Immunfluoreszenz-Basierte Hochdurchsatzanalyse Identifiziert MK5 als Myc-Regulierende Kinase

Der Transkriptionsfaktor Myc spielt eine entscheidende Rolle bei der Tumorentstehung und dem Tumorwachstum. Dabei ist das MYC-Gen in humanen Tumoren selten mutiert, sondern häufig sind Mechanismen der post-transkriptionellen Regulation der Myc- Proteinexpression verändert. Zur Identifikation unbe...

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Bibliographic Details
Main Author: Kreß, Theresia
Contributors: Lingelbach, Klaus (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2011
Online Access:PDF Full Text
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The transcription factor Myc plays an essential role in tumor development, growth and maintenance. The MYC gene itself is mutated only rarely. In most tumors, mechanisms of post-transcriptional regulation of the MYC mRNA or the Myc protein expression are altered. To identify novel mechanisms of post-transcriptional regulation of Myc, a high throughput screening platform was developed and established. High throughput analyses using RNAi libraries targeting the human kinome identified the protein kinase MK5/MAPKAPK5/PRAK as negative regulator of Myc protein expression. Depletion of MK5 increased Myc protein expression whereas overexpression of MK5 reduced Myc levels. MK5 directly phosphorylates the transcription factor FoxO3a at multiple serine residues in vitro: S215, S253, S551 and S555. Phosphorylation of S215 was confirmed in vivo using a phospho-specific antibody. Phosphorylation of S215 induces nuclear localization of FoxO3a. After activation by MK5, FoxO3a binds to the promoter of the miR-34b/c and induces their transcription. miR-34b and miR-34c bind to the 3’-UTR of the MYC mRNA, inhibit its translation and therefore reduce Myc protein expression. In addition, Myc binds to the E-box region within the MK5 promoter and induces its transcription. Therefore, MK5 itself is a direct target of the Myc transcription factor, creating a negative feedback loop consisting of Myc, MK5, FoxO3a and miR-34b/c. This negative feedback loop plays an important role in suppressing the progression of colon carcinogenesis. However, at the transition from well-differentiated to un-differentiated colorectal adenocarcinomas, MK5 mRNA as well as MK5 protein expression is reduced. This leads to increased Myc protein expression, tumor progression as well as metastasis.