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Titel:Functional characterization of the Ustilago maydis protein Acb1 and its derived peptide SDF-2
Autor:Jungmann, Joachim
Weitere Beteiligte: Kahmann, Regine (Prof.Dr.)
Veröffentlicht:2016
URI:https://archiv.ub.uni-marburg.de/diss/z2017/0063
DOI: https://doi.org/10.17192/z2017.0063
URN: urn:nbn:de:hebis:04-z2017-00635
DDC: Biowissenschaften, Biologie
Titel(trans.):Funktionelle Charakterisierung des Ustilago maydis Proteins Acb1 und des produzierten Peptids SDF-2
Publikationsdatum:2017-09-14
Lizenz:https://rightsstatements.org/vocab/InC-NC/1.0/

Dokument

Schlagwörter:
Maisbeulenbrand, Acyl-CoA Bindeprotein, ustilago maydis, acyl-CoA binding protein

Summary:
The plant pathogenic fungus Ustilago maydis secretes effector proteins during the biotrophic interaction with its host plant Zea mays, with the aim being the suppression of the plant immune response virulence. All effector proteins virulence characterized to date were shown to be conventionally secreted proteins. However, over the last decade it could be established that proteins without a signal peptide can also be targeted to the outside of the cell in an ER/Golgi independent manner. A proteomic analysis on the apoplastic fluid of U. maydis infected maize plants identified 65 fungal proteins which did not contain a signal peptide. One of these proteins is the Acyl-CoA binding protein (ACBP) of U. maydis, Acb1, which will be investigated in this thesis. Over the years many studies have been dedicated to the functional analysis of Acyl-CoA binding proteins, in which ACBPs were shown to have many distinct intracellular functions, such as control over the intracellular Acyl-CoA pool size, roles in vesicular trafficking and their relevance for fatty acid biosynthesis. Further, in addition to their metabolic roles, ACBPs were shown to be involved in gene regulation, and also signaling by receptor binding, as was shown for the ACBP of the amoeba Dictyostelium discoideum (AcbA). In this organism it was shown that AcbA is secreted under nutrient limitation and extracellularly processed into the Spore Differentiation Factor 2 (SDF-2), which induces terminal spore differentiation. In this thesis it could be shown that the Acb1 protein of U. maydis is unconventionally secreted, subsequently processed into a SDF-2 like peptide and that this peptide is capable of inducing the sporulation of D. discoideum cells. The study of the protein’s extracellular function in U. maydis, as a full length secreted protein or in its processed form as the SDF-2 like peptide was hampered by the important intracellular functions of Acb1. Deletion of the acb1 gene resulted in strong growth defects and an accumulation of suppressor mutations, which were able to restore growth in a mutation dependent manner. Ultimately it could be shown that a virulence related function of the SDF-2 like peptide in both the solopathogenic strain SG200, as well as the compatible wild type strains FB1 and FB2, has to be considered unlikely. A potential extracellular role of the full length secreted protein has to date been neither confirmed nor ruled out.

Zusammenfassung:
Der Pflanzen-pathogene Pilz Ustilago maydis sekretiert während seiner biotrophen Interaction mit seiner Wirtspflanze Zea mays Effektor-proteine, mit dem Ziel die pflanzliche Immunantwort zu unterdrücken. Für alle bisher charakterisierten Effektoren wurde gezeigt, dass sie konventionell sekretiert werden. Über die letzten Jahrzehnte konnte jedoch gezeigt werden, dass Proteine, die nicht mit einem Signalpeptid transkribiert werden, auf einem ER/Golgi unabhängigen Weg aus der pilzlichen Zelle transportiert werden können. In einer proteomischen Analyse der apoplastischen Flüssigkeit von U. maydis-infizierten Mais Pflanzen konnten 65 pilzliche Proteine identifiziert werden die über kein Signalpeptid verfügen. Eines dieser Proteine ist das Acyl-CoA Binde Protein (ACBP), Acb1, welches in dieser Studie untersucht wurde. Über die vielen Jahre in denen Acyl-CoA Binde Proteine bereits untersucht wurden, konnten viele verschiedene Funktionen der Proteine entdeckt werden. Diese schließen die Kontrolle über intrazelluläre Acyl-CoA Vorräte, eine Funktion bei dem Transport intrazellulärer Vesikel und Beteiligung an der Fettsäuresynthese ein. Des Weiteren wurde gezeigt, dass ACBPs in der Genregulation eine wichtige Rolle spielen können und, wie für das homologe ACBP von Dictyostelium discoideum (AcbA) gezeigt wurde, als Signalstoffe für Rezeptorbindung agieren könne. Für AcbA konnte gezeigt werden, dass es spezifisch unter Nährstoffmangel sekretiert wird und extrazellulär in den Sporen Differenzierungs Faktor 2 (SDF-2) prozessiert wird, der endgültige Sporendifferenzierung einleitet. In dieser Arbeit konnte gezeigt werden, dass das Acb1 Protein von U. maydis auch unkonventionell sekretiert wird und nach der Sekretion in ein SDF-2 ähnliches Peptid prozessiert wird. Für das SDF-2 ähnliche Peptid konnte gezeigt werden, dass es Sporendifferenzierung in D. discoideum in einleitet. Das Untersuchen der extrazellulären Funktionen des in voller Länge sekretierten Proteins, oder des daraus entstehenden Peptids, wurde durch die wichtigen intrazellulären Aufgaben von Acb1 erschwert. Die Deletion des acb1 Gens war durch erhebliche Wachstumsdefekte charakterisiert, welche durch Suppressormutationen ganz oder teilweise unterdrückt werden konnten. Letztendlich konnte eine für die Virulenz relevante Rolle des SDF-2 ähnlichen Peptids in dem solopathogenen Stamm SG200, also auch den kompatiblen Wildtyp Stämmen FB1 und FB2, für unwahrscheinlich erklärt werden. Eine mögliche Rolle des unprozessierten Acb1 Proteins konnte bisher weder bestätigt, noch ausgeschlossen werden.

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