deidra M; Thompson, Rebecca N; Kashlan, Ossama B (2018) The epithelial Na+ channel ? subunit autoinhibitory tract suppresses channel activity by binding the ? subunit's finger-thumb domain interface. J Biol Chem 293:16217-16225|
|Sun, Zhihao; Brodsky, Jeffrey L (2018) The degradation pathway of a model misfolded protein is determined by aggregation propensity. Mol Biol Cell 29:1422-1434|
|Boyd-Shiwarski, Cary R; Shiwarski, Daniel J; Roy, Ankita et al. (2018) Potassium-regulated distal tubule WNK bodies are kidney-specific WNK1 dependent. Mol Biol Cell 29:499-509|
|Kashlan, Ossama B; Kinlough, Carol L; Myerburg, Michael M et al. (2018) N-linked glycans are required on epithelial Na+ channel subunits for maturation and surface expression. Am J Physiol Renal Physiol 314:F483-F492|
|Jackson, Edwin K; Mi, Eric; Ritov, Vladimir B et al. (2018) Extracellular Ubiquitin(1-76) and Ubiquitin(1-74) Regulate Cardiac Fibroblast Proliferation. Hypertension 72:909-917|
Showing the most recent 10 out of 380 publications