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Microarray analysis using fluor-linked nucleotide probes generated from mRNA identifies altered genes in mice mutated in monoamine oxidase A implicated in neurodevelopment and prostate cancer


The advances in molecular biology have enabled a greater understanding of the mechanism underlying cognitive processes and the corresponding disorders.  Transmitting the signals by the nervous system involves a complex interplay between adjacent neurons.  

Prior to the excitation, the membrane of the neurons are kept in a polarized state (-70 miliVolts).  In the synapsis (space between adjacent neurons), the pre-synaptic neuron releases neurotransmitters.  For instance, in the case of sensory neurons, external signals such as light or sound are converted into the release of neurotransmitter.    The binding of the neurotransmitters to the receptors present in the post-synaptic neuron causes it to become depolarized through the action of voltage-gated ion channels, which allows the influx of ions such as sodium into the cell interior.  The rise in potential due to depolarization is followed by the the closure of the sodium channels and the opening of potassium channels to allow outward efflux of potassium ions to repolarize the cell membrane.  This process (i.e. axon potential) involves the sequential activation of ion channels to propagate the signal along the axon, resulting in the nerve conduction with a speed as high as 150 m/s (meter per second) for myelinated neurons.

 The diverse types of neurotransmitters include neuropeptides (ex. somatostatin), amino acids [ex. glutamate, GABA (gamma-aminobutyric acid)], gas (ex. nitric oxide), trace amines (ex. tyramine, tryptamine) in addition to the more commonly known molecules such as acetylcholine.  Among them are monoamines such as serotonin, epinephrine, norepinephrine, dopamine, and histamine.  The neurotransmitter serotonin is involved in a number of important cognitive processes including mood, sleep, thermoregulation, cognition, appetite, learning, vasoconstriction (blood pressure), memory, etc.

 The biosynthesis of serotonin involves converting L-tryptophan to 5-hydroxy-L-tryptophan, followed by its decarboxylation to yield serotonin.  Upon excitation, serotonin is released into synapsis, where it binds to the serotonin receptor (categorized into 7 families) to activate the post-synaptic neuron.  The released serotonin can be cleared from synapsis via serotonin reuptake pump.   Serotonin is degraded by monoamine oxidase (MAO), which also degrades other monoamine neurotransmitters such as norepinephrine and dopamine.  Following the discovery of two forms of MAO in 1968, the question remained as to whether the MAO A and B isoenzymes represent two distinct proteins or a single enzyme that has been differentially modified post-translationally (Johnston, 1968).  The enigma was solved by J. Shih and colleagues (University of Southern California, USA), who determined that they represent two separate polypeptides through the molecular cloning and sequencing of cDNAs encoding human MAO A and B (Bach et al., 1988).

                    

Lower levels of serotonin are associated with depression, and preventing the reuptake of serotonin from synapsis by the adjacent neurons, or suppressing the degradation of serotonin by inhibiting monoamine oxidase represent current antidepressant pharmacological strategies.  However, the use of non-selective MAO inhibitors (blocks both MAO A and B) can be accompanied by an adverse side effect known as 'cheese reaction', which occurs if administered while consuming fermented food (ex. wine, cheese), processed meat, bean products (ex. soy sauce) or chocolate that are rich in tyramine.  While both MAO A and B are expressed in the central nervous system, MAO A is also expressed in gastrointestinal tissues.  The inhibition of MAO A (plus absence of MAO B) in intestine by the drugs allows tyramine (undegraded) to enter circulation and activate medulla to release norepinephrine, which increases blood pressure suddenly via vasoconstriction.

 The identification of the MAO A gene enabled the generation of a 'knockout' mouse lacking the MAO A, which was done in collaboration with Seif and colleagues [Centre National de la Recherche Scientifique (CNRS), France] (Cases et al., 1995).  The MAO A deficiency led to an increased level of serotonin in the brain of mouse pups.  Intriguingly, adult male mice lacking MAO A exhibited enhanced aggression.  The result is reminiscent of Brunner syndrome (Brunner, 1993), which described abnormal aggression by males from a Dutch family carrying mutated (point deletion) MAO A gene (located in X chromosome) and lacking MAO A completely.  Nevertheless, it is thought that multiple factors including genes and environment may play critical role in the development of complex brain functions that underlie human behavior.

 To characterize the above phenomenon, the genes whose expression was altered in MAO A knockout mice were identified.  It entailed the generation of fluor-labeled probes using cDNAs reverse transcribed from mRNAs isolated from the brain of MAO A mutant mouse pups to hybridize to the Affymetrix GeneChip microarrays (Chen et al., 2017).  The differentially expressed genes identified were involved in various neural activities including cognitive function, neurodevelopment, neurotransmission, and apoptosis.

 Several lines of data suggest that MAO A may also play a role in prostate carcinogenesis.  An elevated expression of MAO A was observed in advanced stage prostate cancer (True et al., 2006, Wu et al., 2014).  The promoter of MAO A gene is regulated by androgen (male hormone) (Ou et al., 2006).  In the PTEN knockout mouse model of invasive prostate cancer, inactivating MAO A gene decreased the incidence of invasive cancer (Liao et al., 2018).  Mechanistically, inhibiting MAO A activity using drugs suppressed the signaling by androgen receptor (Gaur et al., 2019).  These results have led to a clinical trial (Phase II) assessing the efficacy of phenelzine (inhibits MAO A and B) in biochemically recurrent prostate cancer (BRPC) (Gross et al, 2021).  BRPC refers to the cases, in which the rise in PSA (prostate specific antigen) level is detected despite the inability to image metastasized cancer.



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References

Bach AW, Shih JC, et al. cDNA cloning of human liver monoamine oxidase A and B: molecular basis of differences in enzymatic properties.  Proc Natl Acad Sci U S A.  85:4934-8 (1988).  PMID: 3387449

Brunner HG, Nelen M, et al. Abnormal behavior associated with a point mutation in the structural gene for monoamine oxidase A.  Science. 262:578-80 (1993).  PMID: 8211186

Cases O, Shih JC, et al.  Aggressive behavior and altered amounts of brain serotonin and norepinephrine in mice lacking MAOA.  Science.  268:1763-6 (1995).  PMID: 7792602

Chen K, Shih JC, et al.  Altered gene expression in early postnatal monoamine oxidase A knockout mice.   Brain Res.  1669:18-26 (2017).  PMID: 28535982

Gaur S, Shih JC, et al.  Effect of Monoamine oxidase A (MAOA) inhibitors on androgen-sensitive and castration-resistant prostate cancer cells.  Prostate. 79:667-677 (2019).  PMID: 30693539

Gross ME, Shih JC, et al. Phase 2 trial of monoamine oxidase inhibitor phenelzine in biochemical recurrent prostate cancer.  Prostate Cancer Prostatic Dis.  24:61-68 (2021).  PMID: 32123315

Johnston JP.  Some observations upon a new inhibitor of monoamine oxidase in brain tissue.  Biochem Pharmacol.  17:1285-97 (1968). PMID: 5659776

Liao CP, Gross ME, Shih JC, et al. Loss of MAOA in epithelia inhibits adenocarcinoma development, cell proliferation and cancer stem cells in prostate.  Oncogene.   37:5175-5190 (2018).  PMID: 29844571

Ou XM, Shih JC., et al. Glucocorticoid and androgen activation of monoamine oxidase A is regulated differently by R1 and Sp1.  J Biol Chem. 281:21512-21525 (2006).  PMID: 16728402

True L, Coleman I, et al.  A molecular correlate to the Gleason grading system for prostate adenocarcinoma.  Proc Natl Acad Sci U S A.   103:10991-6 (2006).  PMID: 16829574

Wu JB, Chung LW, et al.  Monoamine oxidase A mediates prostate tumorigenesis and cancer metastasis.  J Clin Invest. 124:2891-908 (2014).  PMID: 24865426