Biology Reference
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2. UBIQUITINATION OF ARRESTINS
2.1. Mechanisms: Modification by E3 ubiquitin ligase(s)
Ubiquitination of arrestin was first demonstrated for the mammalian
b
-arrestin2. Upon agonist-stimulation of the
b
2
AR, isolated
b
-arrestin2
immunoprecipitates showed a smeary laddering of ubiquitination when
immunoblotted for ubiquitin.
48
This pattern is characteristic of protein
polyubiquitination; however, unlike many polyubiquitinated proteins that
are recognized by the 26S proteasomal proteases and degraded,
b
-arrestin2
ubiquitination does not lead to detectable degradation of the protein.
48
It
was found that the RING domain-containing E3 ubiquitin ligase Mdm2
mediates this ubiquitination, and further that when the process is blocked
with Mdm2 mutants that lack the catalytic domain, but could still bind
b
-arrestin2, there is a significant inhibition of
b
2
AR internalization.
48
Stud-
ies have also revealed that the restoration of Mdm2 expression in Mdm2/
p53 double knock-out mouse embryonic fibroblasts resulted in the rescue
of both isoproterenol-stimulated
b
-arrestin ubiquitination and rapid recep-
tor internalization.
48
These effects are also observed in heterologous cells
such as human embryonic kidney cells (HEK293) in which Mdm2 expres-
sion is depleted using Mdm2-specific siRNA or in Chinese hamster fibro-
blasts (CHW) in which Mdm2 activity is inhibited by exogenous expression
of a truncated Mdm2 lacking the catalytic domain.
48-50
Isoproterenol-
stimulated Mdm2-dependent ubiquitination of
b
-arrestin is therefore an
essential molecular event required for rapid internalization of the
b
2
AR into
clathrin-coated vesicles. The importance of
b
-arrestin2 ubiquitination in
7TMR regulation is corroborated by several alternative experimental
approaches, such as mutant
b
-arrestins that are stabilized and impaired in
ubiquitination
51
and regulation by deubiquitination.
49
In addition to Mdm2, other E3 ubiquitin ligases, namely, Nedd4,
50
AIP4,
52
and Parkin,
53
have also been shown to bind
b
-arrestin2; however,
these interactions do not seem to result in
b
-arrestin ubiquitination.
b
-Arrestin has also been shown to be SUMOylated upon
b
2
AR stimulation.
SUMO or small ubiquitin-like modifier is a ubiquitin-like protein that is also
appended to lysines in substrate proteins.
54
SUMOylation has analogous
effects to ubiquitination in modulating protein interactions, signaling, and
gene transcription, but unlike ubiquitin, it does not promote proteasomal
degradation. Additionally, unlike ubiquitination, SUMOylation is generally
targeted to the canonical motif
c
-K-x-E which represents the recognition
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