Phosphorylation of DARPP-32 and protein phosphatase inhibitor-1 in rat choroid plexus: regulation by factors other than dopamine.
The molecular mechanisms underlying regulation of fluid production by secretory epithelia such as the choroid plexus are poorly understood. Two cAMP-regulated inhibitors of protein phosphatase-1, inhibitor-1 (I1) and a dopamine- and cAMP-regulated phosphoprotein, M(r) = 32,000 (DARPP-32), are enriched in the choroid plexus. We show here that these two phosphoproteins are colocalized in choroid plexus epithelial cells. We have developed a novel method for studying the phosphorylation state of DARPP-32 and I1 in intact cells, using a phosphorylation state-specific monoclonal antibody. Several drugs and hormones that are known to alter fluid secretion and that increase cAMP levels (forskolin, isoproterenol, vasoactive intestinal peptide) or cGMP levels (atrial natriuretic peptide) or that may use additional second messenger pathways (5-HT), increase the phosphorylation of I1 and DARPP-32 in rat choroid plexus. In contrast, dopamine does not alter cAMP and cGMP levels, or I1 and DARPP-32 phosphorylation. Our results indicate that DARPP-32, known to be regulated by dopamine in a number of tissues, can be phosphorylated in response to non-dopaminergic factors, including hormones acting through non-cAMP-dependent pathways. Our results also raise the possibility that inhibition of phosphatase-1, as a result of I1 and DARPP-32 phosphorylation, might be part of a final common pathway in the action of several factors that are known or thought to alter cerebrospinal fluid production.
Snyder
G L
GL
Laboratory of Molecular and Cellular Neuroscience, Rockefeller University, New York, New York 10021.
Girault
J A
JA
Chen
J Y
JY
Czernik
A J
AJ
Kebabian
J W
JW
Nathanson
J A
JA
Greengard
P
P
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D002831:Choroid Plexus / Q000378:metabolism*; D003342:Corpus Striatum / Q000378:metabolism; D004298:Dopamine / Q000502:physiology*; D051937:Dopamine and cAMP-Regulated Phosphoprotein 32; D007150:Immunohistochemistry; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D051937:Dopamine and cAMP-Regulated Phosphoprotein 32; D047908:Intracellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; D011506:Proteins; C021042:protein phosphatase inhibitor-1; D011494:Protein Kinases; D004298:Dopamine