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Biophys. J. BioFAST: First Published May 4, 2007. doi:10.1529/biophysj.106.102244
© 2007 by the Biophysical Society.

A more recent version of this article appeared on August 1, 2007.
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CELL BIOPHYSICS

SINGLE PARTICLE TRACKING OF MEMBRANE PROTEIN DIFFUSION IN A POTENTIAL: SIMULATION, DETECTION, AND APPLICATION TO CONFINED DIFFUSION OF CFTR Cl- CHANNELS

Songwan Jin 1, Peter M. Haggie 2 and Alan S. Verkman 2*

1 University of California-San Francisco
2 Univ. of California - San Francisco

* To whom correspondence should be addressed. E-mail: verkman{at}itsa.ucsf.edu.

Submitted on December 1, 2006
Revised on February 1, 2007
Accepted on 26 March 2007


  Abstract
Confined diffusion of membrane receptors and lipids can result from intramembrane barriers, skeletal interactions, rafts, and other phenomena. We simulated single particle diffusion in 2-dimensions in an arbitrary potential, V(r), based on summation of random and potential gradient-driven motions. Algorithms were applied and verified for detection of potential-driven diffusion, and for determination of V(r) from radial particle density distributions, taking into account experimental uncertainties in particle position and finite trajectory recording. SPT analysis of the diffusion of cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channels in mammalian cells revealed confined diffusion with diffusion coefficient, ~ 0.004 µm2/s. SPT data fitted closely to a spring-like attractive potential, V(r) = kr2, but not to other V(r) forms such as hard-wall or viscoelastic-like potentials. The 'spring constant' k, determined from SPT data was 2.6±0.8 pN/µm, and not altered significantly by modulation of skeletal protein architecture by jasplakinolide. However, k was reduced by a low concentration of latrunculin, supporting the involvement of actin in the spring-like tethering of CFTR. Confined diffusion of membrane proteins is likely a general phenomenon suitable for non-invasive V(r) analysis of force-producing mechanisms. Our data provide first measurement of actin elasticity that does not involve application of an external force.

Key Words: CFTR, SPT, actin, cell cytoskeleton, cystic fibrosis




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Copyright © 2007 by the Biophysical Society.