Cholera toxin and its B subunit do not change cytosolic free calcium concentration.
Using the fluorescent Ca2+ probe Quin-2 it has been reported that cholera toxin (CT) and its B subunit (B-CT) increase cytosolic free Ca2+ concentration ([Ca2+]i) in entherocytes, thymocytes and fibroblasts. In this work we show, however, that the fluorescence increases of Quin-2-loaded cells (rat thymocytes, mouse splenocytes, P-388 macrophages and 3T3 fibroblasts) observed upon addition of CT or B-CT are not caused by an increase in [Ca2+]i. The observed effect appears to be accounted for by EDTA-2Na admixtures (present as conservation agent in all CT and B-CT preparations) which 'unquenches' the fluorescence of Quin-2 acid leaked out from the cells into the extracellular medium and produces influorescent complexes with contaminating heavy metal ions. Thus the mitogenic effect of B-CT is not obviously connected with the cytosolic free Ca2+ increase but is probably due to ganglioside-mediated protein phosphorylation.
Astashkin
E I
EI
All-Union Research Institute of Biotechnology, Moscow, USSR.
Surin
A M
AM
Mikhna
M G
MG
Nikolaeva
I S
IS
Lazarev
A V
AV
Gukovskaya
A S
AS
D000818:Animals; D002118:Calcium / Q000032:analysis / Q000378:metabolism*; D002460:Cell Line; D002772:Cholera Toxin / Q000494:pharmacology*; D003600:Cytosol / Q000032:analysis / Q000378:metabolism*; D005347:Fibroblasts / Q000032:analysis / Q000378:metabolism; D008264:Macrophages / Q000032:analysis / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D010446:Peptide Fragments / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains; D013050:Spectrometry, Fluorescence; D013154:Spleen / Q000032:analysis / Q000166:cytology / Q000378:metabolism; D013950:Thymus Gland / Q000032:analysis / Q000166:cytology / Q000378:metabolism
D010446:Peptide Fragments; C050310:cholera toxin, B subunit (50-64); D002772:Cholera Toxin; D002118:Calcium