Changes in G protein pattern and in G protein-dependent signaling during
erythropoietin- and dimethylsulfoxide-induced differentiation of murine
erythroleukemia cells
F Kesselring, K Spicher and H Porzig
Department of Pharmacology, Universitat Bern, Switzerland.
We have studied the expression of G protein subtypes and the role of G
protein-dependent signaling in two subclones of RED-1 cells, an
erythropoetin(Epo)-sensitive, murine erythroleukemia cell line. Clone 6C8
showed terminal erythroid differentiation in response to a combined
treatment with Epo and dimethylsulfoxide. Clone G3 was resistant to these
inducers, but responded to Epo with enhanced proliferation. We measured G
protein alpha subunit levels by toxin-catalyzed adenosine diphosphate
(ADP)-ribosylation with [32P]-nicotinamide adenine dinucleotide (NAD) and
by semiquantitative immunoblotting with specific antisera. Native RED-1
cells expressed G alpha i2, alpha i3, alpha s, and alpha q/11, but not
alpha i1 and alpha o. Terminal differentiation was associated with a
selective loss (approximately 80%) of G alpha i3 and an increase in a
truncated cytosolic form of G alpha i2, while the membrane levels of alpha
i2, alpha q/11, and alpha s did not change significantly. Treatment of G3
cells with the inducers was without effect on G protein abundance. However,
except for alpha s, G3 cells contained significantly higher levels of the
different G protein alpha subunits tested. Stimulation of G protein-coupled
receptors by thrombin and ADP caused a pertussis toxin (PTX)-inhibitable
transient increase in intracellular Ca2+ that was markedly reduced in
differentiated cells. In G3 cells, but not in 6C8 cells, thrombin also
caused a PTX- sensitive inhibition of isoprenaline-stimulated cyclic
3',5'-adenosine monophosphate (cAMP) formation. Our results show that
specific alterations in G protein expression and function are associated
with erythroid differentiation of erythroleukemia cells but do not prove a
causal relationship. The loss of G alpha i3 may affect cellular responses
that are mediated via P2T purine or thrombin receptors.
Volume 84,
Issue 12,
pp. 4088-4098,
12/15/1994
Copyright © 1994 by The American Society of Hematology
