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isom lab @ the university of miami
pH regulation of cell signaling systems
high content screening
PH REGULATION OF G PROTEIN-COUPLED RECEPTORS
G protein-coupled receptors (GPCRs) are both the largest class of cell surface receptors in humans, and the receptor family most targeted by therapeutic drugs. Major efforts in the lab are focused on understanding how acidic pH changes regulate GPCR-drug interactions and signaling. Our objective is to identify GPCR-drug interactions (in)capable of endomembrane signaling, and (in)sensitive to disease-related acidosis.
PH REGULATION OF CELL SURFACE RECEPTORS
Our lab studies proteins and cell-surface receptors that detect and transduce cellular signals. In certain biological scenarios—cancer, hypoxia, nutrient stress, inflammation, endocytosis—these signaling proteins encounter acidic conditions. Our lab uses experimental and computational approaches to understand how signaling proteins and their constituent networks are regulated by such pH changes.
The need for pH regulation of molecular, cellular, and physiological processes is wide-ranging, as it exists in all domains of life and viruses. At the molecular level, pH changes act to regulate the structure, function, and interaction of proteins, RNA, DNA, lipids, metabolites, and drugs. pH effects in proteins, which are mediated by ionizable side chains and electrostatic forces, are difficult to predict from sequence and structure. In the lab we develop informatics algorithms to address this challenge, and validate our findings at the lab bench using a variety of approaches.
INFORMATICS & PRECISION MEDICINE
Genome, exome, and RNA sequencing continually expand the potential to understand protein function in terms of genetic variation. While some DNA and RNA variation is the causal factor in disease and adverse drug responses, most variations are harmless. We try to understand which variations matter: e.g. cause changes in drug efficacy and pH sensitivity. Using big data informatics and scalable cell-based functional assays, we are developing approaches that will allow us experimentally interrogate how such variations regulate GPCR signaling systems.
PH REGULATION OF STRESS-RESPONSE SIGNALING NETWORKS
A surprising variety of conditions cause a significant drop in cellular pH, such as nutrient limitation, oxidative stress, and ischemia. Recent studies indicate that these acidic pH signals serve a cytoprotective function by mediating unexpected changes in cellular biochemistry and signaling. Examples include changes in the material properties of cytoplasm, and assembly of proteins into phase-separated macromolecular structures. We aim to identify the signaling cascades that regulate these processes.
PHASE-SEPARATED MACROMOLECULAR STRUCTURES
We are also interested in identifying proteins that are predisposed to assemble into phase-separated structures in response to pH changes. This interest is an extension of our studies on protein electroinformatics. Our objective is to identify topological patterns of ionizable side chains that are characteristic of higher-order structures stimulated by pH stress. In the longer term, our goal is to use these insights to develop predictive approaches that assess the scope of assembly formation in proteomes.
Our major computational efforts in the lab are focused on mining protein structural information for actionable intelligence. We use computational geometry as our primary conceptual framework for identifying residue topologies that are predictive of protein function. We develop our own algorithms, primarily in Python, and use the incredible high performance computing infrastructure in place at the University of Miami to routinely undertake millions of calculations on thousands of protein structures.
Our major experimental efforts are organized to broadly support our computational research. We have a variety of protein- and cell-based assays in place for rapidly quantifying biochemical parameters such as protein stability, ligand and drug affinity, receptor-drug interactions, and changes in cellular pH. Our core experimental infrastructure comprises technologies that include a liquid handling robotic screening platform, highly sensitive microplate readers, and novel cell-based model systems.
DAN ISOM, PH.D.
DISOM at MIAMI.EDU