- Signaling mechanisms of G protein-coupled receptors (GPCRs).
- The composition and stability of GPCR signaling complexes.
- Regulation of G protein signaling by RGS proteins.
GPCR signaling complexes in living cells
National Institutes of Health, R01GM078319, 10/07-9/11
Quantifying G-protein signaling complexes with FRAP
National Science Foundation, MCB060024, 8/06-7/09
Most of our efforts are devoted towards understanding the
timing of the molecular events and interactions that underlie G protein
signaling. One goal of this research is to provide kinetic information
to complement and "animate" the rapidly accumulating body of structural
information available for GPCRs, G proteins, effectors and regulators.
Our biophysical studies are generally done in living cells, and make use
of advanced imaging and spectroscopic methods such as fluorescence
recovery after photobleaching (FRAP) and bioluminescence resonance
In recent studies we used FRAP to study the association of
GPCRs with their cognate G proteins prior to receptor activation. Using
this method we have found that some (but not all) GPCRs "preassemble"
with their G proteins, and that this preassembly facilitates signaling.
Using a similar method we were able to show that, contrary to the
widely-accepted model, GPCRs do not always associate with each other as
stable dimers or higher-order oligomers. Using BRET we have shown that
GPCRs that associate with each other at the cell surface can dissociate
when they are internalized, thus demonstrating that endocytosis can
change the quaternary structure of GPCRs.
Graduate Student Supervision (as major advisor):
1996-2001 Huanmian Chen (Ph.D. 2001)
1999-2003 Jonathon Wetherington (Ph.D. 2003)
2002-2003 Chadwick Hales (Ph.D. 2003; co-advisor with J. Goldenring)
2001- Michael Clark
2002- Robert Lober
2004- Gregory Digby
1997 Caroline E. Rick, Ph.D., (visiting fellow, University of Chicago)
1997-2001 Xueyong Wang, Ph.D.
2000-2004 Rebecca Brogan, Ph.D.
Lambert, N.A., Johnston, C.A., Cappell, S.D.,
Kuravi, S., Willard, F.S. and Siderovski, D.P. Regulators of G-protein
Signaling accelerate GPCR signaling kinetics and govern sensitivity
solely by accelerating GTPase activity. Proc. Natl. Acad. Sci. USA,
107(15): 7066-7071, 2010.
Lambert, N.A. GPCR dimers fall apart. Science Signaling, 3(115):pe12, 2010.
Kuravi, S., Lan, T.-H., Barik, A. and Lambert, N.A.
Third-party BRET indicates constitutive association of membrane
proteins: application to class A GPCRs and G proteins. Biophysical
Journal, 98(10):2391-2399, 2010.
Fonseca, J.M. and Lambert, N.A. Instability of a class A GPCR oligomer interface. Molecular Pharmacology, 75(6):1296-9,
Hollins, B., Kuravi, S., Digby, G.J and Lambert, N.A. The c-terminus of GRK3 indicates rapid dissociation of G protein heterotrimers. Cellular
Signalling, 21(6):1015-1021, 2009.
Lambert, N.A. Uncoupling diffusion and binding in FRAP experiments. Nature Methods, 6:183, 2009.
Vilardaga, J.P., B�nemann, M., Feinstein, T.N., Lambert, N.,
Nikolaev, V.O., Engelhardt, S., Lohse, M.J., and Hoffmann, C. GPCR and G
proteins: drug efficacy and activation in live cells. Molecular
Endocrinology, 23(5):590-9, 2009.
Lambert, N.A. Dissociation of heterotrimeric G proteins in cells. Science Signaling, 1(25):re5,
Digby, G.J., Sethi, P.R. and Lambert, N.A. Differential dissociation of G protein heterotrimers. J. Physiol., 586(14):3325-3335,
Qin, K., Sethi, P.R. and Lambert, N.A.
Abundance and stability of complexes containing inactive G
protein-coupled receptors and G proteins. FASEB J., 22(8):2920-2927,
Clark, M.A., Sethi, P.R. and Lambert, N.A.
Active G?q subunits and M3 acetylcholine receptors promote distinct
modes of association of RGS2 with the plasma membrane. FEBS Letters,
Lober, R.M., Pereira, A.M. and Lambert, N.A.
Rapid activation of inwardly-rectifying potassium channels by immobile
G-protein coupled receptors. J. Neurosci., 26(48):12602-8, 2006.
Digby, G.J., Lober, R.M., Sethi, P.R. and Lambert, N.A. Some G protein heterotrimers physically dissociate in living cells. Proc. Natl.
Acad. Sci. USA, 103(47):17789-17794, 2006.
Clark, M.A. and Lambert, N.A. Endogenous RGS
Proteins Regulate the Kinetics of G?q/11-Mediated Modulation of Ion
Channels in CNS Neurons. Molecular Pharmacology, 69:1280-7, 2006.
Wetherington, J.P. and Lambert, N.A. GABAB
receptor activation desensitizes postsynaptic GABAB and A1 adenosine
responses in rat hippocampal neurons. J. Physiol., 544:459-467, 2002.
Smith, R.M., Baibakov, B., Lambert, N.A., and
Vogel, S.S. Low pH Inhibits Compensatory Endocytosis at a Step Between
Depolarization and Calcium Influx. Traffic, 3:397-406, 2002.
Wetherington, J.P. and Lambert, N.A.
Differential desensitization of responses mediated by presynaptic and
postsynaptic A1 adenosine receptors. J. Neurosci., 22:1248-55, 2002.
Chen, H. and Lambert, N.A. Endogenous RGS
proteins regulate presynaptic inhibition at rat hippocampal synapses.
Proc. Natl. Acad. Sci. USA, 97:12810-12815, 2000.
Smith, R.M., Baibakov, B., Ikebuchi, Y., White, B.H., Lambert, N.A.,
Kaczmarek, L.K. and Vogel S.S., Exocytotic insertion of calcium
channels constrains compensatory endocytosis to sites of exocytosis, J.
Cell Biol., 148:755-768, 2000.
Wang, X. and Lambert, N.A. GABAB receptors
couple to potassium and calcium channels on identified lateral perforant
pathway projection neurons, J. Neurophysiol., 83:1073-1078, 2000.
Vogel, S.S., Smith, R.M., Baibakov, B., Ikebuchi, Y. and Lambert, N.A. Calcium influx is required for endocytotic membrane retrieval, Proc. Natl. Acad.
Sci. USA, 96:5019-5024, 1999.
Chen, H. and Lambert, N.A. Inhibition of
dendritic calcium influx by activation of G-protein-coupled receptors in
the hippocampus, J. Neurophysiol., 78:3484 3488, 1997.
Lambert, N.A. and Wilson, W.A.
High-threshold Ca2+ currents in hippocampal interneurones and their
selective inhibition by activation of GABAB receptors. J. Physiol.,
Lambert, N. and Grover, L. The mechanism of biphasic GABA responses. Science, 269:928-929, 1995
Lambert, N.A. and Wilson, W.A. Heterogeneity
in presynaptic regulation of GABA release from hippocampal inhibitory
neurons. Neuron, 11:1057-1067, 1993.
Grover, L.M., Lambert, N.A., Schwartzkroin,
P.A. and Teyler, T.J. Role of HCO3- ions in depolarizing GABAA receptor
mediated responses in pyramidal cells of rat hippocampus. J.
Neurophysiol., 69:1541-1555, 1993.
Lambert, N.A., Borroni, A.M., Grover, L.M.
and Teyler, T.J. Hyperpolarizing and depolarizing GABAA
receptor-mediated dendritic inhibition in area CA1 of rat hippocampus.
J. Neurophysiol., 66:1538-1548, 1991.
Lambert, N.A. and Teyler, T.J. Adenosine
depresses excitatory but not fast inhibitory synaptic transmission in
area CA1 of the rat hippocampus. Neurosci. Lett., 122:50 52, 1991.