Molecular mechanisms of the Cystic Fibrosis transmembrane conductance regulator (CFTR)

PV2

Diagram legend: Current record from excised, inside-out patch from oocyte expressing WT CFTR. Gating is regulated by protein kinase A (PKA) -mediated phosphorylation and by ATP. On the right, conductance levels corresponding to 0, 1, 2, or 3 simultaneously open channels are indicated.

Dr Paola Vergani
Lecturer Tel: 020 7679 7908
Email: p.vergani@ucl.ac.uk
Lab member: Dr Daniella Muallem

Dr Paola Vergani graduated in 1991 with a degree in Biological Sciences from the University of Pavia, Italy. She obtained her PhD from the University of Milan, studying transport mechanisms in yeast. After doing a first postdoctoral research period at Wye College, University of London, she moved to the Rockefeller University in New York where she started working on the structure/function of CFTR. In 2006 she joined the UCL Pharmacology Department as a lecturer.

Cystic fibrosis is the most common life-threatening inherited disease in the UK. We study the protein whose dysfunction causes cystic fibrosis, CFTR. It belongs to the superfamily of ABC proteins, which couple hydrolytic cycles at conserved nucleotide-binding domains (NBDs) to diverse cellular functions. CFTR is unique among ABC proteins in that its transmembrane domains comprise an ion channel. Opening and closing (gating) of the ion-permeation pathway is “remotely” controlled by ATP binding and hydrolysis at its NBDs . With our experiments we try to answer the question: how are conformational signals, originating at the catalytic site in the NBD, transmitted to the channel gate?

PV3

A CFTR topology: two NBDs (NBD1, green, and NBD2, blue), two TMDs, and the R-domain. B Crystal structure of an ATP-bound bacterial NBD homodimer (Chen et al., 2003 Molecular Cell 12: 651-661), and schematic representation of an NBD1/NBD2 “head-to-tail” dimer. C Proposed mechanism of ATP-dependent gating.

In alignments of homologous sequences one can discern sets of positions at which amino acid distribution has varied in a concerted way. This correlation in evolutionary space likely reflects conservation of a network of energetically coupled residues that mediates transmission of signals from one domain to another in individual ABC proteins. We use the evolutionary record to select targets for mutagenesis and, using patch-clamp techniques to record single channel activity, we follow how the energetic coupling between pairs of side-chains changes during the gating cycle. Since it is likely that ABC proteins share a common conformational coupling mechanism, we can use CFTR as a “model” ABC protein, exploiting the exceptional resolution afforded by single channel recording to study a basic mechanism shared with other, harder to study, ABC proteins.

Recent Publications:

  • P. Vergani, A. C. Nairn and D. C. Gadsby (2003). On the mechanism of MgATP-dependent gating of CFTR Cl- channels. J. Gen. Physiol., 121:17-36.
  • C. Basso, P. Vergani, A. C. Nairn and D. C. Gadsby (2003). Prolonged non-hydrolytic interaction of nucleotide with CFTR’s N-terminal nucleotide binding domain and its role in channel gating. J. Gen. Physiol., 122:333-48
  • P. Vergani, S. W. Lockless, A. C. Nairn and D. C. Gadsby. (2005) CFTR channel opening by ATP-driven tight dimerization of its nucleotide binding domains. Nature, 433:876-880.
  • P. Vergani, C. Basso, M. Mense, A. C. Nairn, D. C. Gadsby (2005). Control of the CFTR channel’s gates. Biochemical Society Transactions, 33(Pt 5):1003-7.
  • D. C. Gadsby, P. Vergani, and L. Csanády (2006). The ABC protein turned chloride channel whose failure causes cystic fibrosis. Nature, 440:477-83.
  • M. Mense, P. Vergani, D.M. White, G. Altenberg, A. C. Nairn, D. C. Gadsby (2006). In vivo phosphorylation of CFTR promotes formation of a nucleotide-binding domain heterodimer. EMBO J., 25:4728-39