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Fraser J. Moss, PhD
View Curriculum Vitae (pdf)

Mailing Address:
Robbins E518
Phone: 216-368-5405
Fax: 216-368-5586

Research Interests

Molecular mechanisms sensing and regulating physiological pH

My research interests focus on understanding the molecular physiology of solute transporters, channels, and signaling proteins involved in acid-base regulation and gas transport.

In my research studying the function of sodium-coupled bicarbonate cotransporters (NCBTs), in their roles regulating intracellular and whole-body pH and transepithelial transport I determine the extracellular pH (pHo) dependence, kinetics, and substrate specificity (HCO3vs CO3=) of the NCBTs, e.g. NBCe1-A, and investigate which NCBT residues are responsible for imparting substrate specificity, electrogenicity, HCO3-independent Na conductance. I employ two-electrode voltage clamp electrophysiology to record the currents and ion-sensitive microelectrodes to record the intracellular pH (pHi) or sodium concentration in cells heterologously expressing NCBTs.  Out-of-equilibrium (OOE) solution perfusion exquisitely controls the extracellular environment of cells heterologously expressing NCBTs, allowing independent control of extracellular [CO2], [HCO3-] or pH.

My related acid-base physiology research focuses on the molecular processes underlying how the proximal tubule senses Δ[CO2]BL and Δ[HCO3-]BL and transduces these changes to modulate the rate of H+ secretion (JH) or bicarbonate reabsorption (JHCO3) during either respiratory or metabolic acidosis. Receptor protein tyrosine phosphatase γ (RPTPγ) is a major candidate for the CO2/HCO3- sensor. Changes in extracellular [CO2] and [HCO3-] result in changes in intracellular phosphatase activity. I use Förster resonance energy transfer (FRET) imaging to monitor the oligomerization or dissociation of RPTPγ as extracellular [CO2] or [HCO3-] changes in live cells, and also the how RPTPγ’s interactions with downstream signaling targets (e.g ErbB1 & ErbB2) changes as a consequence. Data recorded from samples exposed to in-equilibrium and OEE CO2/HCO3- solutions determines whether the highest RPTPγ phosphatase activity is activated by high CO2 or HCO3-, and which ligand is bound when RPTPγ is either a monomer or a dimer. The NCBTs are one of the major targets whose function is likely modulated by the activity of RPTPγ and its downstream signaling cascade.

I also study the movement of gas via protein channels (“gas channels”) across biological membranes. We developed the first assay for gas flux, based on maintaining the neutral buoyancy of a Xenopus oocyte previously injected with a 200-nl N2 bubble. During the neutral buoyancy assay (NBA) we apply pressure to the air above a saline solution, causing the bubble to constrict sufficiently that the oocyte falls to a depth of ~5 cm. As gas molecules dissolve in and diffuse through the cytoplasm, and eventually exit the cell, the bubble tends to collapse. A feedback system (camera, computer, digitally controlled pressure regulator) reacts by lowering the neutral-buoyancy air pressure (PNB) to maintain the oocyte at a 5-cm depth. The pressure inside the bubble (PBubble > PNB by a fixed amount) falls proportionally with the decreasing number of air molecules. The rate at which PNB falls thus reflects gas efflux from the bubble and can be converted into gas efflux in nmoles/s. We are also able to measure gas influx and can quantify N2, O2, or CO2 influx into an oocyte expressing NCBTs or other gas channel proteins (e.g. aquaporins or rhesus proteins) relative to control oocytes.

Specific Projects
  1. Molecular mechanism for sensing [CO2] and [HCO3] by RPTPγ

    HCO3 reabsorption (JHCO3) from renal proximal tubules (PT) is acutely regulated by basolateral [CO2] and [HCO3], not by extracellular pH (pHo). More recently we reported that the knockout of receptor protein tyrosine phosphatase γ (RPTPγ), normally present in the PT basal membrane, eliminates the CO2 and HCO3 sensitivities of JHCO3, as well as the normal defense to whole-body metabolic acidosis (MAc). The RPTPγ intracellular region has both a D1 phosphatase domain and a D2 blocking domain. When RPTPγ dimerizes, the D2 domain of one monomer blocks the D1 domain of the other. The extracellular region possess a carbonic anhydrase (CA) like domain (CALD) that is strikingly similar to classic CAs. However, the CALD lacks the amino acid residues believed necessary for CA activity. If the CALD is no longer capable of interconverting CO2 and HCO3, we hypothesize that it can sense CO2 or HCO3 and that the identity of the ligand bound to the CALD favors either dimerization or monomerization of the intracellular RPTPγ phosphatase domains. To detect the interaction of two RPTPγ monomers, we fuse the protein with the pH- and halide-insensitive GFP variants Aquamarine (Aq) to serve as a Förster resonance energy transfer (FRET) donor, and Citrine (Cit) to serve as a FRET acceptor, and coexpress the fusion proteins in HEK cells. We subject the cells to solutions representing different acid-base disturbances and acquire data from coexpressing cells, normalizing FRET for donor and acceptor expression levels (NFRET). We are able to vizualize changes in RPTPγ dimerization as reported by changes in the NFRET in response to each acid-base disturbance.  In parallel experiments, we co-transfect cells with RPTPγ-Aq and ErbB1-Cit (a candidate downstream target for RPTPγ) and are able to measure changes in NFRET in response to acid-base disturbances that are reciprocal to those measured for RPTPγ dimerization. We hypothesize that HCO3 and CO2 compete at the CALD to control RPTPγ dimerization state (and presumably phosphatase activity). We also hyothesize that changes in the RPTPγ dimerization state in response to [CO2]o or [HCO3]o but not pH, influence its interaction with downstream effector molecules including ErbB1.

  2. Is NBCe1 an exchanger rather than a cotransporter?

    I am recording the membrane potential (Vm), slope conductance and pHi from oocytes expressing NBCe1 that are exposed to different OOE solutions designed to probe the dependence of the transport direction on different extracellular ions and HCO3.

  3. Structural determinants of NBCe1 electrogenicity

    To determine the structural elements of NBCe1 that are essential for electrogenicity, with emphasis on the contribution of extracellular loop 4.

  4. The directional dependence of DIDS block of NCBTs

    Identifying and examining the roles of extracellular DIDS binding sites and determining the directional dependence of DIDS block in NBCe1 and other Na+-coupled HCO3 transporters.

  5. The role of carbonic anhydrase II on HCO3- -initiated transport through the SLC4A4 transporter NBCe1

    Testing the metabolon hypothesis by investigating what influence, if any, of soluble carbonic anhydrase II (CAII) or CAII fused to the C-terminus of NBCe1A has on transporter slope conductance.

Honors and Awards
  • 2021-Present   Co-Investigator R01 DK128315, NIH
  • 2019-Present   Co-Investigator Department of Defense—Air Force Research Laboratories, FA8650-19-C-6103
  • 2017-Present   Key personnel & co-author NIH R01 DK113197
  • 2016-Present   Key personnel on ONR grant N000141612535
  • 2014-2019        Key personnel on ONR grant N000141512060
  • 2013                    American Physiological Society Postdoctoral Travel Award IUPS 2013
  • 2005-2007        American Heart Association (Western States Affiliate) Postdoctoral Fellowship.
  • 2005-2009        Key personnel & co-authored NIH R01 HL079350
  • 2005-2007        Most Valuable Scientist –
  • 1998-2001        GlaxoSmithKline/Medical Research Council CASE Ph. D. Scholarship
  • US Patent           Moss FJ, CD Son, R Srinivasan & HA Lester. Methods and systems for detection of stoichiometry by Förster resonance energy transfer. 2014.
Featured Publications
  • Moss FJ & WF Boron. Carbonic anhydrase enhances activity of endogenous Na-H exchangers and not the Na+/HCO3 cotransporter, NBCe1, when expressed in Xenopus oocytes. J Physiol (Lond) 598:5821–5856, 2020. PMID: 32969493, PMCID: PMC7747792, doi: 10.1113/JP280143
  • Moss FJ, P Mahinthichaichan, D Lodowski, T. Kowatz, E Tajkhorshid, A Engel, WF Boron & A Vahedi-Faridi. Aquaporin-7: A dynamic aquaglyceroporin with higher water and glycerol transport capacity than its bacterial homolog GlpF. Frontiers in Physiology 11, 2020. PMID: 32695023, PMCID: PMC7339978 doi: 10.3389/fphys.2020.00728.
  • Zhao P, Geyer RR, Salameh AI, Wass AB, Taki S, Huffman DE, Meyerson HJ, Gros G, Occhipinti R, Moss FJ, Boron WF. Role of channels in the oxygen permeability of red blood cells. Version: 2. bioRxiv [preprint]. 2020 August; [revised 2020 September]: 2020.08.28.265066Available from: doi: 10.1101/2020.08.28.265066.
  • McHugh DR, CU Cotton, FJ Moss, M Vitko, DM Valerio, TJ Kelley, S Hao, A Jafri, ML Drumm, WF Boron, RC Stern, K McBennett & CA Hodges. Linaclotide improves gastrointestinal transit in cystic fibrosis mice by inhibiting sodium/hydrogen exchanger 3. American Journal of Physiology-Gastrointestinal and Liver Physiology 315:G868–G878, 2018. PMID: 30118317 doi: 10.1152/ajpgi.00261.2017
  • Michenkova M, S Taki, MC Blosser, HJ Hwang, T Kowatz, FJ Moss, R Occhipinti, X Qin, S Sen, E Shinn, D-K Wang, B Zeise, P Zhao, N Malmstadt, A Vahedi-Faridi, E Tajkhorshid & WF. Boron. CO2 Transport across membranes. Interface Focus, 11:2, 2021. PMID: 33633837, PMCID: PMC7898146, doi:
  • Moss FJ, B Zeise, D Huffman, S O’Neill & WF Boron. Quantitation of a neutral-buoyancy assay (NBA) to estimate transmembrane N2 flux. FASEB J 34, S1:1–1, 2020. doi: 10.1096/fasebj.2020.34.s1.06055.
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