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P. Michael Iuvone, Audra D. Brown, Rashidul Haque, Joan Weller, Jolanta B. Zawilska, Shyam S. Chaurasia, Minhui Ma, David C. Klein; Retinal Melatonin Production: Role of Proteasomal Proteolysis in Circadian and Photic Control of Arylalkylamine N-Acetyltransferase. Invest. Ophthalmol. Vis. Sci. 2002;43(2):564-572.
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purpose. Dynamic day–night changes in melatonin synthesis are regulated by
changes in the activity of serotonin N-acetyltransferase
(arylalkylamine N-acetyltransferase [AA-NAT]).
Similarly, a light-induced decrease in AA-NAT activity at
night rapidly suppresses melatonin synthesis. The purpose of the
current study was to test the hypothesis that in vivo changes of AA-NAT
activity in chicken retina homogenates parallel changes in AA-NAT
protein. This led to examination of the role of proteasomal proteolysis
in the regulation of retinal AA-NAT activity and protein levels.
methods. Chickens, entrained to a 12-hour light–12-hour dark cycle, were
assessed under various lighting conditions, in some cases after in vivo
intravitreal administration of the protein synthesis inhibitor
cycloheximide or lactacystin, an inhibitor of the 20S proteasome.
Tissue homogenates were prepared, AA-NAT enzyme activity was measured,
and immunoreactive protein was estimated by Western blot using an
anti-chicken AA-NAT1-21 serum.
results. The abundance of AA-NAT protein in both the retina and pineal gland
exhibited a daily rhythm that was statistically indistinguishable from
that of AA-NAT’s activity measured in tissue homogenates. Acute
exposure to light at night rapidly decreased AA-NAT protein and
activity in a parallel fashion. Administration of cycloheximide at
night decreased retinal AA-NAT activity in darkness and enhanced the
effect of light. The light-evoked suppression of retinal AA-NAT protein
and activity was blocked by intravitreal injection of lactacystin,
which also was found to increase AA-NAT activity, either at night or
during the daytime.
conclusions. AA-NAT activity measured in tissue homogenates reflects the steady
state level of enzyme protein. AA-NAT protein in the retina turns over
rapidly, reflecting a balance of de novo synthesis and proteasomal
proteolysis. The suppressive effects of light at night are due
primarily to enhanced AA-NAT proteolysis.
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