Publikationsserver der Universitätsbibliothek Marburg
Titel:Die Analyse einer Ribokinase als neue myogenese-relevante Komponente und erste Hinweise auf eine Beteiligung von Ca2+-abhängigen Faktoren in der Myogenese von Drosophila melanogaster
Autor:Griemert, Barbara
Weitere Beteiligte: Renkawitz-Pohl, Renate (Prof. Dr.)
Veröffentlicht:2010
URI:https://archiv.ub.uni-marburg.de/diss/z2010/0146
URN: urn:nbn:de:hebis:04-z2010-01467
DOI: https://doi.org/10.17192/z2010.0146
DDC: Biowissenschaften, Biologie
Titel (trans.):Analysis of a ribokinase as a new myogenesis-relevant component and first indications for participation of Ca2+-dependent factors in myogenesis in Drosophila melanogaster
Publikationsdatum:2010-06-22
Lizenz:https://rightsstatements.org/vocab/InC-NC/1.0/

Dokument

Schlagwörter:
Muskelentwicklung, Myogenesis, Developmental Biology, Entwicklungsbiologie

Zusammenfassung:
Die EMS-induzierte Mutante E832 (P3-106/2) zeigt Störungen in der Entwicklung der somatischen und viszeralen larvalen Muskulatur in der Embryonalentwicklung von Drosophila melanogaster (Reichert 2004; Wierach 2005). Ziel der vorliegenden Arbeit war die weiterführende Analyse dieser Mutante, die Charakterisierung und Identifizierung des für den mutanten Phänotyp verantwortlichen Gens. Bei Komplementationsanalysen unter Zuhilfenahme von charakterisierten Deletionen zur Eingrenzung des Genortes zeigte sich, dass womöglich drei Letalmutationen in myogenese-relevanten Genen auf dem E832 -Chromosom vorliegen. Aufgrund dieser Annahme wurde eine Entkopplung durchgeführt, um die Mutationen mit Hilfe von Rekombinations-Ereignissen zu trennen. Die resultierenden Fliegenstämme wurden mit Immunhistologie- und Komplementationstests analysiert. Zwei Regionen (32F1-33A2 und 33E1-33E3) konnten für einen somatischen Muskelphänotyp verantwortlich gemacht werden. Die Manifestierung des viszeralen Phänotyps erwies sich nach der Entkopplung als schwierig und somit lag der Schwerpunkt auf der Untersuchung der somatisch-relevanten Mutationen in der Region 33E1-33E3. Ein Großteil der in dieser Region liegenden Gene wurde anschließend kloniert, um mit Hilfe des UAS-Gal4-Systems Rettungs-Experimente durchzuführen. Ebenfalls konnte die mRNA-Expression mittels in situ-Hybridisierungen analysiert und zusammen mit den Rettungs-Experimenten einige Gene als Kandidaten für eine Relevanz in der Muskelentwicklung ausgeschlossen werden, weil Expressionsmuster in anderen Geweben wie Tracheen (CG31862) und Garlandzellen (CG31763 und Pkd2) erhalten wurden. Nur eines der Kandidatengene (CG17010) ist während der Mesoderm-Entwicklung exprimiert. Für CG17010 ist die Kodierungskapazität für eine Ribokinase annotiert und erhielt daher die Bezeichnung D-Rbks33E. D-Rbks33E (CG17010) zeigte eine Rettung des mutanten, somatischen Phänotyps in 33E1-33E3. Die mRNA dieses Gens ist in der Embryogenese weitestgehend ubiquitär exprimiert, wobei eine erhöhte Transkriptmenge im somatischen Mesoderm zu beobachten ist. Die Sequenzierung des mutierten Allels von D-Rbks33E ergab die Deletion einer Base. Auf Proteinebene führt dies zum Verlust der ATP-Binderstelle der D-Rbks33E. Erste immunhistologische Analysen zeigen, dass die Zelladhäsion in der Mutante korrekt verläuft und deshalb ein Defekt im nachfolgenden Prozess während der Myoblastenfusion vorliegen muss. Auf der Suche nach weiteren möglichen fusionsrelevanten Proteinen wurde nach Ca2+-abhängigen Faktoren und hier speziell nach Ca2+-Bindeproteinen gesucht. Es konnte ein Kandidat, Kosh, identifiziert werden. In situ-Hybridisierungen mit kosh ergaben durch die spezifische Expression erste Anhaltspunkte auf eine Funktion in den fusionskompetenten Myoblasten während der Fusion mit den Founderzellen. Somit ist Kosh eines der ersten Proteine, welches anzeigt, dass Kalzium neben einer physiologischen auch eine entwicklungsbiologische Rolle für die Muskulatur in Drosophila spielen könnte. Folglich wurden mit Hilfe vorliegender Untersuchungen zwei neue Komponenten aufgedeckt, welche in die Signalkaskaden der Myogenese einzugliedern sind.

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