Pablo Artigas, Ph.D.
Associate Professor of Cell Physiology & Molecular Biophysics
Department of Cell Physiology and Molecular Biophysics
Texas Tech University Health Sciences Center
3601 4th Street, STOP 6551
Lubbock, Texas 79430
Phone: (806) 743-1142
Lab: (806) 743-3170
FAX: (806) 743-1512
Our research focuses on understanding the function, mechanisms and pharmacology of the proteins that transport ions across membranes. They are essential for the electrical signaling in the cardiovascular and nervous systems, and as such, they represent the target of several pharmacological agents used for the treatment of a number of cardiovascular and neural diseases.
In particular, we are focusing our efforts on understanding how the Na/K and H/K pumps operate at the molecular level, as well as their role in health and disease. These two pumps form the P-type IIC ATPase family. As indicated by their names, the Na/K pump couples ATP hydrolysis to extrusion of three Na+ ions in exchange for two K+ ions, against each ion’s electrochemical gradient, while the H/K pump hydrolyses ATP to export H+ and import K+. Both pump types are formed by association of a catalytic α- and an auxiliary β-subunit, which are frequently associated with a FXYD regulatory subunit, as shown in the figure.
These subunits present tissue specific distribution. Four genes encode four Na/K pump α subunits and three genes encode three different Na/K pump β subunits. There are two genes encoding H/K pump’s α subunits; the α-subunit expressed in the stomach forms the gastric H/K pump when associated to the specific gastric β subunit, while the α-subunit expressed in the apical membrane of other epithelia forms the non-gastric (aka colonic) H/K pump by association with the β1 subunit of the Na/K pump.
The gastric H/K pump is responsible for acidification of the stomach and is the target of the widely used antacid drug, omeprazole. The non-gastric H/K pump is involved in K+ reabsorption and recently has been implicated in the development of infections in cystic fibrosis patients. The Na/K pump is present in nearly all animal cells. Na/K pumps formed by the α1 subunit are expressed in all cells, while pumps formed by the α2 subunit are specific to muscle (all three types) and glia, and α3 pumps are expressed in neurons and in the heart. Mutations to all three α subunits are known to produce several types of illnesses, including Conn’s syndrome (primary aldosteronism) and Charcot-Marie-Tooth disease type 2 (α1), migraine (α2), and alternating hemiplegia of childhood and some forms of Parkinsonism (α3).
We use an array of techniques that include electrophysiology (patch-clamp, two-electrode voltage clamp and cut-open oocyte), molecular biology (transcriptome analysis, site-directed mutagenesis, non-sense suppression, heterologous protein expression and western blot), biochemistry (chemical modification, measurement of ATPase activity, radiolabeled ion uptake) and fluorescence microscopy (voltage clamp fluorometry, immunocytochemistry).
We are thankful to all agencies (federal and private) that have supported our research.
Active: National Science Foundation, American Heart Association (Predoctoral Fellowship to
Past: South Plains Foundation, American Heart Association.
Associate Professor, Department of Cell Physiology and Molecular Biophysics and CMPR, TTUHSC
Associate Professor, Department of Molecular Physiology and Biophysics, University of Iowa
Professor, Director, School of Biological Sciences, Illinois State University
Professor, Department of Biochemistry and Molecular Biophsics, University of Chicago
Professor, Department of Biomedicine, Aahrus University, Denmark
DECIPHERING THE DETERMINANTS OF IONIC SELECTIVITY AND PHARMACOLOGY OF THE NA/K AND H/K PUMPS
From the basic science point of view, it remains unclear how the Na/K pump selects Na+ over the more concentrated K+ when facing intracellularly or how it selects K+ over the more concentrated Na+ when facing extracellularly. It is also unclear what determines the selectivity of Na+ or H+ which make the Na/K and the H/K pumps different. We (Holm et al, 2017) have recently uncovered a single residue that may be responsible for the 3Na+/2K+ stoichiometry that determines the electrogenicity of the pump. We are currently using a combination of regular mutagenesis, molecular dynamics simulations (in collaboration with Benoît Roux) and incorporation of unnatural amino acids (through a process called non-sense suppression) to untangle the mechanisms of ion selectivity and stoichiometry at the atomic level. From a translational science point of view, we are trying to develop new drugs to selectively inhibit or modulate different types of pumps in order to treat disease.
ELUCIDATING THE PHYSIOLOGICAL ROLE OF FXYD PROTEINS
During the early 2000s, it was demonstrated that a group of proteins that have a single transmembrane segment regulate the function of the Na/K pump. Despite efforts by several groups, the function of each of these regulators remains unknown. For instance, in heart cells, FXYD1 is the most phosphorylated plasma membrane protein in response to adrenaline. Yet, the mice in which FXYD1 has been “knocked out” survive without major apparent phenotypes and problems. This illustrates that some of the important functions of this (the most studied FXYD) and the other six FXYD proteins in the human body are poorly understood. We are using electrophysiology, co-immunoprecipitation, and Western blotting to study the function of these proteins in different tissues, including the the adrenal gland, cardiovascular and central nervous system.
DETERMINING THE PATHOPHYSIOLOGICAL MECHANISMS OF NA/K PUMP MUTATIONS THAT INDUCE ILLNESS
We aim to understand the functional consequences of mutations that affect the Na/K pump α1 subunit to decipher how these mutations trigger pathophysiological effects. We have recently demonstrated (Meyer et al, 2017) that most Na/K pump mutations found in aldosterone-producing adenomas (benign tumors of the adrenal gland) from patients with primary aldosteronism (Conn’s syndrome) lead to a loss-of-function instead of the previously proposed gain-of-function. We are evaluating other illness-inducing mutations in this important α subunit, as well as the importance of the function of different Na/K pump types (α1β1, α2β2, etc.) in the adrenal gland and other tissues. For this project, we use heterologous expression and patch clamp electrophysiology of Xenopus oocytes, adrenal glands, and other tissue slices.
INVESTIGATING THE ROLE OF NA/K PUMPS WITH SPECIAL MODIFICATIONS IN THE ADAPTATION OF ANIMALS TO DIFFERENT ENVIRONMENTS
The brine shrimp (crustaceans of the order Anostraca, genus Artemia) is one of only two animals known to live in the Great Salt Lake of Utah at ~ 14% salinity (about four times that of seawater). Brine shrimp survive in salinities up to 10 times of that in normal sea water. It is known that they have two α subunits for the Na/K pump. One of these α subunits has several modifications around the ion-binding sites, which make it very different to most Na/K pumps found in other animals. Teleost fish present similar modifications, but when adapting to fresh water rather than high salinity. Our laboratory is trying to uncover the reasons for these special Na/K pump modifications, and what adaptive advantage, if any, is gained by the animals that carry them. The importance of other ion-transport proteins in adaptation is also being investigated. In this project, we use transcriptome analysis and several other molecular biology techniques, immunostaining, radioactive isotope uptake, heterologous protein expression in oocytes, and electrophysiology.
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Pablo Artigas, Ph.D.
(806) 743 - 1142