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J Biol Chem, Vol. 275, Issue 2, 1479-1484, January 14, 2000
From the Recent studies implicate a role in hepatocyte
organic anion transport of a plasma membrane protein that has been
termed oatp1 (organic anion transport protein 1). Little is known
regarding mechanisms by which its transport activity is modulated
in vivo. In previous studies (Campbell, C. G., Spray,
D. C., and Wolkoff, A. W. (1993) J. Biol.
Chem. 268, 15399-15404), we demonstrated that hepatocyte uptake
of sulfobromophthalein was down-regulated by extracellular ATP. We have
now found that extracellular ATP reduces the
Vmax for transport of sulfobromophthalein by
rat hepatocytes; Km remains unaltered. Reduced
transport also results from incubation of hepatocytes with the
phosphatase inhibitors okadaic acid and calyculin A. Immunoprecipitation of biotinylated cell surface proteins indicates
that oatp1 remains on the cell surface after exposure of cells to ATP
or phosphatase inhibitor, suggesting that loss of transport activity is
not caused by transporter internalization. Exposure of
32P-loaded hepatocytes to extracellular ATP results in
serine phosphorylation of oatp1 with the appearance of a single major
tryptic phosphopeptide; oatp1 from control cells is not phosphorylated.
This phosphopeptide comigrates with one of four phosphopeptides
resulting from incubation of cells with okadaic acid. These studies
indicate that the phosphorylation state of oatp1 must be an important
consideration when assessing alterations of its functional expression
in pathobiological states.
Down-regulation by Extracellular ATP of Rat Hepatocyte Organic
Anion Transport Is Mediated by Serine Phosphorylation of Oatp1*
,, and Department of Molecular Pharmacology and
§ Marion Bessin Liver Research Center, Albert Einstein
College of Medicine, Bronx, New York 10461
*
This work was supported by National Institutes of Health
Grants DK-23026, DK-41296, and CA-56677.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
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