Ruolo del silenziamento epigenetico di miR-132 attraverso la metilazione del DNA nella patogenesi del cancro alla prostata
Author(s)
Cortelli, Silvia
Date Issued
May 5, 2013
Type
Doctoral Thesis
Abstract
Epigenetic refers to heritable changes in gene expression not involving the DNA sequence. Part of
the epigenome involves the methylation of cytosine in regions rich of CpG dinucleotides
called“CpG islands”.
The CpG islands are placed at the promoter regions and their methylation silenced the downstream
gene transcription.
In cancer aberrant promoter methylation has begun to explain the deregulated expression of small,
endogenous RNAs called microRNA (or miRNA). MicroRNAs inhibit mRNA translation to fine
tune gene network. In prostate cancer, the downregulation of microRNAs may contribute to cancer
initiation and development, and the cause may be their promoter hypermethylation.
To screen for epigenetically silenced miRNAs in prostate cancer, prostate normal epithelial (PrEC)
and carcinoma cells (PC3, DU145 and LNCaP) were treated with the demethylation agent 5-aza-2’-
deoxycytidine (AZA) and subsequently examined the expression changes of 650 miRNAs using
megaplex stemloop RT-qPCR.
After applying a selection strategy it was analyzed the methylation status of CpG islands upstream
a subset of miRNAs by methylation specific PCR (MSP). The CpG islands of miR-18b, miR-132,
miR-34b/c, miR-148a, miR-450a and miR-542-3p exhibited methylation patterns congruent with
their expression modulations in response to AZA.
Methylation analysis of these CpG islands in a panel of 50 human prostate carcinoma specimens
and 24 normal controls revealed miR-132 to be methylated in 42% of human cancer cases in a
positively manner correlated to total Gleason and tumor stage.
Expression analysis of miR-132 in our tissue panel confirmed its downregulation through
methylation in tumor. Re-expression of miR-132 in PC3 cells induced cell detachment followed by
cell death (anoikis).
Two proteins involved in cellular adhesion [heparin-binding epidermal growth factor (HB-EGF)
and Talin2] were confirmed as direct targets of miR-132.
These results demonstrate that miR-132 is a methylation-silenced miRNA with an antimetastatic
role in prostate cancer controlling cellular adhesion, furthermore taking into account the use of
miR-132 expression or CpG island methylation as a biomarker for PCa progression and its potential
therapeutic strategy for advanced PCa.
Additional information
Dottorato di ricerca in Genetica e biologia cellulare
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