Zellbasierte Hochdurchsatzanalysen identifizieren eine zentrale, wachstumsregulierende Rolle von Cofilin-1 im humanen Pankreaskarzinom
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Philipps-Universität Marburg
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Abstract
Due to the poor prognosis of the human ductal adenocarcinoma of the pancreas
(PDAC) it is of major importance to identify novel target genes with high functional
relevance to provide a better diagnosis or new therapeutic approaches.
In this study, 91 potential candidate genes identified as differentially expressed in
carcinogenic pancreatic tissues in previous high-throughput analyses were functionally
characterized. To this end, the technology of “reverse transfection“ was established in
our lab to perform the analyses in a highly parallelized microarray approach.
Overexpression and knockdown experiments were carried out in transformed and nontransformed
cell lines for all candidates to screen for subcellular localisation of the
gene products and their impact on cellular functions like apoptosis, proliferation and
differentiation.
Data evaluation led to the selection of 10 genes which showed significant and
reproducible effects in the parallelized experiments. One of them was Cofilin-1 (CFL1),
a member of the ADF/Cofilin family of actin-binding proteins. Parallelized microarray
analyses revealed a previously undocumented growth-regulatory effect of this gene in
pancreatic cancer. Subsequent detailed characterization demonstrated a strong
overexpression of CFL1 in PDAC on mRNA as well as on protein level. RNAi-mediated
knockdown of CFL1 in 4 different cell lines led to a significant inhibition of cell growth
whereas apoptotic cascades were not affected. Furthermore, a significant decrease of
anchorage-independent growth was observed in soft agar assays. Flow cytometric
analyses indicated an attenuation of the G1 to S phase transition.
Aside from its growth-regulatory function, migration assays revealed an additional
inhibitory effect of CFL1 on cell motility identified in “time lapse” and wound healing
assays. In contrast to this, a directional migration analyzed in Boyden Chamber
analyses remained unaffected.
Based on these data and the current state of the literature, the results of this study
present for the first time a direct growth-regulatory effect of CFL1 in pancreatic cancer.
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Dates
Created: 2013Issued: 2013-11-25Updated: 2013-11-25
Faculty
Medizin
Publisher
Philipps-Universität Marburg
Language
ger
Data types
DoctoralThesis
Keywords
pancreatic adenocarcinomaPankreaskarzinomCofilin-1Hochdurchsatzanalyse
DFG-subjects
Bauchspeicheldrüsenkrebs
DDC-Numbers
500
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Kirchhoff, Sandra (1045692727): Zellbasierte Hochdurchsatzanalysen identifizieren eine zentrale, wachstumsregulierende Rolle von Cofilin-1 im humanen Pankreaskarzinom. : Philipps-Universität Marburg 2013-11-25. DOI: https://doi.org/10.17192/z2013.0666.
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This item has been published with the following license: In Copyright