title: Degradation of Serotonin N-Acetyltransferase, a Circadian Regulator, by the N-end Rule Pathway.
abstract: Serotonin N-acetyltransferase (AANAT) converts serotonin to N-acetylserotonin (NAS), a distinct biological regulator and the immediate precursor of melatonin, a circulating hormone that influences circadian processes, including sleep. N-terminal sequences of AANAT enzymes vary among vertebrates. Mechanisms that regulate the levels of AANAT are incompletely understood. Previous findings were consistent with the possibility that AANAT may be controlled through its degradation by the N-end rule pathway. By expressing the rat and human AANATs and their mutants not only in mammalian cells but also in the yeast Saccharomyces cerevisiae, and by taking advantage of yeast genetics, we show here that two "complementary" forms of rat AANAT are targeted for degradation by two "complementary" branches of the N-end rule pathway. Specifically, the N(α)-terminally acetylated (Nt-acetylated) Ac-AANAT is destroyed through the recognition of its Nt-acetylated N-terminal Met residue by the Ac/N-end rule pathway, whereas the non-Nt-acetylated AANAT is targeted by the Arg/N-end rule pathway, which recognizes the unacetylated N-terminal Met-Leu sequence of rat AANAT. We also show, by constructing lysine-to-arginine mutants of rat AANAT, that its degradation is mediated by polyubiquitylation of its Lys residue(s). Human AANAT, whose N-terminal sequence differs from that of rodent AANATs, is longer-lived than its rat counterpart and appears to be refractory to degradation by the N-end rule pathway. Together, these and related results indicate both a major involvement of the N-end rule pathway in the control of rodent AANATs and substantial differences in the regulation of rodent and human AANATs that stem from differences in their N-terminal sequences.
authors.affiliation: From the Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125.
authors.forename: Brandon
authors.initials: B
authors.lastname: Wadas
authors.affiliation: the Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, Michigan 48109.
authors.forename: Jimo
authors.initials: J
authors.lastname: Borjigin
authors.affiliation: the Department of Immunology, University of Connecticut School of Medicine, Farmington, Connecticut 06030, and.
authors.forename: Zheping
authors.initials: Z
authors.lastname: Huang
authors.affiliation: From the Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125.
authors.forename: Jang-Hyun
authors.initials: JH
authors.lastname: Oh
authors.affiliation: the Department of Life Sciences, Pohang University of Science and Technology, Pohang, Gyeongbuk, 790-784, South Korea.
authors.forename: Cheol-Sang
authors.initials: CS
authors.lastname: Hwang
authors.affiliation: From the Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125, avarsh@caltech.edu.
authors.forename: Alexander
authors.initials: A
authors.lastname: Varshavsky
mesh_terms: D000107:Acetylation; D000818:Animals; D047068:Arylalkylamine N-Acetyltransferase / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D009154:Mutation; D059748:Proteolysis; D051381:Rats; D012441:Saccharomyces cerevisiae; D054875:Ubiquitination / Q000502:physiology*
chemical_list: C478879:AANAT protein, human; C520223:Aanat protein, rat; D047068:Arylalkylamine N-Acetyltransferase
keywords: acetyltransferase; circadian; proteolysis; serotonin; ubiquitin