Abstract

Genes that regulate essential cellular functions, such as apoptosis or proliferation, must be subject to tight transcriptional control. MiRNAs located in introns of these genes could fulfill important regulatory functions by directly or indirectly controlling the expression of their host genes. The present work dealt with the question of whether intronically localized miRNAs can exert a regulatory influence on their host genes and whether these control mechanisms are impaired or even inactivated in tumors. This was investigated in two known tumorigenic kinases, AKT2 and MAP2K4, and the intronically located miRNA-641 and miRNA-744 by real-time PCR, RNA interference experiments, protein analysis, immunohistochemistry, reporter gene assays, established long-term cell cultures, primary cell lines from glioblastoma tissue and glioblastoma biopsies. It could be shown that both kinases -indeed- are inhibited by their intronic miRNAs via indirect negative feedback: miR-641 inhibits phosphorylation via the targets PIK3R3, MAPKAP1 and NFAT5 and thus the activity of its tumor host gene AKT2; miRNA-744 inhibits the activity of the SMAD and MAPK signaling pathway by targeting TGFB1. We further found that both miRNAs are strongly repressed in glioblastomas as compared to normal brain tissue. In contrast, the miRNA target genes PIK3R3, NFAT5 and TGFB1 were induced in tumor tissue. The present doctoral thesis was thus able to demonstrate an important principle of expressional control: intronic miRNAs can regulate their host genes via complex functional networks. The disruption of these negative feedback mechanisms can significantly influence intracellular signalling promote tumour progression. These results may contribute to the development of miRNA-based future therapeutic strategies.