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On novel functions of cholinesterases

Allebrandt, Karla Viviani (2006)
On novel functions of cholinesterases.
Technische Universität Darmstadt
Ph.D. Thesis, Primary publication

Abstract

The non-specificity of cholinesterases to cholinergic innervated tissues, their early onset during embryogenesis of many organisms, and their non-cholinolytic aryl acylamidase activity, indicate that these enzymes are involved with physiological processes other than the termination of nervous impulse. In this study, cholinesterases expression and function were investigated during the development of two model organisms, chicken (Gallus gallus) and zebrafish (Danio rerio), with the focus on non-cholinolytic and non-catalytic events. In chicken, the pineal organ was investigated taking into consideration: a) its similarity to the eye, as earlier studies suggested a relevance of cholinesterases to retina embryogenesis, b) its relevance on controlling physiological functions following a circadian rhythm, and c) its disfunction in pathological states, which also present altered cholinesterases expression, like Alzheimer’s disease. Indeed, in this study, a remarkable developmentally regulated switch from butyrylcholinesterase (BChE) to acetylcholinesterase (AChE) expression during pineal embryogenesis was found, in association with cell proliferation and differentiation, respectively. Even more, AChE-positive cells were shown to guide the pineal epithelium remodeling (leading to follicles development), indicating it plays a pivotal role in pineal embryogenesis. Besides, the appearance of follicular supportive cells correlated with this remodeling onset, followed by photoreceptor cells differentiation, indicating that these events are interconnected. Furthermore, AChE was demonstrated to be active in cells undergoing apoptosis during pineal embryogenesis, corroborating earlier in vitro studies indicating its involvement with the apoptotic process. However, the mechanism of action of cholinesterases in most of these developmental events is not clear, in particular whether the function could be structural or non-cholinolytic. Using zebrafish as a second model organism, a non-cholinolytic activity of AChE was investigated, from the time its transcription begins until larval development of this organism. This study revealed a particular profile of the AChE-associated aryl acylamidase activity (AAA) during development of zebrafish. AAA was particularly more pronounced than the esterase activity during zebrafish embryogenesis, indicating a relevance of this activity during early development. This non-cholinolytic activity was further investigated in human recombinant BChE wild-type and mutant proteins to address its catalytic power in enzymes with low cholinergic functionality. Altogether, these three studies on novel functions of cholinesterases address aspects of these enzymes also in relation to serotonin, as follow: a) cholinesterases are implicated in the development of the pineal gland, an organ controlling serotonin metabolism; b) a temporal high sensitivity of zebrafish embryos towards serotonin administration correlated with AChE expression onset during their blastula period, and c) serotonin directly interacts with cholinesterases, demonstrated through a non-competitive inhibition of the AAA activity on purified recombinant human BChE. This PhD work, therefore, presents strong evidence of the AChE involvement with morphogenesis, with further implications of its expression for pineal cells differentiation and apoptosis. It also writes further history on the little investigated side activity of cholinesterases, the aryl acylamidase, and supports a link between cholinergic and serotonergic systems.

Item Type: Ph.D. Thesis
Erschienen: 2006
Creators: Allebrandt, Karla Viviani
Type of entry: Primary publication
Title: On novel functions of cholinesterases
Language: English
Referees: Layer, Prof. Dr. Paul G. ; Himstedt, Prof. Dr. Werner
Advisors: Layer, Prof. Dr. Paul G.
Date: 31 January 2006
Place of Publication: Darmstadt
Publisher: Technische Universität
Refereed: 4 November 2005
URL / URN: urn:nbn:de:tuda-tuprints-6446
Abstract:

The non-specificity of cholinesterases to cholinergic innervated tissues, their early onset during embryogenesis of many organisms, and their non-cholinolytic aryl acylamidase activity, indicate that these enzymes are involved with physiological processes other than the termination of nervous impulse. In this study, cholinesterases expression and function were investigated during the development of two model organisms, chicken (Gallus gallus) and zebrafish (Danio rerio), with the focus on non-cholinolytic and non-catalytic events. In chicken, the pineal organ was investigated taking into consideration: a) its similarity to the eye, as earlier studies suggested a relevance of cholinesterases to retina embryogenesis, b) its relevance on controlling physiological functions following a circadian rhythm, and c) its disfunction in pathological states, which also present altered cholinesterases expression, like Alzheimer’s disease. Indeed, in this study, a remarkable developmentally regulated switch from butyrylcholinesterase (BChE) to acetylcholinesterase (AChE) expression during pineal embryogenesis was found, in association with cell proliferation and differentiation, respectively. Even more, AChE-positive cells were shown to guide the pineal epithelium remodeling (leading to follicles development), indicating it plays a pivotal role in pineal embryogenesis. Besides, the appearance of follicular supportive cells correlated with this remodeling onset, followed by photoreceptor cells differentiation, indicating that these events are interconnected. Furthermore, AChE was demonstrated to be active in cells undergoing apoptosis during pineal embryogenesis, corroborating earlier in vitro studies indicating its involvement with the apoptotic process. However, the mechanism of action of cholinesterases in most of these developmental events is not clear, in particular whether the function could be structural or non-cholinolytic. Using zebrafish as a second model organism, a non-cholinolytic activity of AChE was investigated, from the time its transcription begins until larval development of this organism. This study revealed a particular profile of the AChE-associated aryl acylamidase activity (AAA) during development of zebrafish. AAA was particularly more pronounced than the esterase activity during zebrafish embryogenesis, indicating a relevance of this activity during early development. This non-cholinolytic activity was further investigated in human recombinant BChE wild-type and mutant proteins to address its catalytic power in enzymes with low cholinergic functionality. Altogether, these three studies on novel functions of cholinesterases address aspects of these enzymes also in relation to serotonin, as follow: a) cholinesterases are implicated in the development of the pineal gland, an organ controlling serotonin metabolism; b) a temporal high sensitivity of zebrafish embryos towards serotonin administration correlated with AChE expression onset during their blastula period, and c) serotonin directly interacts with cholinesterases, demonstrated through a non-competitive inhibition of the AAA activity on purified recombinant human BChE. This PhD work, therefore, presents strong evidence of the AChE involvement with morphogenesis, with further implications of its expression for pineal cells differentiation and apoptosis. It also writes further history on the little investigated side activity of cholinesterases, the aryl acylamidase, and supports a link between cholinergic and serotonergic systems.

Alternative Abstract:
Alternative abstract Language

Die Unspezifität von Cholinesterasen gegenüber cholinergisch aktiviertem Gewebe, ihre frühe Entstehung während der Embryogenese vieler Organismen, sowie ihre nicht-cholinergische Arylacylamidase Aktivität, weisen darauf hin, dass diese Enzyme mit noch anderen physiologischen Prozessen als der Beendigung von Nervenimpulsen zusammenhängen. In dieser Arbeit wurden die Cholinesterase Expression und Funktion, mit dem Fokus auf nicht-cholinolytischen und nicht-katalytischen Ereignissen, während der Entwicklung zweier Modell Organismen, dem Huhn (Gallus gallus) und dem Zebrafisch (Danio rerio), untersucht. Im Huhn wurde die Pinealdrüse untersucht. Dabei flossen die folgenden Punkte in die Betrachtung mit ein: a) die Ähnlichkeit zum Auge – frühere Studien deuteten auf eine Rolle von Cholinesterasen bei der Embryogenese der Retina hin; b) die Relevanz bei der Kontrolle von physiologischen Funktionen die einem zirkadischen Rhythmus folgen; c) die Dysfunktion pathologischer Zustände mit veränderter Cholinesterase Expression wie z.B. der Alzheimer Krankheit. Tatsächlich wurde in der vorliegenden Arbeit ein bemerkenswerter entwicklungsgesteuerter Wechsel von Butyrylcholinesterase- (BChE) zu Acetylcholinesterase-Expression (AChE), in Zusammenhang mit Zellproliferation beziehungsweise –differenziation, gefunden. Weiterhin wurde gezeigt, dass AChE positive Zellen die Remodellierung des Pineal Epithels leiten (was zur Follikelentwicklung führt) und damit eine entscheidende Rolle bei der Embryogenese der Pinealdrüse spielen. Das Auftauchen von follikularen Unterstützungszellen in Korrelation mit dem Beginn der Remodellierung, gefolgt von Photorezeptordifferenzierung, weist drauf hin, dass diese Ereignisse zusammen hängen. Zusätzlich wurde gezeigt, dass AChE aktiv ist in Zellen, die Apoptose während der Embryogenese der Pinealdrüse durchlaufen, was die in in-vitro Experimenten entdeckte Rolle beim apoptotischen Prozess untermauert. Der Mechanismus jedoch, nach dem die Cholinesterasen in den meisten der Entwicklungsereignisse wirken, ist nicht bekannt. Insbesondere die Frage, ob die Funktion struktureller - oder nicht-cholinolytischer Natur ist, ist unklar. In Zebrafischen, als zweitem Modellorganismus, wurde eine nicht-cholinolytische Aktivität von AChE vom Beginn der Transkription bis zur Larvenentwicklung untersucht. Durch diese Studie wurde ein spezielles Profil der mit AChE zusammenhängenden Arylacylamidase Aktivität (AAA) während der Entwicklung von Zebrafischen entdeckt. Insbesondere war während der Embryogenese von Zebrafischen die AAA stärker ausgeprägt als die Esterase Aktivität. Damit scheint diese [AAA] Aktivität eine Bedeutung für die frühe Entwicklung zu haben. Weiterhin wurde diese nicht-cholinolytische Aktivität in menschlicher, rekombinanter BChE (wild-type) und mutierten Proteinen untersucht, um ihre katalytische Wirksamkeit in Enzymen mit niedriger cholinergischer Funktionalität und zu ermitteln. Die Ergebnisse dieser Studie deuten darauf hin, dass die Esterase und AAA Aktivitäten von verschiedenen katalytisch aktiven Stellen auf der Cholinesterase ausgehen, und das nicht-cholinergisch aktive, mutiert Enzyme AAA Aktivität zeigen können. Gemeinsam behandeln diese 3 Studien über neuer Funktionen von Cholinesesterasen auch Aspekte in Zusammenhang mit Serotonin: A) Cholinesterasen werden in der Entwicklung der Pineal Drüse benötigt, einem Organ, welches den Serotonin Metabolismus kontrolliert; B) eine temporäre, hohe Empfindlichkeit von Zebrafisch Embryos gegenüber Serotoningabe in Zusammenhang mit AchE Expressionsbeginn während der Blastula Phase; und C) Serotonin wechselwirkt direkt mit Cholinesterasen, was durch nicht-kompetitive Inhibition der AAA-Aktivität gegenüber gereinigter, rekombinanter menschlicher BChE demonstriert wurde. Diese Dissertation zeigt starke Indizien für den Zusammenhang von AChE mit Morphogenese, mit weiteren Auswirkungen der AChE Expression die Pinealzellen Differentiation und Apoptose. Des Weiteren wird das Wissen über die wenig untersuchte Arylacylamidase, einer Nebenaktivität von Cholinesterase, erweitert - sowie eine Verbindung zwischen dem cholinergischen und dem serontonergischen System hergestellt.

German
Uncontrolled Keywords: Pineal, Aryl acylamidase, photoreceptors, differentiation
Classification DDC: 500 Science and mathematics > 570 Life sciences, biology
Divisions: 10 Department of Biology
Date Deposited: 17 Oct 2008 09:22
Last Modified: 05 Mar 2013 09:26
PPN:
Referees: Layer, Prof. Dr. Paul G. ; Himstedt, Prof. Dr. Werner
Refereed / Verteidigung / mdl. Prüfung: 4 November 2005
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