Arylalkylamine N-Acetyltransferase: “the Timezyme”
2006; Elsevier BV; Volume: 282; Issue: 7 Linguagem: Inglês
10.1074/jbc.r600036200
ISSN1083-351X
Autores Tópico(s)Circadian rhythm and melatonin
ResumoArylalkylamine N-acetyltransferase controls daily changes in melatonin production by the pineal gland and thereby plays a unique role in biological timing in vertebrates. Arylalkylamine N-acetyltransferase is also expressed in the retina, where it may play other roles in addition to signaling, including neurotransmission and detoxification. Large changes in activity reflect cyclic 3′,5′-adenosine monophosphate-dependent phosphorylation of arylalkylamine N-acetyltransferase, leading to formation of a regulatory complex with 14-3-3 proteins. This activates the enzyme and prevents proteosomal proteolysis. The conserved features of regulatory systems that control arylalkylamine N-acetyltransferase are a circadian clock and environmental lighting. Arylalkylamine N-acetyltransferase controls daily changes in melatonin production by the pineal gland and thereby plays a unique role in biological timing in vertebrates. Arylalkylamine N-acetyltransferase is also expressed in the retina, where it may play other roles in addition to signaling, including neurotransmission and detoxification. Large changes in activity reflect cyclic 3′,5′-adenosine monophosphate-dependent phosphorylation of arylalkylamine N-acetyltransferase, leading to formation of a regulatory complex with 14-3-3 proteins. This activates the enzyme and prevents proteosomal proteolysis. The conserved features of regulatory systems that control arylalkylamine N-acetyltransferase are a circadian clock and environmental lighting. Arylalkylamine N-acetyltransferase (AANAT; EC 2.1.3.87) 2The abbreviations used are: AANAT, arylalkylamine N-acetyltransferase; AcCoA, acetyl coenzyme A; CRE, cAMP response element; CREB, cyclic AMP response element-binding protein; DAT, dopamine N-acetyltransferase; GNAT, Gcn5-related N-acetyltransferase; PCE, photoreceptor conserved element; PKA, cAMP-dependent protein kinase; PKA/14-3-3 sequence, PKA phosphorylation site nested within a nascent 14-3-3 binding motif; SCN, suprachiasmatic nucleus of the hypothalamus.2The abbreviations used are: AANAT, arylalkylamine N-acetyltransferase; AcCoA, acetyl coenzyme A; CRE, cAMP response element; CREB, cyclic AMP response element-binding protein; DAT, dopamine N-acetyltransferase; GNAT, Gcn5-related N-acetyltransferase; PCE, photoreceptor conserved element; PKA, cAMP-dependent protein kinase; PKA/14-3-3 sequence, PKA phosphorylation site nested within a nascent 14-3-3 binding motif; SCN, suprachiasmatic nucleus of the hypothalamus. plays a unique role in vertebrate biology by controlling the rhythmic production of melatonin in the pineal gland (Fig. 1) (1.Arendt J. Melatonin and the Mammalian Pineal Gland. Chapman and Hall, London1995Google Scholar). Activity increases 10- to 100-fold at night, causing an increase in the production and release of melatonin. The dynamics of AANAT activity are remarkable: the doubling time is ∼15 min at night, and the halving time of the decrease that follows a night → light transition is ∼3.5 min (2.Klein D.C. Weller J.L. Science. 1970; 169: 1093-1095Crossref PubMed Scopus (639) Google Scholar, 3.Klein D.C. Weller J.L. Science. 1972; 177: 532-533Crossref PubMed Scopus (282) Google Scholar). Circulating melatonin levels parallel changes in synthesis and release, due to rapid clearance by the liver (1.Arendt J. Melatonin and the Mammalian Pineal Gland. Chapman and Hall, London1995Google Scholar). The rhythmic pattern of activity in the melatonin pathway is a conserved feature of vertebrate biology, consistent with the role of melatonin as the vertebrate hormone of time, i.e. high levels signal night and low levels signal day. Although this signaling pattern is conserved, it is used in a variety of species-dependent ways to optimally control seasonal and daily changes in physiology (1.Arendt J. Melatonin and the Mammalian Pineal Gland. Chapman and Hall, London1995Google Scholar). The unique role that AANAT plays in vertebrate time keeping justifies the title of "the Timezyme." Our knowledge of the biological chemistry of AANAT is summarized in this overarching review. The AANAT Family−Aanat and Aanat homologs form the AANAT family, which is part of the large Gcn5-related acetyltransferase (GNAT) superfamily (4.Vetting M.W. S. de Carvalho L.P. Yu M. Hegde S.S. Magnet S. Roderick S.L. Blanchard J.S. Arch. Biochem. Biophys. 2005; 433: 212-226Crossref PubMed Scopus (467) Google Scholar, 5.Dyda F. Klein D.C. Hickman A.B. Annu. Rev. Biophys. Biomol. Struct. 2000; 29: 81-103Crossref PubMed Scopus (375) Google Scholar). All GNAT family members share common structural features associated with acetyl coenzyme A (AcCoA) binding; in addition, each member family has unique features reflecting substrate specificity for each of a wide range of substrates (e.g. aminoglycosides, diamines, puromycin, histones, and arylalkylamines). The GNAT superfamily also includes another arylalkylamine N-acetyltransferase family, represented by dopamine N-aceyltransferase (Dat), which is expressed in Drosophila melanogaster (6.Hintermann E. Grieder N.C. Amherd R. Brodbeck D. Meyer U.A. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 12315-12320Crossref PubMed Scopus (75) Google Scholar). Dat is not involved time keeping; rather, it functions in cuticle sclerotization and neurotransmission (6.Hintermann E. Grieder N.C. Amherd R. Brodbeck D. Meyer U.A. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 12315-12320Crossref PubMed Scopus (75) Google Scholar). DAT and AANAT family members have not been found in the same genome. AANAT family members have only been found in Gram-positive bacteria, fungi, algae, cephalochordates, and vertebrates; family members are not found in higher plants, insects, nematodes, or urochordates (7.Iyer L.M. Aravind L. Coon S.L. Klein D.C. Koonin E.V. Trends Genet. 2004; 20: 292-299Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar). This distribution may be explained by the initial appearance of the ancestral Aanat in Gram-positive bacteria, reflecting evolution from a GNAT family member. Following this, independent horizontal gene transfer events may have taken place (7.Iyer L.M. Aravind L. Coon S.L. Klein D.C. Koonin E.V. Trends Genet. 2004; 20: 292-299Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar, 8.Coon S.L. Roseboom P.H. Baler R. Weller J.L. Namboodiri M.A. Koonin E.V. Klein D.C. Science. 1995; 270: 1681-1683Crossref PubMed Scopus (309) Google Scholar). For example, an Aanat homolog may have been horizontally transferred from Gram-positive bacteria into the germ cell line of an ancestral vertebrate (7.Iyer L.M. Aravind L. Coon S.L. Klein D.C. Koonin E.V. Trends Genet. 2004; 20: 292-299Abstract Full Text Full Text PDF PubMed Scopus (166) Google Scholar, 9.Coon S.L. Klein D.C. Mol. Cell. Endocrinol. 2006; 252: 2-10Crossref PubMed Scopus (71) Google Scholar). Gene loss may also have influenced the distribution of Aanat family members. Aanat Homologs−AANAT homolog proteins lack several features that characterize AANATs (Fig. 2A) (9.Coon S.L. Klein D.C. Mol. Cell. Endocrinol. 2006; 252: 2-10Crossref PubMed Scopus (71) Google Scholar), including differences within the catalytic core, which explain why AANAT is >1000-fold more active than the yeast homolog (10.Liu B. Sutton A. Sternglanz R. J. Biol. Chem. 2005; 280: 16659-16664Abstract Full Text Full Text PDF PubMed Scopus (31) Google Scholar), and the presence of flanking regulatory regions. These changes appear to have occurred at a point in evolution after the emergence of cephalochordates and before the emergence of vertebrates. The AANAT homolog in fungi acetylates polyamines and arylalkylamines at relatively similar rates and has been referred to as a polyamine acetyltransferase (10.Liu B. Sutton A. Sternglanz R. J. Biol. Chem. 2005; 280: 16659-16664Abstract Full Text Full Text PDF PubMed Scopus (31) Google Scholar, 11.Ganguly S. Mummaneni P. Steinbach P.J. Klein D.C. Coon S.L. J. Biol. Chem. 2001; 276: 47239-47247Abstract Full Text Full Text PDF PubMed Scopus (50) Google Scholar). This pattern of substrate preference is consistent with a broad role in substituted alkylamine detoxification. Acetylation detoxifies amines by promoting elimination and blocking further reactions. Accordingly, it appears that the highly active and precisely regulated "Timezyme" may have evolved from a relatively sluggish detoxifying enzyme. Aanats−Aanats are expressed at high levels in the pinealocyte and retinal photoreceptor (12.Klein D.C. Coon S.L. Roseboom P.H. Weller J.L. Bernard M. Gastel J.A. Zatz M. Iuvone P.M. Rodriguez I.R. Begay V. Falcon J. Cahill G.M. Cassone V.M. Baler R. Recent Prog. Horm. Res. 1997; 52: 307-358PubMed Google Scholar), reflecting their evolutionary origin from a common ancestral photodetector (supplemental Fig. 1); many other genes dedicated to photodetection or melatonin synthesis share this pattern (13.Klein D.C. J. Biol. Rhythms. 2004; 19: 264-279Crossref PubMed Scopus (92) Google Scholar, 14.Klein D.C. Chronobiol. Int. 2006; 23: 5-20Crossref PubMed Scopus (67) Google Scholar). A single Aanat occurs among vertebrate classes other than fish, which have three Aanats (Aanat1a, Aanat1b, and Aanat2). This multiplicity reflects genome and gene duplication events in the fish line (9.Coon S.L. Klein D.C. Mol. Cell. Endocrinol. 2006; 252: 2-10Crossref PubMed Scopus (71) Google Scholar). Aanat1s are preferentially expressed in the fish retina and have similar affinity for phenylethylamines and indolylethylamines. Aanat2 is preferentially expressed in the fish pineal gland; the encoded protein prefers indolylethylamines over phenylethylamines (15.Coon S.L. Begay V. Deurloo D. Falcon J. Klein D.C. J. Biol. Chem. 1999; 274: 9076-9082Abstract Full Text Full Text PDF PubMed Scopus (96) Google Scholar, 16.Appelbaum L. Toyama R. Dawid I.B. Klein D.C. Baler R. Gothilf Y. Mol. Endocrinol. 2004; 18: 1210-1221Crossref PubMed Scopus (45) Google Scholar). Whereas it is clear that AANAT in the pineal gland functions in the melatonin pathway, the role of AANAT in the retina is less apparent. Although there is evidence that retinal AANAT is involved in local signaling via melatonin or N-acetylserotonin (17.Iuvone P.M. Tosini G. Pozdeyev N. Haque R. Klein D.C. Chaurasia S.S. Prog. Retin. Eye Res. 2005; 24: 433-456Crossref PubMed Scopus (299) Google Scholar), several findings suggest there are other functions. Arguing against a role of melatonin in signaling is that the last enzyme in melatonin synthesis, hydroxyindole-O-methyltransferase (Fig. 1), is absent from primate and ungulate retinas, whereas both express high levels of Aanat (13.Klein D.C. J. Biol. Rhythms. 2004; 19: 264-279Crossref PubMed Scopus (92) Google Scholar, 14.Klein D.C. Chronobiol. Int. 2006; 23: 5-20Crossref PubMed Scopus (67) Google Scholar, 18.Coon S.L. Del Olmo E. Young III, W.S. Klein D.C. J. Clin. Endocrinol. Metab. 2002; 87: 4699-4706Crossref PubMed Scopus (83) Google Scholar, 19.Bernard M. Donohue S.J. Klein D.C. Brain Res. 1995; 696: 37-48Crossref PubMed Scopus (57) Google Scholar). In addition, the findings that fish retina AANAT has a distinctly broader substrate preference than that in the pineal gland and that activity increases during the day, not night (20.Besseau L. Benyassi A. Moller M. Coon S.L. Weller J.L. Boeuf G. Klein D.C. Falcon J. Exp. Eye Res. 2006; 82: 620-627Crossref PubMed Scopus (66) Google Scholar), also point toward non-signaling roles. Candidate functions/roles include neurotransmission, which is consistent with the finding that AANAT acetylates dopamine, a retinal transmitter (21.Zilberman-Peled B. Ron B. Gross A. Finberg J.P. Gothilf Y. Brain Res. 2006; 1073–1074: 220-228Crossref PubMed Scopus (31) Google Scholar); acetylation of dopamine might also lead to a novel messenger. Another possible role is detoxification (supplemental Fig. 2); this is especially relevant in the retina because arylalkylamines can undergo Schiff base conjugation with retinaldehyde, the visual chemical required for photon capture (13.Klein D.C. J. Biol. Rhythms. 2004; 19: 264-279Crossref PubMed Scopus (92) Google Scholar, 14.Klein D.C. Chronobiol. Int. 2006; 23: 5-20Crossref PubMed Scopus (67) Google Scholar). Depletion of retinaldehyde by this route would erode photosensitivity. Moreover, the resulting bis-retinyl products are potentially toxic by virtue of photo-oxidation (22.Sparrow J.R. Boulton M. Exp. Eye Res. 2005; 80: 595-606Crossref PubMed Scopus (492) Google Scholar). Accordingly, in this context, retinal AANAT can be seen as an adjunct member of the retinoid cycle by preventing retinoid depletion (23.Blomhoff R. Blomhoff H.K. J. Neurobiol. 2006; 66: 606-630Crossref PubMed Scopus (676) Google Scholar). This effect of detoxification may have been the selective factor that led to the acquisition of Aanat by the ancestral photodetector (13.Klein D.C. J. Biol. Rhythms. 2004; 19: 264-279Crossref PubMed Scopus (92) Google Scholar, 14.Klein D.C. Chronobiol. Int. 2006; 23: 5-20Crossref PubMed Scopus (67) Google Scholar). AANAT is a globular ∼23-kDa cytosolic protein that forms a reversible regulatory complex with 14-3-3 proteins (8.Coon S.L. Roseboom P.H. Baler R. Weller J.L. Namboodiri M.A. Koonin E.V. Klein D.C. Science. 1995; 270: 1681-1683Crossref PubMed Scopus (309) Google Scholar, 12.Klein D.C. Coon S.L. Roseboom P.H. Weller J.L. Bernard M. Gastel J.A. Zatz M. Iuvone P.M. Rodriguez I.R. Begay V. Falcon J. Cahill G.M. Cassone V.M. Baler R. Recent Prog. Horm. Res. 1997; 52: 307-358PubMed Google Scholar, 24.Ganguly S. Gastel J.A. Weller J.L. Schwartz C. Jaffe H. Namboodiri M.A. Coon S.L. Hickman A.B. Rollag M. Obsil T. Beauverger P. Ferry G. Boutin J.A. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8083-8088Crossref PubMed Scopus (163) Google Scholar, 25.Ganguly S. Weller J.L. Ho A. Chemineau P. Malpaux B. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 1222-1227Crossref PubMed Scopus (179) Google Scholar, 26.Klein D.C. Ganguly S. Coon S. Weller J.L. Obsil T. Hickman A. Dyda F. Biochem. Soc. Trans. 2002; 30: 365-373Crossref PubMed Google Scholar, 27.Obsil T. Ghirlando R. Klein D.C. Ganguly S. Dyda F. Cell. 2001; 105: 257-267Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar, 28.Aitken A. Semin. Cancer Biol. 2006; 16: 162-172Crossref PubMed Scopus (633) Google Scholar, 29.Gardino A.K. Smerdon S.J. Yaffe M.B. Semin. Cancer Biol. 2006; 16: 173-182Crossref PubMed Scopus (215) Google Scholar, 30.Pozdeyev N. Taylor C. Haque R. Chaurasia S.S. Visser A. Thazyeen A. Du Y. Fu H. Weller J. Klein D.C. Iuvone P.M. J. Neurosci. 2006; 26: 9153-9161Crossref PubMed Scopus (38) Google Scholar) (Fig. 2, B and C and supplemental Fig. 3). Catalytic Core−The catalytic core of AANAT forms a cavity encompassing the arylalkylamine and AcCoA binding pockets. As with other GNAT superfamily members (5.Dyda F. Klein D.C. Hickman A.B. Annu. Rev. Biophys. Biomol. Struct. 2000; 29: 81-103Crossref PubMed Scopus (375) Google Scholar), AcCoA binds by contacts between the pantethiene moiety and the edge of a rigid β sheet directing the thioacetyl group into the center of the enzyme; the adenosine moiety is at the surface (31.Wolf E. De Angelis J. Khalil E.M. Cole P.A. Burley S.K. J. Mol. Biol. 2002; 317: 215-224Crossref PubMed Scopus (49) Google Scholar, 32.Hickman A.B. Namboodiri M.A. Klein D.C. Dyda F. Cell. 1999; 97: 361-369Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar) (Fig. 2). Arylalkylamines bind in a funnel-shaped pocket formed by three protein loops, which contact the aromatic ring of substrates via aromatic residues. This positions the protonated amine group of substrates close to the AcCoA thioacetyl group (31.Wolf E. De Angelis J. Khalil E.M. Cole P.A. Burley S.K. J. Mol. Biol. 2002; 317: 215-224Crossref PubMed Scopus (49) Google Scholar, 32.Hickman A.B. Namboodiri M.A. Klein D.C. Dyda F. Cell. 1999; 97: 361-369Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar, 33.Hickman A.B. Klein D.C. Dyda F. Mol. Cell. 1999; 3: 23-32Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar). AANAT preferentially acetylates arylethylamines, including the naturally occurring amines serotonin, tyramine, dopamine, tryptamine, dopamine, and phenylethylamine, and synthetic naphthyl-, benzofuryl-, and benzothienylethylamines (supplemental Fig. 4). Diamines and monoamines are relatively poor substrates of AANAT as are arylamines and arylalkylamines with 1-carbon side chains (8.Coon S.L. Roseboom P.H. Baler R. Weller J.L. Namboodiri M.A. Koonin E.V. Klein D.C. Science. 1995; 270: 1681-1683Crossref PubMed Scopus (309) Google Scholar, 34.Ferry G. Ubeaud C. Dauly C. Mozo J. Guillard S. Berger S. Jimenez S. Scoul C. Leclerc G. Yous S. Delagrange P. Boutin J.A. Protein Expr. Purif. 2004; 38: 84-98Crossref PubMed Scopus (12) Google Scholar, 35.Mesangeau C. Yous S. Chavatte P. Ferry G. Audinot V. Boutin J.A. Delagrange P. Bennejean C. Renard P. Lesieur D. J. Enzyme Inhib. Med. Chem. 2003; 18: 119-125Crossref PubMed Scopus (13) Google Scholar, 36.Ferry G. Loynel A. Kucharczyk N. Bertin S. Rodriguez M. Delagrange P. Galizzi J.P. Jacoby E. Volland J.P. Lesieur D. Renard P. Canet E. Fauchere J.L. Boutin J.A. J. Biol. Chem. 2000; 27539799Abstract Full Text Full Text PDF Google Scholar, 37.Ferry G. Loynel A. Kucharczyk N. Bertin S. Rodriguez M. Delagrange P. Galizzi J.P. Jacoby E. Volland J.P. Lesieur D. Renard P. Canet E. Fauchere J.L. Boutin J.A. J. Biol. Chem. 2000; 275: 8794-8805Abstract Full Text Full Text PDF PubMed Scopus (50) Google Scholar, 38.Khalil E. Cole P.A. J. Am. Chem. Soc. 1998; 120: 6195-6196Crossref Scopus (65) Google Scholar). Loops 2 and 3 are relatively rigid compared with loop 1, which is floppy (32.Hickman A.B. Namboodiri M.A. Klein D.C. Dyda F. Cell. 1999; 97: 361-369Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar, 33.Hickman A.B. Klein D.C. Dyda F. Mol. Cell. 1999; 3: 23-32Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar). In the absence of AcCoA, a portion of loop 1 can extend into and occupy the AcCoA binding domain. Loop 1 can be displaced by AcCoA (39.De Angelis J. Gastel J. Klein D.C. Cole P.A. J. Biol. Chem. 1998; 273: 3045-3050Abstract Full Text Full Text PDF PubMed Scopus (110) Google Scholar), favoring completion of the arylalkylamine binding pocket (Fig. 2, B and C, and supplemental movie 1). Loop 1 is longer in AANAT than in homologs due to the addition of a conserved tripeptide (64CPL66; residue numbering is based on ovine AANAT) (Fig. 2A), which is likely to impact binding or catalysis or both. Mechanism of Enzyme Action−Acetyl transfer is initiated by deprotonation of the protonated amine. This is indirectly facilitated by neighboring histidines (His120, 122) (Fig. 2A) (40.Scheibner K.A. De Angelis J. Burley S.K. Cole P.A. J. Biol. Chem. 2002; 277: 18118-18126Abstract Full Text Full Text PDF PubMed Scopus (55) Google Scholar) that are part of a "proton wire" conducting protons to the surface through a water-filled channel (32.Hickman A.B. Namboodiri M.A. Klein D.C. Dyda F. Cell. 1999; 97: 361-369Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar). Deprotonation leads to attack on the thioester bond of AcCoA and formation of a transient ternary complex. This decomposes into the N-acetylated arylalkylamine and CoAS– (coenzyme AS–). The former is ejected by the hydrophilic to hydrophobic conversion due to amine acetylation. Ejection of CoASH from the binding pocket requires tyrosine (Tyr168)-dependent protonation of CoAS (32.Hickman A.B. Namboodiri M.A. Klein D.C. Dyda F. Cell. 1999; 97: 361-369Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar, 40.Scheibner K.A. De Angelis J. Burley S.K. Cole P.A. J. Biol. Chem. 2002; 277: 18118-18126Abstract Full Text Full Text PDF PubMed Scopus (55) Google Scholar). A synthetic mimic of the ternary complex, CoA-S-N-acetyltryptamine, is a highly potent and specific AANAT inhibitor (27.Obsil T. Ghirlando R. Klein D.C. Ganguly S. Dyda F. Cell. 2001; 105: 257-267Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar, 31.Wolf E. De Angelis J. Khalil E.M. Cole P.A. Burley S.K. J. Mol. Biol. 2002; 317: 215-224Crossref PubMed Scopus (49) Google Scholar, 32.Hickman A.B. Namboodiri M.A. Klein D.C. Dyda F. Cell. 1999; 97: 361-369Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar, 38.Khalil E. Cole P.A. J. Am. Chem. Soc. 1998; 120: 6195-6196Crossref Scopus (65) Google Scholar, 41.Coon S.L. Weller J.L. Korf H.W. Namboodiri M.A. Rollag M. Klein D.C. J. Biol. Chem. 2001; 276: 24097-24107Abstract Full Text Full Text PDF PubMed Scopus (43) Google Scholar). AANAT Regulatory Regions−Evolutionary acquisition of the flanking regulatory regions represents a pivotal change in AANAT biology because it enabled cAMP to control rapid activation/inactivation and protection/degradation switching (26.Klein D.C. Ganguly S. Coon S. Weller J.L. Obsil T. Hickman A. Dyda F. Biochem. Soc. Trans. 2002; 30: 365-373Crossref PubMed Google Scholar, 42.Zheng W. Zhang Z. Ganguly S. Weller J.L. Klein D.C. Cole P.A. Nat. Struct. Biol. 2003; 10: 1054-1057Crossref PubMed Scopus (55) Google Scholar, 43.Zheng W. Schwarzer D. Lebeau A. Weller J.L. Klein D.C. Cole P.A. J. Biol. Chem. 2005; 280: 10462-10467Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar) by phosphorylating the Ser/Thr residues in the PKA/14-3-3 motifs (PKA-dependent phosphorylation site nested within a nascent 14-3-3 binding motif) (Fig. 2A). The N-terminal regulatory region is ∼30 residues in length. The primary structure of the PKA/14-3-3 motif 28RRHpTLP33 resembles that of a canonical "mode I" 14-3-3 binding motif (RSX(pS/pT)XP) whereas the three-dimensional configuration of this sequence in the AANAT/14-3-3 complex resembles that of a bound peptide with a "mode II" motif (RX(Y/F)X(pS/pT)XP) (27.Obsil T. Ghirlando R. Klein D.C. Ganguly S. Dyda F. Cell. 2001; 105: 257-267Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar). The sequence external to this site is disordered; the only conserved feature is a lysine (Lys10), which is thought to be critical for proteosomal proteolysis (44.Gastel J.A. Roseboom P.H. Rinaldi P.A. Weller J.L. Klein D.C. Science. 1998; 279: 1358-1360Crossref PubMed Scopus (252) Google Scholar). The C-terminal regulatory region contains the PKA/14-3-3 motif 200RRNpSG(C/R)205. This represents a "mode III" 14-3-3 binding motif ((pS/pT)X1–2-COOH). AANAT can bind to 14-3-3 proteins via either PKA/14-3-3 motif; however, binding via both is required for activation (24.Ganguly S. Gastel J.A. Weller J.L. Schwartz C. Jaffe H. Namboodiri M.A. Coon S.L. Hickman A.B. Rollag M. Obsil T. Beauverger P. Ferry G. Boutin J.A. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8083-8088Crossref PubMed Scopus (163) Google Scholar, 25.Ganguly S. Weller J.L. Ho A. Chemineau P. Malpaux B. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 1222-1227Crossref PubMed Scopus (179) Google Scholar, 27.Obsil T. Ghirlando R. Klein D.C. Ganguly S. Dyda F. Cell. 2001; 105: 257-267Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar, 45.Ganguly S. Coon S.L. Klein D.C. Cell Tissue Res. 2002; 309: 127-137Crossref PubMed Scopus (206) Google Scholar). Formation of the AANAT/14-3-3 Complex−Dark-dependent phosphorylation of AANAT and binding to 14-3-3 occurs in vivo as a function of environmental lighting (24.Ganguly S. Gastel J.A. Weller J.L. Schwartz C. Jaffe H. Namboodiri M.A. Coon S.L. Hickman A.B. Rollag M. Obsil T. Beauverger P. Ferry G. Boutin J.A. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8083-8088Crossref PubMed Scopus (163) Google Scholar, 25.Ganguly S. Weller J.L. Ho A. Chemineau P. Malpaux B. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 1222-1227Crossref PubMed Scopus (179) Google Scholar, 28.Aitken A. Semin. Cancer Biol. 2006; 16: 162-172Crossref PubMed Scopus (633) Google Scholar, 29.Gardino A.K. Smerdon S.J. Yaffe M.B. Semin. Cancer Biol. 2006; 16: 173-182Crossref PubMed Scopus (215) Google Scholar, 30.Pozdeyev N. Taylor C. Haque R. Chaurasia S.S. Visser A. Thazyeen A. Du Y. Fu H. Weller J. Klein D.C. Iuvone P.M. J. Neurosci. 2006; 26: 9153-9161Crossref PubMed Scopus (38) Google Scholar, 46.Aitken A. Plant Mol. Biol. 2002; 50: 993-1010Crossref PubMed Scopus (141) Google Scholar). PKA/14-3-3 motifs bind to amphipathic grooves on the inner surface of the half-pipe 14-3-3 dimer (24.Ganguly S. Gastel J.A. Weller J.L. Schwartz C. Jaffe H. Namboodiri M.A. Coon S.L. Hickman A.B. Rollag M. Obsil T. Beauverger P. Ferry G. Boutin J.A. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8083-8088Crossref PubMed Scopus (163) Google Scholar, 25.Ganguly S. Weller J.L. Ho A. Chemineau P. Malpaux B. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 1222-1227Crossref PubMed Scopus (179) Google Scholar). In addition, a significant contribution to binding comes from multiple contacts outside these regions (Fig. 2, B and C). Biochemical studies with the 14-3-3 ζ isoform argue that one AANAT binds to a single 14-3-3 dimer. However, structural studies with a mono-phosphorylated truncated AANAT (pT31 AANAT2–201) indicate that two AANATs can bind to a single 14-3-3 ζ dimer, each contacting 14-3-3 via the N-terminal PKA/14-3-3 motif (Fig. 2, B and C). The physiological stoichiometry of AANAT/14-3-3 complexes is unknown. Binding of AANAT to 14-3-3 activates the enzyme by favoring an optimal configuration of Loop 1, in which affinity for arylalkylamines is increased. It also shields AANAT from proteins involved in dephosphorylation and degradation and may also prevent thiol-dependent inactivation (24.Ganguly S. Gastel J.A. Weller J.L. Schwartz C. Jaffe H. Namboodiri M.A. Coon S.L. Hickman A.B. Rollag M. Obsil T. Beauverger P. Ferry G. Boutin J.A. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 8083-8088Crossref PubMed Scopus (163) Google Scholar, 25.Ganguly S. Weller J.L. Ho A. Chemineau P. Malpaux B. Klein D.C. Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 1222-1227Crossref PubMed Scopus (179) Google Scholar, 27.Obsil T. Ghirlando R. Klein D.C. Ganguly S. Dyda F. Cell. 2001; 105: 257-267Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar, 30.Pozdeyev N. Taylor C. Haque R. Chaurasia S.S. Visser A. Thazyeen A. Du Y. Fu H. Weller J. Klein D.C. Iuvone P.M. J. 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Chem. 2005; 280: 11544-11551Abstract Full Text Full Text PDF PubMed Scopus (50) Google Scholar, 51.Appelbaum L. Vallone D. Anzulovich A. Ziv L. Tom M. Foulkes N.S. Gothilf Y. J. Mol. Endocrinol. 2006; 36: 337-347Crossref PubMed Scopus (49) Google Scholar, 52.Kikuchi T. Raju K. Breitman M.L. Shinohara T. Mol. Cell. Biol. 1993; 13: 4400-4408Crossref PubMed Scopus (105) Google Scholar, 53.Burke Z. Wells T. Carter D. Klein D. Baler R. J. Neurochem. 1999; 73: 1343-1349Crossref PubMed Scopus (39) Google Scholar). PCEs bind members of the orthodenticle CRX/OTX transcription factor family, which are expressed in the pinealocyte and retina throughout life (16.Appelbaum L. Toyama R. Dawid I.B. Klein D.C. Baler R. Gothilf Y. Mol. Endocrinol. 2004; 18: 1210-1221Crossref PubMed Scopus (45) Google Scholar, 50.Appelbaum L. Anzulovich A. Baler R. Gothilf Y. J. Biol. Chem. 2005; 280: 11544-11551Abstract Full Text Full Text PDF PubMed Scopus (50) Google Scholar, 51.Appelbaum L. Vallone D. Anzulovich A. 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Furukawa T. Nat. Neurosci. 2003; 6: 1255-1263Crossref PubMed Scopus (445) Google Scholar). In addition, E-box elements contribute by blocking ectopic expression (16.Appelbaum L. Toyama R. Dawid I.B. Klein D.C. Baler R. Gothilf Y. Mol. Endocrinol. 2004; 18: 1210-1221Crossref PubMed Scopus (45) Google Scholar). 3A. Humphries, T. Wells, R. Baler, D. C. Klein, and D. A. Carter, unpublished observations made with transgenic mice. Circadian Rhythmicity and Photic "Turnoff"−Nearly all vertebrate AANAT control systems have two elements that regulate dynamics: a circadian clock and a photic turnoff mechanism. The clock is an autonomous ∼24-h oscillator constructed of transcriptional/translational feedback loops that act through interactions with E-boxes in clock gene promoters (61.Debruyne J.P. Noton E. Lambert C.M. Maywood E.S. Weaver D.R. Reppert S.M. Neuron. 2006; 50: 465-477Abstract Full Text Full Text PDF PubMed Scopus (341) Google Scholar, 62.Etchegaray J.P. Yang X. DeBruyne J.P. Peters A.H. Weaver D.R. Jenuwein T. Reppert S.M. J. Biol. Chem. 2006; 281: 21209-21215Abstract Full Text Full Text PDF PubMed Scopus (135) Google Scholar, 63.Reppert S.M. Weaver D.R. Nature. 2002; 418: 935-941Crossref PubMed Scopus (3334) Google Scholar). The clock is entrained to the environmental lighting cycle by light. Turnoff mechanisms cause rapid suppressive effects of light by decreasing cAMP levels, leading to dissociation of the AANAT/14-3-3 complex, followed by inactivation and proteosomal proteolytic destruction (supplemental Fig. 3) (44.Gastel J.A. Roseboom P.H. Rinaldi P.A. Weller J.L. Klein D.C. Science. 1998; 279: 1358-1360Crossref PubMed Scopus (252) Google Scholar). An exception to this general bipartite clock/turnoff model is found in salmonoid fish, in which AANAT activity is not regulated by a clock but only by a photic turnoff mechanism (64.Falcon J. Gothilf Y. Coon S.L. Boeuf G. Klein D.C. J. Neuroendocrinol. 2003; 15: 378-382Crossref PubMed Scopus (83) Google Scholar). This general model of regulation takes two forms, as regards the location of the clock and photodetectors (65.Klein D.C. Baler R. Roseboom P.H. Weller J.L. Bernard M. Gastel J.A. Zatz M. Iuvone P.M. Bega´y V. Falco´n J. Cahill G. Cassone V.M. Coon S.L. Lydic R. Baghdoyan H.A. Handbook of Behavioral State Control: Cellular and Molecular Mechanisms. CRC Press, Boca Raton, FL1998: 45-60Google Scholar): submammalian pinealocytes and vertebrate retinal photoreceptors or mammalian pinealocytes. Submammalian Pinealocytes and Vertebrate Retinal Photoreceptors−In submammalian pinealocytes and in retinal photoreceptors, the clock and AANAT are located in the same cell (17.Iuvone P.M. Tosini G. Pozdeyev N. Haque R. Klein D.C. Chaurasia S.S. Prog. Retin. Eye Res. 2005; 24: 433-456Crossref PubMed Scopus (299) Google Scholar, 50.Appelbaum L. Anzulovich A. Baler R. Gothilf Y. J. Biol. 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Mammalian Pinealocytes−The clock controlling the mammalian pinealocyte is located in the suprachiasmatic nucleus of the hypothalamus (SCN), which receives photic input from the retina via the retinohypothalamic tract. Light acts through this tract to entrain the clock and also acts downstream of the clock to block SCN stimulation of the pineal gland. The SCN is hardwired to the pineal gland by a neural pathway that traverses the brain, spinal cord, and superior cervical ganglia. Postganglionic projections from the latter innervate the pineal gland. At night, SCN stimulation releases norepinephrine into the pineal perivascular space (26.Klein D.C. Ganguly S. Coon S. Weller J.L. Obsil T. Hickman A. Dyda F. Biochem. Soc. Trans. 2002; 30: 365-373Crossref PubMed Google Scholar, 45.Ganguly S. Coon S.L. Klein D.C. Cell Tissue Res. 2002; 309: 127-137Crossref PubMed Scopus (206) Google Scholar, 72.Klein D.C. Ciba Found Symp. 1985; 117: 38-56PubMed Google Scholar). Norepinephrine activates adenylyl cyclase via β1-adrenergic receptors and increases intracellular Ca2+ and protein kinase C activity via α1B-adrenergic receptors (73.Sugden D. Vanecek J. Klein D.C. Thomas T.P. Anderson W.B. Nature. 1985; 314: 359-361Crossref PubMed Scopus (314) Google Scholar, 74.Ho A.K. Thomas T.P. Chik C.L. Anderson W.B. Klein D.C. J. Biol. Chem. 1988; 263: 9292-9297Abstract Full Text PDF PubMed Google Scholar), thereby potentiating β1-adrenergic receptor activation of adenylyl cyclase (75.Klein D.C. Sugden D. Weller J.L. Proc. Natl. Acad. Sci. U. S. A. 1983; 80: 599-603Crossref PubMed Scopus (244) Google Scholar). This "cross-talk" causes a large and rapid increase in cAMP (73.Sugden D. Vanecek J. Klein D.C. Thomas T.P. Anderson W.B. Nature. 1985; 314: 359-361Crossref PubMed Scopus (314) Google Scholar, 76.Ho A.K. Klein D.C. J. Biol. Chem. 1987; 262: 11764-11770Abstract Full Text PDF PubMed Google Scholar, 77.Sugden L.A. Sugden D. Klein D.C. J. Biol. 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Sassone-Corsi P. Trends Neurosci. 1997; 20: 487-492Abstract Full Text Full Text PDF PubMed Scopus (137) Google Scholar). Other factors appear to modulate Aanat transcription, including [Ca2+]i (87.Yu L. Schaad N.C. Klein D.C. J. Neurochem. 1993; 60: 1436-1443Crossref PubMed Scopus (34) Google Scholar), an unidentified rapidly turning over protein repressor (88.Ho A.K. Terriff D.L. Price D.M. Chik C.L. Chronobiol. Int. 2006; 23: 361-367Crossref PubMed Scopus (1) Google Scholar) and endogenous clock control of cAMP production (17.Iuvone P.M. Tosini G. Pozdeyev N. Haque R. Klein D.C. Chaurasia S.S. Prog. Retin. Eye Res. 2005; 24: 433-456Crossref PubMed Scopus (299) Google Scholar, 67.Chaurasia S.S. Pozdeyev N. Haque R. Visser A. Ivanova T.N. Iuvone P.M. Mol. Vis. 2006; 12: 215-223PubMed Google Scholar, 89.Chansard M. Liang J. Iwahana E. Baker T. Whittaker J. Fukuhara C. J. Pineal Res. 2006; 41: 85-94Crossref PubMed Scopus (5) Google Scholar, 90.Fukuhara C. Yamazaki S. Liang J. J. Neurochem. 2005; 93: 156-162Crossref PubMed Scopus (23) Google Scholar). In summary, Aanat expression requires ongoing activation via PCEs. In addition, in some cases, a rhythmic pattern of transcription is conferred by one or two mechanisms: cAMP-dependent activation via CREs (e.g. rat pinealocytes) or activation by clock gene products via E-boxes (e.g. zebrafish and chicken pinealocytes, rat and chicken retinal photoreceptors); both mechanisms operate in chicken photoreceptors. Vertebrate AANAT protein and activity are regulated via binding to 14-3-3 proteins. Continued research on AANAT is expected to lead to the development of drugs and genetic treatments that control activity of the enzyme. As a result, it may be possible to modulate endocrine function, sleep, mood, and behavior through effects on pineal melatonin synthesis and to improve vision by promoting detoxification of amines in the retina. I wish to express appreciation to Drs. P. Michael Iuvone, Yoav Gothilf, and Steven L. Coon for their careful review of this paper and constructive contributions. Download .pdf (.97 MB) Help with pdf files
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