The overall goal of this project is to understand the structural basis of CLCA1 activation and its role in airway biology in health and disease, in order to translate this knowledge into treatments for asthma and COPD, a current unmet need. CLCA1 is a potent modulator of calcium-activated chloride channels (CaCCs) and also a central mediator in mucous cell metaplasia, the process that leads to mucus overproduction. In this project we will investigate the structural and biochemical basis of CLCA activation of CaCCs, and investigate the role that CLCA1-mediated channel activation plays in mucous cell metaplasia. In our preliminary results, we demonstrate that CLCA proteins contain a consensus cleavage site that is recognized by a unique zinicin metalloprotease domain located within the N-terminus of CLCA itself. Furthermore, we show that this self- cleavage is required for hCLCA1 to activate CaCCs. These data suggest that CLCA1 is synthesized in a full- length """"""""inactive"""""""" form and that self-cleavage is required to produce an """"""""active"""""""" form of the protein. This project will focus on the structural and biochemical analysis of regulation of the metalloprotease domain activity, since it is necessary to produce the active form. We will then characterize the structura changes that occur upon activation by determining the structures of the full-length """"""""inactive"""""""" and """"""""active"""""""" forms of CLCA1. Finally, we will address the functional role of CLCA1 features in CaCC activation and the role of the channel in mucous cell metaplasia. Understanding how CLCA1 activity is regulated by its own metalloprotease domain and the downstream functional consequences of this regulation will facilitate the design of CLCA1 inhibitors for anti- mucus therapeutics.

Public Health Relevance

The protein CLCA1 is central to airway biology in health and disease as it is a potent modulator of calcium- activated chloride channels and mediates the overproduction of mucus, both through unknown mechanisms of action. The work outlined here will provide a detailed understanding of CLCA1 function in both of these processes, primarily using structural biology techniques. This information will be crucial for the development of CLCA1-trageting anti-mucus treatments for asthma and COPD.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
1R01HL119813-01
Application #
8563052
Study Section
Lung Cellular, Molecular, and Immunobiology Study Section (LCMI)
Program Officer
Banks-Schlegel, Susan P
Project Start
2013-08-01
Project End
2018-04-30
Budget Start
2013-08-01
Budget End
2014-04-30
Support Year
1
Fiscal Year
2013
Total Cost
$361,760
Indirect Cost
$123,760
Name
Washington University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Sala-Rabanal, Monica; Yurtsever, Zeynep; Berry, Kayla N et al. (2017) Modulation of TMEM16A channel activity by the von Willebrand factor type A (VWA) domain of the calcium-activated chloride channel regulator 1 (CLCA1). J Biol Chem 292:9164-9174
Kober, Daniel L; Brett, Tom J (2017) TREM2-Ligand Interactions in Health and Disease. J Mol Biol 429:1607-1629
Kober, Daniel L; Alexander-Brett, Jennifer M; Karch, Celeste M et al. (2016) Neurodegenerative disease mutations in TREM2 reveal a functional surface and distinct loss-of-function mechanisms. Elife 5:
Liu, Shenbin; Feng, Jing; Luo, Jialie et al. (2016) Eact, a small molecule activator of TMEM16A, activates TRPV1 and elicits pain- and itch-related behaviours. Br J Pharmacol 173:1208-18
Meyer, Peter A; Socias, Stephanie; Key, Jason et al. (2016) Data publication with the structural biology data grid supports live analysis. Nat Commun 7:10882
Yurtsever, Zeynep; Patel, Dhara A; Kober, Daniel L et al. (2016) First comprehensive structural and biophysical analysis of MAPK13 inhibitors targeting DFG-in and DFG-out binding modes. Biochim Biophys Acta 1860:2335-2344
Kober, Daniel L; Yurtsever, Zeynep; Brett, Thomas J (2015) Efficient Mammalian Cell Expression and Single-step Purification of Extracellular Glycoproteins for Crystallization. J Vis Exp :e53445
Sala-Rabanal, Monica; Yurtsever, Zeynep; Nichols, Colin G et al. (2015) Secreted CLCA1 modulates TMEM16A to activate Ca(2+)-dependent chloride currents in human cells. Elife 4:
Wu, Kangyun; Byers, Derek E; Jin, Xiaohua et al. (2015) TREM-2 promotes macrophage survival and lung disease after respiratory viral infection. J Exp Med 212:681-97
Zhang, Yong; Mao, Dailing; Roswit, William T et al. (2015) PARP9-DTX3L ubiquitin ligase targets host histone H2BJ and viral 3C protease to enhance interferon signaling and control viral infection. Nat Immunol 16:1215-27

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