title: Aspartic acid substitutions in monoamine oxidase-A reveal both catalytic-dependent and -independent influences on cell viability and proliferation.
abstract: Post-translational influences could underlie the ambiguous roles of monoamine oxidase-A (MAO-A) in pathologies such as depression, cancer and Alzheimer disease. In support of this, we recently demonstrated that the Ca²⁺-sensitive component of MAO-A catalytic activity is inhibited by a pro-survival p38 (MAPK)-dependent mechanism. We substituted three aspartic acid (D) residues in human MAO-A that reside in putative Ca²⁺-binding motifs and overexpressed the individual proteins in the human HEK293 cell line. We assayed the overexpressed proteins for catalytic activity and for their influence on cell viability (using MTT conversion and trypan blue exclusion) and proliferation/DNA synthesis [using bromodeoxyuridine (BrdU) incorporation]. Innate MAO-A catalytic activity (and the capacity for generating hydrogen peroxide) was unaffected by the D61A substitution, but inhibited moderately or completely by the D248A and D328G substitutions, respectively. The Ca²⁺-sensitive activities of wild-type and D248A MAO-A proteins were enhanced by treatment with the selective p38(MAPK) inhibitor, SB203580, but was completely abrogated by the D61A substitution. Monoamine oxidase-A(D61A) was toxic to cells and exerted no effect on cell proliferation, while MAO-A(D248A) was generally comparable to wild-type MAO-A. As expected, the catalytic-dead MAO-A(D328G) was not cytotoxic, but unexpectedly enhanced both MTT conversion and BrdU staining. Variant-dependent changes in Bax and Bcl-2/Bcl-XL protein expression were observed. A different pattern of effects in N2-a cells suggests cell line-dependent roles for MAO-A. A catalytic-dependent mechanism influences MAO-A-mediated cytotoxicity, whereas a catalytic-independent mechanism contributes to proliferation. Context-dependent inputs by either mechanism could underlie the ambiguous pathological contributions of MAO-A.
authors.affiliation: Cell Signalling Laboratory, Department of Psychiatry, University of Saskatchewan, 107 Wiggins Rd., Saskatoon, SK, S7N 5E5, Canada.
authors.forename: Zelan
authors.initials: Z
authors.lastname: Wei
authors.forename: Tamara
authors.initials: T
authors.lastname: Satram-Maharaj
authors.forename: Bradley
authors.initials: B
authors.lastname: Chaharyn
authors.forename: Kelly
authors.initials: K
authors.lastname: Kuski
authors.forename: Paul R
authors.initials: PR
authors.lastname: Pennington
authors.forename: Xia
authors.initials: X
authors.lastname: Cao
authors.forename: Jennifer
authors.initials: J
authors.lastname: Chlan
authors.forename: Darrell D
authors.initials: DD
authors.lastname: Mousseau
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism*; D001973:Bromodeoxyuridine / Q000378:metabolism; D002118:Calcium / Q000494:pharmacology; D002384:Catalysis / Q000187:drug effects; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D047468:Immunoprecipitation; D051379:Mice; D008995:Monoamine Oxidase / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed / Q000379:methods; D009154:Mutation / Q000235:genetics*; D009447:Neuroblastoma / Q000473:pathology; D018384:Oxidative Stress / Q000187:drug effects / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D012701:Serotonin / Q000493:pharmacokinetics; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D013482:Superoxide Dismutase / Q000378:metabolism; D000072105:Superoxide Dismutase-1; D014162:Transfection; D014316:Tritium / Q000493:pharmacokinetics; D051020:bcl-X Protein / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D007093:Imidazoles; D019253:Proto-Oncogene Proteins c-bcl-2; D011725:Pyridines; C000606290:SOD1 protein, human; D051020:bcl-X Protein; D014316:Tritium; D001224:Aspartic Acid; D012701:Serotonin; C000606291:Sod1 protein, mouse; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D008995:Monoamine Oxidase; C579458:monoamine oxidase A, human; D001973:Bromodeoxyuridine; C093642:SB 203580; D002118:Calcium