Abstract
We examined here the role of second messenger-dependent kinases and
beta-arrestins in short-term regulation of the PTH receptor (PTHR)
signaling. The inhibition of protein kinase C (PKC) in COS-7 cells
transiently expressing PTHR, led to an approximately 2-fold increase
in PTH-stimulated inositol phosphate (IP) and cAMP production. The
inhibition of protein kinase A increased cAMP production 1.5-fold
without affecting IP signaling. The effects of PKC inhibition on
PTHR-mediated G(q) signaling were strongly decreased for a carboxy-terminally
truncated PTHR (T480) that is phosphorylation deficient. PKC inhibition
was associated with a decrease in agonist-stimulated PTHR phosphorylation
and internalization without blocking PTH-dependent mobilization of
beta-arrestin2 to the plasma membrane. Overexpression of beta-arrestins
strongly decreased the PTHR-mediated IP signal, whereas cAMP production
was impaired to a much lower extent. The regulation of PTH-stimulated
signals by beta-arrestins was impaired for the truncated T480 receptor.
Our data reveal mechanisms at, and distal to, the receptor regulating
PTHR-mediated signaling pathways by second messenger-dependent kinases.
We conclude that regulation of PTHR-mediated signaling by PKC and
beta-arrestins are separable phenomena that both involve the carboxy
terminus of the receptor. A major role for PKC and beta-arrestins
in preferential regulation of PTHR-mediated G(q) signaling by independent
mechanisms at the receptor level was established.
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