Robert D PhairIntegrative Bioinformatics Inc. · Consulting Services
Robert D Phair
PhD
About
75
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7,241
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Introduction
Additional affiliations
April 1983 - June 1996
July 1979 - June 1996
Education
September 1973 - December 1977
September 1964 - June 1968
Publications
Publications (75)
We need to effectively combine the knowledge from surging literature with complex datasets to propose mechanistic models of SARS-CoV-2 infection, improving data interpretation and predicting key targets of intervention. Here, we describe a large-scale community effort to build an open access, interoperable and computable repository of COVID-19 mole...
Background
It is increasingly recognized that intestinal cells can store lipids after a meal, yet the effect of this phenomenon on lipid absorption patterns in insulin resistance remains unknown.Methods
The kinetics of meal fat appearance were measured in insulin-sensitive (IS, n = 8) and insulin-resistant (IR, n = 8) subjects after sequential, iso...
Abstract Systems biology has experienced dramatic growth in the number, size, and complexity of computational models. To reproduce simulation results and reuse models, researchers must exchange unambiguous model descriptions. We review the latest edition of the Systems Biology Markup Language (SBML), a format designed for this purpose. A community...
Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a debilitating noncommunicable disease brandishing an enormous worldwide disease burden with some evidence of inherited genetic risk. Absence of measurable changes in patients’ standard blood work has necessitated ad hoc symptom-driven therapies and a dearth of mechanistic hypotheses re...
The saga of fluorescence recovery after photobleaching (FRAP) illustrates how disparate technical developments impact science. Starting with the classic 1976 Axelrod et al. work in Biophysical Journal, FRAP (originally fluorescence photobleaching recovery) opened the door to extraction of quantitative information from photobleaching experiments, la...
Genetically encoded fluorescent proteins, combined with fluorescence microscopy, are widely used in cell biology to collect kinetic data on intracellular trafficking. Methods for extraction of quantitative information from these data are based on the mathematics of diffusion and tracer kinetics. Current methods, though useful and powerful, depend o...
Studies of lipoprotein kinetics almost exclusively rely on steady-state approaches to modeling. Herein, we have used a non-steady state experimental design to examine the role of cholesteryl ester transfer protein in mediating high density lipoprotein triglyceride flux in vivo in rhesus macaques and therefore we developed an alternative strategy to...
Mechanistic modeling has the potential to transform how cell biologists contend with the inescapable complexity of modern biology. I am a physiologist-electrical engineer-systems biologist who has been working at the level of cell biology for the past 24 years. This perspective aims 1) to convey why we build models, 2) to enumerate the major approa...
Insulin control of fatty acid metabolism has long been deemed dominated by suppression of adipose lipolysis. The goal of the present study was to test the hypothesis that this single role of insulin is insufficient to explain observed fatty acid dynamics.
Fatty acid kinetics were measured during a meal tolerance test and insulin sensitivity assesse...
Seen from the perspective of funding organizations, investors, and the general public, the productivity of our world-wide biomedical research enterprise is declining despite increased investment. This opinion piece suggests a cause and a solution. The cause is the enormous complexity of human biology and pathophysiology. The unsolved human diseases...
The endomembrane system of eukaryotic cells uses membrane-enclosed carriers to move diverse macromolecules among different membrane-bound compartments, a requirement for cells to secrete and take up molecules from their environment. Two recycling pathways-biosynthetic and endocytic, each with specific lipid components-make up this system, with the...
Mechanistic gene network model. Diagram of the full mechanistic gene network model. Black arrows represent processes (chemical reactions, transport, or binding). Rectangles represent states. A state is a molecule or complex in a physiologic place. Places are represented by the background ivory or mauve bands of color and are labeled at the bottom o...
Corresponding equations of the computational model. Text file displaying the corresponding equations of the computational model shown in Additional file 1.
A Berkeley Madonna model file modified slightly from the model file generated automatically by ProcessDB. A Berkeley Madonna model file (MODE3680_20080326_final.mmd) containing all the equations and parameters for both WT and Tabby experiments, as well as the fits of the experimental data. This file can be run and examined using the free trial vers...
Berkeley Madonna variable names corresponding to those of the model diagram. Table showing Berkeley Madonna variable names corresponding to those of the model diagram shown in Additional file 1.
Additional time course fits of the WT and Tabby experimental data by the full mechanistic model. Time course fits of the WT and Tabby experimental data by the full mechanistic model that are not included in Figures 4, 5, 8 of the main text. Note, Eda is not included among the genes because differences in Eda expression in WT and Tabby mice are not...
Ectodysplasin-A appears to be a critical component of branching morphogenesis. Mutations in mouse Eda or human EDA are associated with absent or hypoplastic sweat glands, sebaceous glands, lacrimal glands, salivary glands (SMGs), mammary glands and/or nipples, and mucous glands of the bronchial, esophageal and colonic mucosa. In this study, we util...
The prevailing view of intra-Golgi transport is cisternal progression, which has a key prediction--that newly arrived cargo exhibits a lag or transit time before exiting the Golgi. Instead, we find that cargo molecules exit at an exponential rate proportional to their total Golgi abundance with no lag. Incoming cargo molecules rapidly mix with thos...
We imaged transcription in living cells using a locus-specific reporter system, which allowed precise, single-cell kinetic measurements of promoter binding, initiation and elongation. Photobleaching of fluorescent RNA polymerase II revealed several kinetically distinct populations of the enzyme interacting with a specific gene. Photobleaching and p...
DNA lesions interfere with DNA and RNA polymerase activity. Cyclobutane pyrimidine dimers and photoproducts generated by ultraviolet irradiation cause stalling of RNA polymerase II, activation of transcription-coupled repair enzymes, and inhibition of RNA synthesis. During the S phase of the cell cycle, collision of replication forks with damaged D...
Secretory protein trafficking relies on the COPI coat, which by assembling into a lattice on Golgi membranes concentrates cargo at specific sites and deforms the membranes at these sites into coated buds and carriers. The GTPase-activating protein (GAP) responsible for catalyzing Arf1 GTP hydrolysis is an important part of this system, but the mech...
A quantitative understanding of human folate metabolism is needed.
The objective was to quantify and interpret human folate metabolism as it might occur in vivo.
Adults (n = 13) received 0.5 nmol [(14)C]pteroylmonoglutamate (100 nCi radioactivity) plus 79.5 nmol pteroylmonoglutamate in water orally. (14)C was measured in plasma, erythrocytes, urine...
Genome structure and gene expression depend on a multitude of chromatin-binding proteins. The binding properties of these
proteins to native chromatin in intact cells are largely unknown. Here, we describe an approach based on combined in vivo
photobleaching microscopy and kinetic modeling to analyze globally the dynamics of binding of chromatin-as...
We have described procedures for collecting, processing, and analyzing kinetic data obtained by photobleaching microscopy of GFP-tagged chromatin proteins in nuclei of cultured living cells. These procedures are useful for characterizing the in vivo binding of chromatin proteins to their natural template--unperturbed, native chromatin in an intact...
In mitosis, chromosome, cytoskeleton, and organelle dynamics must be coordinated for successful cell division. Here, we present evidence for a role for Arf1, a small GTPase associated with the Golgi apparatus, in the orchestration of mitotic Golgi breakdown, chromosome segregation, and cytokinesis. We show that early in mitosis Arf1 becomes inactiv...
We have analyzed the kinetics of assembly and elongation of the mammalian RNA polymerase I complex on endogenous ribosomal
genes in the nuclei of living cells with the use of in vivo microscopy. We show that components of the RNA polymerase I machinery
are brought to ribosomal genes as distinct subunits and that assembly occurs via metastable inter...
Cytosolic coat proteins that bind reversibly to membranes have a central function in membrane transport within the secretory pathway. One well-studied example is COPI or coatomer, a heptameric protein complex that is recruited to membranes by the GTP-binding protein Arf1. Assembly into an electron-dense coat then helps in budding off membrane to be...
The ability to visualize protein dynamics and biological processes by in vivo microscopy is revolutionizing many areas of biology. These methods generate large, kinetically complex data sets, which often cannot be intuitively interpreted. The combination of dynamic imaging and computational modelling is emerging as a powerful tool for the quantitat...
The endocytic itineraries of lipid raft markers, such as glycosyl phosphatidylinositol (GPI)-anchored proteins and glycosphingolipids, are incompletely understood. Here we show that different GPI-anchored proteins have different intracellular distributions; some (such as the folate receptor) accumulate in transferrin-containing compartments, others...
The endocytic itineraries of lipid raft markers, such as glycosyl phosphatidylinositol (GPI)-anchored proteins and glycosphingolipids,
are incompletely understood. Here we show that different GPI-anchored proteins have different intracellular distributions;
some (such as the folate receptor) accumulate in transferrin-containing compartments, others...
The mammalian cell nucleus contains numerous sub-compartments, which have been implicated in essential processes such as transcription and splicing. The mechanisms by which nuclear compartments are formed and maintained are unclear. More fundamentally, it is not known how proteins move within the cell nucleus. We have measured the kinetic propertie...
This chapter describes a method that employs digital time-lapse imaging to obtain virtually continuous sets of data on the transit of vesicular stomatitis virus G protein– green fluorescent protein (VSVG-GFP) through the compartments of the secretory pathway in single cells after shift to permissive temperature. The VSVG-GFP distribution within the...
Quantitative imaging and photobleaching were used to measure ER/Golgi recycling of GFP-tagged Golgi proteins in interphase cells and to monitor the dissolution and reformation of the Golgi during mitosis. In interphase, recycling occurred every 1.5 hr, and blocking ER egress trapped cycling Golgi enzymes in the ER with loss of Golgi structure. In m...
Quantitative time-lapse imaging data of single cells expressing the transmembrane protein, vesicular stomatitis virus ts045 G protein fused to green fluorescent protein (VSVG–GFP), were used for kinetic modeling of protein traffic through the various compartments of the secretory pathway. A series of first order rate laws was sufficient to accurate...
![TABLE 1 . Plasma cholesterol levels and plasma [3H]cholesteryl...](profile/Robert-Phair/publication/13773015/figure/tbl1/AS:601618416427030@1520448416530/Plasma-cholesterol-levels-and-plasma-3Hcholesteryl-linoleate-kinetics_Q320.jpg)
Niemann-Pick C disease (NP-C) is a rare inborn error of metabolism with hepatic involvement and neurological sequelae that usually manifest in childhood. Although in vitro studies have shown that the lysosomal distribution of LDL-derived cholesterol is defective in cultured cells of NP-C subjects, no unusual characteristics mark the plasma lipoprot...
Increasingly, successful research on metabolic systems relies on teams of specialists. Because of the enormous complexity of these systems, many experimental groups have sought collaborations with theoreticians for data analysis and modeling. Predictably, cultural differences in scientific approach, methodology, assumptions, and language have led t...
Steady-state cytosolic calcium (Ca2+i) concentration in a vascular smooth muscle cell is determined by Ca2+ influx and Ca2+ extrusion across the plasma membrane, yet no means for determining the absolute magnitude of these transmembrane Ca2+ fluxes in the basal state of the resting cell has been devised. We now report a method that combines fluores...
Established cell lines are now widely used in experiments concerning vascular smooth muscle (VSM) function; however, considerable evidence suggests that cultured VSM cells are functionally different from VSM cells in intact blood vessels. In order to test the hypothesis that calcium signaling mechanisms are comparable in these two preparations, we...
Since the platelet-derived growth factor (PDGF)-induced increase in cellular inositol 1,4,5-trisphosphate (InsP3) has been found to decay to basal levels soon after the onset of PDGF exposure, it has been argued that activation of Ca2+ release from intracellular stores must be similarly transient. The possibility remains, however, that PDGF-induced...
Studies were carried out in three normolipidemic non-obese men with insulin-dependent diabetes mellitus (IDDM) and three normal men, to assess whether the clearance of postprandial Sf 100-400 lipoproteins is decreased in IDDM. Sf greater than 100 lipoproteins isolated from plasma 4.5 h after fat ingestion were labeled with 125I and injected into th...
We combined techniques of 45Ca efflux and computer-assisted kinetic analysis to investigate the effects of forskolin and caffeine on intracellular Ca2+ metabolism in intact rabbit aortic segments. When either 1 microM forskolin or 10 mM caffeine was present during the 45Ca load and efflux, the amount of 45Ca released from the tissue was reduced dur...
Evidence for and against the theory that cell calcium is causally involved in the pathogenesis of atherosclerosis is presented and evaluated. In particular, it is argued that: (1) arterial calcium is increased in atherosclerosis; (2) this increase in tissue calcium content is largely intracellular; (3) this increased intracellular calcium content i...
Several lines of evidence, including the reported ability of calcium channel blockers to prevent atherogenesis in cholesterol-fed rabbits, suggest that calcium mediates one or more of the pathologic changes in atherosclerosis. Moreover, it has long been known that calcium accumulates in atherosclerotic blood vessels. To test the hypothesis that a s...
A new method, based on computer-assisted kinetic analysis of 45Ca efflux data, was used to measure calcium contents and fluxes for extracellular and intracellular compartments in intact segments of rabbit aorta. After a 1-hour loading period, efflux data were collected for 8 hours using a flow-through tissue chamber. These long-term effluxes were n...
Previous work from this laboratory revealed the presence of at least three distinct intracellular calcium compartments in intact segments of rabbit aorta. In this study one of these intracellular compartments is shown to be sensitive to dinitrophenol and to increased extracellular phosphate. Intact aortic segments were loaded with 45Ca in bicarbona...
Reduction of body weight is commonly used to decrease the plasma lipids of patients with primary endogenous hypertriglyceridemia, but the effects of stabilized weight reduction on lipoprotein compositions and distribution are not well known. Since lipoprotein structures are perhaps as important in normal and abnormal metabolism and atherogenesis as...
The involvement of prostaglandins in the effects of arachidonic acid (20:4n-6) on insulin and glucagon release was investigated, using the isolated, perfused rat pancreas model. 20:4n-6, the substrate for dienoic prostaglandins, or 20:3n-3, a fatty acid that cannot be metabolized to prostaglandins were perfused over 55 min. 20: 4n-6 evoked triphasi...
Adrenal secretory rates of cortisol and arterial concentrations of adrenocorticotropin (ACTH) were measured in conscious trained dogs subjected to intravenous infusion of ACTH. To investigate the causal relation of ACTH to the secretion of cortisol, a mechanistic mathematical model based on current hypotheses of adrenocortical function was construc...
If newly formed adenosine is the mediator of active hyperemia in skeletal muscle, tissue adenosine must increase and remain elevated during sustained muscle contraction. We tested this prediction using isolated canine anterior calf muscles. Muscle samples were obtained before and during contraction by punch biopsy and adenosine was measured by spec...
A period of prolonged vasodilation follows flow-restricted exercise of skeletal muscle. We tested the hypothesis that adenosine participates in mediating this vascular response. Vascularly isolated, anterior calf muscles of anesthetized dogs were stimulated to contract at a rate of 4 twitches/sec. Blood flow was held constant at 12.5 +/- 1.3 ml/min...
The kinetics of apolipoproteins B and C were studied in 14 normal and hyperlipoproteinemic subjects after injection of exogenously (125)I-labeled very low density lipoprotein (VLDL) particles. Plasma radioactivities of apoB and apoC were determined over a period of 4 days in VLDL (d < 1.006) and total radioactivity in intermediate (IDL) (1.006 < d...
A model is proposed for the metabolism of plasma lipoprotein apoproteins based on studies of a hyperlipoproteinemic subject who received 2.5 mCi[3H]leucine intravenously. Measurements included apoprotein specific activities (apo-B and apo-C) of very low density lipoprotein (VLDL) and of three low density lipoprotein (LDL) subspecies, Sf 17 LDL, Sf...
Influence of capillary permeability on local cerebral blood flow (LCBF) estimated by the autoradiograhpic diffusible-indicator method was analyzed by computer simulation. Its influence increases with increasing flow. With normal perfusion rates in gray matter, capillary permeability coefficient x surface area (PS value) must exceed 0.12 cm3-s(-1)-g...
The measurement of local tissue blood flow has been an important contribution to our present knowledge of the physiology and pathophysiology of cerebral blood flow. The first and most widely used technique for this purpose is the autoradiographic diffusible indicator technique based on the principles of inert gas exchange developed by Kety (3). Its...
A mathematical model describing human iodide metabolism has been developed and studied utilizing Program SAAM 25 of Berman and Weiss. The compartmental model simulates known aspects of thyroid physiology, including return of trapped iodide from thyroid to plasma, first come-first served turnover of intrathyroidal iodine, leakage of iodotyrosine iod...
Cellular modeling is essential. Systems with thousands of interacting parts are simply beyond the capacity of human information processing (1), and computational approaches to cellular modeling are widely regarded as one of the grand challenges of 21 st century biology (2). This challenge is being approached from the perspective of genomics and pro...
The authors have investigated VLDL assembly by examining the intracellular distribution, residence kinetics in the endoplasmic reticulum (ER) and Golgi, and secretion of three VLDL apoproteins (apo). Primary hepatocytes from estrogen-induced chicks were labeled with ³H-leucine for 4h to reach constant levels of cellular labeled apoB, apoAI and apo...
Questions
Questions (12)
Many mitochondrial proteins are encoded by the nuclear genome. Do different cell types supply the same set of proteins to their mitochondria? Or do different cell types fine-tune mitochondrial function to suit their own physiological function?
A year ago I asked if the cytosolic pH (pHcyto) was equal to the pH in the mitochondrial intermembrane space (pHims). One person responded, and we agreed that the outer mitochondrial membrane is no barrier to the movement of protons so pHcyto = pHims. But two people do not make a scientific consensus.
In recent decades it has become clear that the outer mitochondrial membrane has a vital role in programmed cell death, but this does not address its normal physiological role.
Questions for discussion:
- Is there a membrane potential across the OUTER mitochondrial membrane?
- Is the OMM VDAC channel ever closed?
- What transmembrane proteins (besides VDAC) reside in the OMM?
- Several enzymes localize to the cytosolic face of the OMM. Examples: MAO, KMO. Do you know of others? Is there a teleological reason for these localizations?
- What else does the OMM do?
Does a red blood cell have the minimum machinery to support attachment, entry, replication, and export of a +strand RNA virus like SARS-CoV-2?
Can you cite a reference?
Thank you.
As far as I know, the "reference" genome in a genome browser is usually the genome of a single sequenced individual.
I have often wondered if there is a special track in genome browsers consisting of the major allele at every position for that population (EAS, EUR, SAS, etc). It's true this track would change as more and more individuals are sequenced, but wouldn't such a track be widely useful?
When speaking of Mitchell's chemiosmotic hypothesis, it's common to consider the proton gradient as measured from the mitochondrial intermembrane space to the mitochondrial matrix. But textbooks say the outer mitochondrial membrane is freely permeable to molecules smaller than ~5kDa. Does this mean that the pH of the mitochondrial intermembrane space is equal to the cytosolic pH?
