Properties of single drosophila trpl channels expressed in sf9
insect cells.
Kunze, Diana L., William G. Sinkins, Luis Vaca, and William P.
Schilling.
Rammelkamp Center for Education and Research, MetroHealth Campus,
Case Western Reserve University, Cleveland OH 44109-1998, Instituto
de Fisiologia Celular, Universidad Nacional Autonoma de Mexico,
Ciudad Universitaria, Mexicao, D.F. 04510
APStracts 3:0245C, 1996.
The trp-like (trpl) gene is thought to encode an ion channel important
for signal transduction in Drosophila photoreceptor cells. Consistent
with this hypothesis, heterologous expression of the trpl-encoded
protein (Trpl) is associated with the appearance of an outwardly
-rectifying, non-selective cation current. In the present study,
single channels were recorded in cell-attached, inside-out, and
outside-out membrane patches from Sf9 insect cells infected with
recombinant baculovirus containing trpl cDNA under control of the
polyhedrin promoter. The single channel current-voltage relationship
was linear from -100 to +80 mV with a slope conductance of 89-110 pS.
However, the probability of opening, P(o), was voltage-sensitive,
increasing at positive potentials contributing to the outwardly
rectifying properties of the whole-cell currents. The single channels
were 1) never observed in Sf9 cells infected with recombinant
baculovirus containing the B2 bradykinin receptor cDNA or in non
-infected Sf9 cells, 2) appear at the same time post-infection as the
Trpl whole-cell current, 3) were non-selective with respect to Na+,
Ca2+, and Ba2+, 4) were blocked by 1-2 mM La3+ and Gd3+ (but not 10
(M), and 5) were blocked by 4-8 mM Mg2+. The single Trpl channel
activity increased spontaneously with time after patch formation and
the activity was further increase by application of bradykinin to
cells expressing both the B2 bradykinin receptor and the Trpl
protein. These results suggest that this single channel activity
reflects expression of the Trpl protein and provides conclusive
evidence that trpl encodes a non-selective cation channel consistent
with its proposed role in Drosophila phototransduction.
Received 20 May 1996; accepted in final form 19 July 1996.
APS Manuscript Number C275-6.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 21 August 1996