Single particle tracking of acyl carrier protein (ACP)-tagged TrkA receptors in PC12nnr5 cells

Andrea Callegari; Stefano Luin; Laura Marchetti; Alessandro Duci; Antonino Cattaneo; Fabio Beltram

doi:10.1016/j.jneumeth.2011.10.019

Single particle tracking of acyl carrier protein (ACP)-tagged TrkA receptors in PC12nnr5 cells

J Neurosci Methods. 2012 Feb 15;204(1):82-86. doi: 10.1016/j.jneumeth.2011.10.019. Epub 2011 Oct 28.

Authors

Andrea Callegari¹, Stefano Luin², Laura Marchetti², Alessandro Duci³, Antonino Cattaneo³, Fabio Beltram⁴

Affiliations

¹ NEST Laboratory, Scuola Normale Superiore and Istituto Nanoscienze - CNR, Piazza San Silvestro 12, I-56126 Pisa, Italy; IIT@NEST, Center for Nanotechnology Innovation, Piazza San Silvestro 12, I-56126 Pisa, Italy; Scuola Normale Superiore, Piazza dei Cavalieri 7, I-56127 Pisa, Italy. Electronic address: a.callegari@sns.it.
² NEST Laboratory, Scuola Normale Superiore and Istituto Nanoscienze - CNR, Piazza San Silvestro 12, I-56126 Pisa, Italy; Scuola Normale Superiore, Piazza dei Cavalieri 7, I-56127 Pisa, Italy.
³ Scuola Normale Superiore, Piazza dei Cavalieri 7, I-56127 Pisa, Italy.
⁴ NEST Laboratory, Scuola Normale Superiore and Istituto Nanoscienze - CNR, Piazza San Silvestro 12, I-56126 Pisa, Italy; IIT@NEST, Center for Nanotechnology Innovation, Piazza San Silvestro 12, I-56126 Pisa, Italy; Scuola Normale Superiore, Piazza dei Cavalieri 7, I-56127 Pisa, Italy.

PMID: 22061422
DOI: 10.1016/j.jneumeth.2011.10.019

Abstract

There is a wide interest in studying the membrane mobility of Nerve Growth Factor (NGF) tropomyosin receptor kinase A (TrkA) at the single molecule level, in order to elucidate its diverse signaling responses related to different receptor functions. Here we present an experimental strategy based on the acyl carrier protein (ACP) tag in order to study the dynamics of the high-affinity NGF receptor TrkA in the membrane of PC12nnr5 cells. We present a single-particle tracking (SPT) study using highly photostable semiconductor quantum dots (Qdots) conjugated to ACP-tagged TrkA receptors. We demonstrate that ACP-TrkA shows biochemical and biological properties identical to those of its unmodified counterpart and that single receptor molecules in living cells display distinct diffusive regimes and a highly heterogeneous dynamics.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Acyl Carrier Protein / metabolism*
Animals
Cell Line, Tumor
Male
Microscopy, Fluorescence / methods*
Molecular Imaging / methods*
Molecular Probe Techniques*
PC12 Cells
Prostatic Neoplasms / metabolism*
Quantum Dots*
Rats
Receptor, trkA / metabolism*
Staining and Labeling

Substances

Acyl Carrier Protein
Receptor, trkA