Martina Gentzsch, PhD

Associate Professor

Specialty Areas:
Processing, routing and regulation of ion channels.
Personalized therapeutics for rescue of mutant CFTR.
Preclinical assays for evaluation of drugs targeting CFTR, ENaC, and CaCC.
 
Chronology:
Ph.D. Summa Cum Laude: Cell Biology, University of Regensburg, Germany, 1997
Postdoctoral Fellow: Mayo Clinic, Scottsdale, AZ, 1998-2001
Research Associate: Mayo Clinic College of Medicine, Scottsdale, AZ, 2001-2005
Assistant Professor of Cell and Developmental Biology, UNC, 2005-2012
Director, Correction Core: Cystic Fibrosis Research Center, UNC, 2012-2015
Assistant Professor of Cell Biology and Physiology: UNC, 2012-present
Director, Pre-Clinical Core: UNC, 2015-present
Director, CFTR Functional Analysis Core: UNC, 2015-present
 
Research Focus:

A major goal of our studies is to assess potential targets that are of clinical interest for cystic fibrosis (CF) therapy. In particular, our research focuses on CFTR processing, intracellular trafficking, and rescue. We have developed reliable methodology to track apical stability of mutant CFTR in physiologically relevant primary human bronchial epithelial (HBE) and human nasal epithelial (HNE) cultures. We are applying these techniques and electrophysiological assays to evaluate CFTR rescue by small-molecule compounds in primary and conditionally reprogrammed epithelial nasal, bronchial, and intestinal cultures and tissues. Furthermore, we have developed and utilize state-of-the-art pre-clinical assays to investigate the efficacy of novel therapeutic approaches to CF in relevant in vitro, ex vivo, and in vivo models of CF including organoids from different tissues.

In summary, we are designing and optimizing bio-assays for personalized CFTR-targeting therapeutics and are assessing the effects of novel therapeutic approaches to CF in relevant models of CF that we previously established. Currently we are investigating 1) means to rescue rare CFTR mutations, 2) mechanistic details on how environmental factors such as disease status, bacterial infection, carbon dioxide levels, pH, and lipid composition affect CFTR rescue, 3) the consequences of CFTR rescue on mucociliary clearance in CF airways, and 4) diverse roles of CFTR in multiple organs with particular emphasis on assessing differences in CFTR modulator action in cells from various tissues.

Research Methods:

  • Analysis of processing, stability, and trafficking of ion channels, including CFTR and ENaC, by biochemical methods i.e. Western Blot, Immunoprecipitation, Surface Biotinylation, Pulse-Chase Experiments, On-Cell Westerns
  • Evaluation of pharmacological rescue of mutant CFTR in primary and conditionally reprogrammed human nasal, bronchial, and intestinal cultures
  • Analysis of electrophysiological properties of cell cultures and tissues
  • Sphere assays of ion channel function
  • 2D and 3D cultures of primary and conditionally reprogrammed human nasal epithelial and bronchial epithelial cells
  • Protein analysis of nasal, bronchial, and intestinal tissue
  • Analysis of protein glycosylation and secretion
  • Endocytosis and recycling assays
  • Transient transfection and generation of stable cell lines
  • Confocal immunofluorescence and live-cell microscopy
  • Viral gene delivery and protein expression
  • Silencing (retroviral and lentiviral vectors)
  • Sample preparation for mass spectrometry-based proteomics
  • Proteolysis of ENaC in the Xenopus oocyte model
Rescue of CFTR. The mutant CFTR protein, F508del, is retained at the ER in differentiated primary human bronchial epithelial (HBE) cells.
Rescue of CFTR. The mutant CFTR protein, F508del, is retained at the ER in differentiated primary human bronchial epithelial (HBE) cells. Corrector compounds promote transfer of mutant CFTR from the ER to the apical membrane, whereas potentiator compounds increase activity of CFTR at the apical surface. The insets show confocal immunofluorescence of CFTR (green) expressed in primary HBE cells. F508del CFTR is retained within the cell (bottom panel) while wild-type CFTR is at the cell surface (top panel). Cilia are stained in red and nuclei in blue.
Correction of mutant CFTR in CF HBE cells (P67L/∆F508).
Correction of mutant CFTR in CF HBE cells (P67L/∆F508). Short-circuit currents were measured in Ussing chambers. Amiloride was added to inhibit ENaC, forskolin to stimulate CFTR, VX-770 to potentiate CFTR, and CFTR inhibitor-172 to inhibit CFTR. The blue stripe, treatment with VX-809 (5 µM) + VX-770 (1 µM), resulted in the highest level of CFTR function for this particular mutant.
CFTR impedes proteolysis of ENaC.
CFTR impedes proteolysis of ENaC. Expression of increasing amounts of CFTR (shown in red) diminishes appearance of matriptase-induced proteolytic fragments of α-ENaC (shown in green). CFTR, ENaC, and matriptase were co-expressed in Xenopus oocytes. Molecular weight markers (250, 150, 100, 75 and 50 kDa) are shown in yellow.
Virally delivered expression of CFTR in epithelial cells.
Virally delivered expression of CFTR in epithelial cells. Wild-type CFTR resides at the apical membrane in primary HBE cells. Cilia are stained in blue, virally delivered CFTR-expressing cells in green, and CFTR is shown in red.
Human primary nasal cells grown as 3D sphere cultures.
Human primary nasal cells grown as 3D sphere cultures. Nasal epithelial cells obtained from individuals can be cultured into 3D spheres and used for studying ion channel function. Apical membrane and cilia are stained in red, cells in green, and nuclei in blue.
Human primary bronchial sphere grown in matrigel.
Human primary bronchial sphere grown in matrigel. Bronchial cells obtained from individuals can be cultured into 3D spheres for studying ion channel function. When grown in matrigel, the apical surface faces the inside of the sphere. This section is stained with AB PAS. Bar=50µm.

Collaborations:

Dawn Bowles, PhD (Duke University, Surgery):
CFTR function in cardiomyocytes.

Nikolay V. Dokholyan, PhD (Department of Biochemistry and Physics, UNC):
Structure of ENaC and CFTR.

Doug Cyr, Ph.D. (Cell Biology and Physiology, UNC):
Rescue of rare CFTR Mutations.

Camille Ehre, PhD (Department of Pediatrics, UNC):
CFTR rescue, mucin production.

Charles Esther, MD, PhD (Department of Pediatric Pulmonology, UNC):
CFTR-targeting drug concentrations in cultures and tissues.

M. Gregory Forest, PhD (Mathematics and Biomedical Engineering, UNC):
Human nasal epithelial organoids for predicting effectiveness of CFTR modulators.

Barbara Grubb, PhD (Department of Medicine, UNC):
Measurement of ion transport in mouse models.

Jennifer Guimbellot, MD, PhD (Department of Pediatrics, UAB):
Nasosphere assays to measure of CFTR function.

Michael Knowles, MD (Department of Medicine, UNC):
Genetic modifiers of CFTR function.

Silvia Kreda (Department of Medicine, UNC):
CFTR rescue and mucin secretion

Scott Magness, PhD (Departments of Medicine, Biomedical Engineering, Cell Biology and Physiology, UNC):
Ion channel properties of intestinal 2D and 3D cultures.

Marianne Muhlebach, MD (Department of Pediatric Pulmonology, UNC)
Collection of nasal epithelial tissue to generate 2D and 3D culture models.

Sean Murphy, PhD (Wake Forest Institute for Regenerative Medicine):
Airway tissue engineering.

Wanda O’Neal, PhD (Department of Medicine, UNC):
Viral vectors for expression and knock down of CFTR and ENaC.

Lawrence Ostrowski, PhD (Department of Medicine, UNC):
CFTR rescue and mucocilary clearance.

Scott H. Randell, PhD (Departments of Medicine and Cell Biology and Physiology, UNC):
Properties of primary and conditionally reprogrammed bronchial 2D and 3D cultures.

Carla Ribeiro (Department of Medicine, UNC):
Inflammation and CFTR rescue.

John R. Riordan, PhD (Department of Biochemistry and Physics, UNC):
CFTR: structure, function and processing.

Robert Tarran, PhD (Department of Cell Biology and Physiology, UNC):
Modulation of CFTR trafficking. Regulation of ENaC function.

Monroe J. Stutts, PhD (Department of Medicine, UNC):
ENaC-CFTR interaction and regulation of ENaC .

 

Selected Bibliography:

Armirotti A, Tomati V, Matthes E, Veit G, Cholon DM, Phuan PW, Braccia C, Guidone D, Gentzsch M, Lukacs GL, Verkman AS, Galietta LJV, Hanrahan JW, Pedemonte N (2109) Bioactive Thymosin Alpha-1 Does Not Influence F508del-CFTR Maturation and Activity. Sci Rep. 16;9(1):10310.

Tomati V, Caci E, Ferrera L, Pesce E, Sondo E, Cholon DM, Quinney NL, Boyles SE, Armirotti A, Ravazzolo R, Galietta LJ, Gentzsch M, Pedemonte N (2019) Thymosin α-1 does not correct F508del-CFTR in cystic fibrosis airway epithelia. JCI Insight. 4(7).

Clancy JP, Cotton CU, Donaldson SH, Solomon GM, VanDevanter DR, Boyle MP, Gentzsch M, Nick JA, Illek B, Wallenburg JC, Sorscher EJ, Amaral MD, Beekman JM, Naren AP, Bridges RJ, Thomas PJ, Cutting G, Rowe S, Durmowicz AG, Mense M, Boeck KD, Skach W, Penland C, Joseloff E, Bihler H, Mahoney J, Borowitz D, Tuggle KL (2019) CFTR modulator theratyping: Current status, gaps and future directions. J Cyst Fibros. 18(1), 22-34.

Gentzsch M, Cholon DM, Quinney NL, Boyles SE, Martino MEB,  Ribeiro CMP (2018) The cystic fibrosis airway milieu enhances rescue of F508del in a pre-clinical model. Eur Respir J. 52(6).

Moorefield EC, Blue RE, Quinney NL, Gentzsch M, Ding S (2018) Generation of renewable mouse intestinal epithelial cell monolayers and organoids for functional analyses. BMC Cell Biol. 19(1), 15.

Kota P, Gentzsch M, Dang Y, Boucher R, Stutts M (2018) The N-terminus of alpha-ENaC mediates ENaC cleavage and activation by furin. J Gen Physiol. 150(8), 1179-1187.

Gentzsch M, Mall MA (2018) Ion channel modulators in cystic fibrosis. Chest. 154(2), 383-393. Review.

Cholon DM, Gentzsch M (2018) Recent progress in translational cystic fibrosis research using precision medicine strategies. J Cyst Fibros. 17(2S), S52-S60.

Tomati V, Caci E, Ferrera L, Pesce E, Sondo E, Cholon DM, Quinney NL, Boyles SE, Armirotti A, Ravazzolo R, Galietta LJ, Gentzsch M, Pedemonte N (2018) Thymosin α-1 does not correct F508del-CFTR in cystic fibrosis airway epithelia. JCI Insight. 3(3).

Guimbellot JS, Leach JM, Chaudhry IG, Quinney NL, Boyles SE, Chua M, Aban I, Jaspers I, Gentzsch M (2017) Nasospheroids permit measurements of CFTR-dependent fluid transport. JCI Insight. 2(22).

Gentzsch M, Boyles SE, Cheluvaraju C, Chaudhry IG, Quinney NL, Cho C, Dang H, Liu X, Schlegel R, Randell SH (2017) Pharmacological rescue of conditionally reprogrammed cystic fibrosis bronchial epithelial cells. Am J Respir Cell Mol Biol. 56(5), 568-574.

Gentzsch M, Ren HY, Houck SA, Quinney NL, Cholon DM, Sopha P, Chaudhry IG, Das J, Dokholyan NV, Randell SH, Cyr DM (2016) Restoration of R117H CFTR folding and function in human airway cells through combination treatment with VX-809 and VX-770. Am J Physiol Lung Cell Mol Physiol. 311(3), L550-9.

Watson MJ, Lee SL, Marklew AJ, Gilmore RC, Gentzsch M, Sassano MF, Gray MA, Tarran R (2016) The cystic fibrosis transmembrane conductance regulator (CFTR) uses its C-terminus to regulate the A2B adenosine receptor. Sci Rep. 6, 27390.

Cholon DM, Esther CR Jr, Gentzsch M (2016) Efficacy of lumacaftor-ivacaftor for the treatment of cystic fibrosis patients homozygous for the F508del-CFTR mutation. Expert Rev Precis Med Drug Dev. 1(3), 235-243.

Cholon DM, Quinney NL, Fulcher ML, Esther CR, Das J, Dokholyan NV, Randell SH, Boucher RC, Gentzsch M (2014) Potentiator ivacaftor abrogates pharmacological correction of ∆F508 CFTR in cystic fibrosis. Sci Transl Med. 6, 246ra96.

Kota P, Buchner G, Chakraborty H, Dang YL, He H, Garcia GJM, Kubelka J, Gentzsch M, Stutts MJ, Dokholyan NV (2014) The N-terminal domain allosterically regulates cleavage and activation of the epithelial sodium channel. J Biol Chem. 289, 23029-42.

Aleksandrov AA, Kota P, Cui L, Jensen T, Alekseev AE, Reyes S, He L, Gentzsch M, Aleksandrov LA, Dokholyan NV, Riordan JR (2012) Allosteric modulation balances thermodynamic stability and restores function of ∆F508 CFTR. J Mol Biol. 419, 41-60.

Kota P, García-Caballero A, Dang H, Gentzsch M, Stutts MJ, Dokholyan NV (2012) Energetic and structural basis for activation of the epithelial sodium channel by matriptase. Biochemistry 51, 3460−9.

Clunes LA, Davies CM, Coakley RD, Aleksandrov AA, Henderson AG, Zeman KL, Worthington EN, Gentzsch M, Kreda SM, Cholon D, Bennett WD, Riordan JR, Boucher RC, Tarran R (2012) Cigarette smoke exposure induces CFTR internalization and insolubility, leading to airway surface liquid dehydration. FASEB J. 26, 533-45.

Kreda SM, Gentzsch M (2011) Imaging CFTR protein localization in cultured cells and tissues. In Cystic fibrosis diagnosis and protocols. Vol. 742 of Methods of Molecular Biology. Amaral M, Kunzelmann K (eds). pp. 15-33. Humana Press.

Johnson JS, Gentzsch M, Zhang L, Ribeiro CM, Kantor B, Kafri T, Pickles RJ, Samulski RJ (2011) AAV exploits subcellular stress associated with inflammation, endoplasmic reticulum expansion, and misfolded proteins in models of cystic fibrosis. PLoS Pathog. 7, e1002053.

Hegedus T, Gentzsch M, Riordan JR (2011) CFTR: Understanding the cause and influencing the outcome of a major genetic disease. In: The ABC transporters of human physiology and disease. Chapter 10. Linton K, Holland B (eds). pp. 269-309. World Scientific Publishing.

Cholon DM, O’Neal WK, Randell SH, Riordan JR, Gentzsch M (2010) Apical stability and endocytic trafficking of wild-type and mutant CFTR in primary cultures of human airway epithelia. Am J Physiol Lung Cell Mol Physiol. 298, L304-14.

Gentzsch M, Dang H, Dang Y, Garcia-Caballero A, Suchindran H, Boucher RC, Stutts MJ (2010) The cystic fibrosis transmembrane conductance regulator impedes proteolytic stimulation of the epithelial Na+ channel. J Biol Chem. 285, 32227-32. “Paper of the Week”

Aleksandrov AA, Kota P, Aleksandrov L, He L, Jensen T, Cui L, Gentzsch M, Dokholyan NV, Riordan JR (2010) Regulatory insertion removal restores maturation, stability and function of ∆F508 CFTR. J Mol Biol. 401, 194-210.

Young A, Gentzsch M, Jia Y, Abban CY, Meneses PI, Bridges RJ, Bradbury NA (2009) Dynasore inhibits removal of wild-type and ∆F508 CFTR from the plasma membrane. Biochem J. 421, 377-85.

Hutt DM, Herman, DM, Rodrigues, APC, Noel S, Pilewski JM, Matteson J, Hoch B, Kellner W, Kelly JW, Richarson J, Thomas PJ, Matsumura Y, Skach WR, Gentzsch M, Riordan JR, Sorscher EJ, Okiyonad T, Lukacs GL, Frizzell RA, Manning G, Gottesfeld JM, Balch WE (2009) Reduced histone deacetylase 7 activity restores function to misfolded CFTR in cystic fibrosis. Nat Chem Biol. 6, 25-33.

Fulcher ML, Gabriel SE, Olsen JC, Tatreau JR, Gentzsch M, Livanos E, Saavedra MT, Salmon P, Randell SH (2009) Novel human bronchial epithelial cell lines for cystic fibrosis research. Am J Physiol Lung Cell Mol Physiol. 29, L82-91.

Chang XB, Mengos A, Hou YX, Cui L, Jensen TJ, Aleskandrov A, Riordan JR, Gentzsch M (2008) Role of N‑linked oligosaccharides in the biosynthetic processing of CFTR. J Cell Sci. 121, 2814-23.

Gentzsch M, Choudhury A, Chang X-B, Pagano R, Riordan JR (2007) Misassembled mutant ∆F508 CFTR in the distal secretory pathway alters cellular lipid trafficking. J Cell Sci. 120, 447-55.

Thelin, WR, Chen, Y, Gentzsch M, Kreda S, Sallee J, Scarlett C, Borchers C, Jacobson, K, Stutts KJ, Milgram S (2007) A direct interaction with filamins modulates the stability and plasma membrane expression of CFTR. J Clin Invest. 117, 364-74.

Cui L, Aleksandrov L, Chang X-B, Hou YX, He L, Hegedus T, Gentzsch M, Aleksandrov A, Balch WE, Riordan JR (2007) Domain interdependence in the biosynthetic assembly of CFTR. J Mol Biol. 365, 981-94.

Grubb BR, Gabriel SE, Mengos A, Gentzsch M, Randell SH, Van Heeckeren AM, Knowles MR, Drumm ML, Riordan JR, Boucher RC (2006) SERCA pump inhibitors do not correct biosynthetic arrest of DF508 CFTR in cystic fibrosis. Am J. Respir Cell Mol Biol. 34, 355-63.

Hegedus T, Aleksandrov A, Cui L, Gentzsch M, Chang X-B, Riordan JR (2006) ∆F508 CFTR with two altered RXR motifs escapes from ER quality control but its channel activity is thermally sensitive. Biochim Biophys Acta. 1758, 565-72.

Gentzsch M, Chang X-B, Cui L, Wu Y, Ozols VV, Choudhury AK, Pagano RE, Riordan JR (2004) Endocytic trafficking routes of wild-type and DF508 CFTR. Mol Biol Cell. 15, 2684-96.

Gentzsch M, Cui L, Mengos A, Chang XB, Chen JH, Riordan JR (2003) The PDZ-binding chloride channel ClC-3B localizes to the Golgi and associates with cystic fibrosis transmembrane conductance regulator-interacting PDZ proteins. J Biol Chem. 278, 6440-9.

Gentzsch M, Aleksandrov A, Aleksandrov L, Riordan JR (2002) Functional analysis of the C‑terminal boundary of the second nucleotide binding domain of the cystic fibrosis transmembrane conductance regulator and structural implications. Biochem J. 366, 541-8.

Gentzsch M, Riordan JR (2001) Localization of sequences within the C-terminal domain of CFTR which impact maturation and stability. J Biol Chem. 276, 1291-8.

Complete Bibliography:

http://www.ncbi.nlm.nih.gov/pubmed?term=Gentzsch%20M%5BAuthor%5D&cmd=DetailsSearch

Martina Gentzsch