Get 20M+ Full-Text Papers For Less Than $1.50/day. Start a 14-Day Trial for You or Your Team.

Learn More →

A three‐dimensional colocalization RNA interference screening platform to elucidate the alternative lengthening of telomeres pathway

A three‐dimensional colocalization RNA interference screening platform to elucidate the... A high‐content colocalization RNA interference screen based on automatic three‐color confocal fluorescence microscopy was developed to analyze the alternative lengthening of telomeres (ALT) pathway. Via this pathway telomerase‐negative cancer cells can maintain their telomeres and with it their unlimited proliferative potential. A hallmark of ALT cells is the colocalization of promyelocytic leukemia (PML) nuclear bodies with telomeres to form ALT‐associated PML nuclear bodies (APBs). In our screen, the presence of APBs was used as a marker to identify proteins required for the ALT mechanism. A cell‐based assay and an automatic confocal image acquisition procedure were established. Using automatic image analysis based on 3D parametric intensity models to identify APBs, we conducted an unbiased and quantitative analysis of nine different candidate genes. A comparison with the literature and manual analysis of the gene knockdown demonstrates the reliability of our approach. It extends the available repertoire of high‐content screening to studies of cellular colocalizations and allows the identification of candidate genes for the ALT mechanism that represent possible targets for cancer therapy. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Biotechnology Journal Wiley

A three‐dimensional colocalization RNA interference screening platform to elucidate the alternative lengthening of telomeres pathway

Loading next page...
 
/lp/wiley/a-three-dimensional-colocalization-rna-interference-screening-platform-OQbVB0mWKc

References (39)

Publisher
Wiley
Copyright
Copyright © 2012 Wiley Subscription Services, Inc., A Wiley Company
ISSN
1860-6768
eISSN
1860-7314
DOI
10.1002/biot.201000474
pmid
21648092
Publisher site
See Article on Publisher Site

Abstract

A high‐content colocalization RNA interference screen based on automatic three‐color confocal fluorescence microscopy was developed to analyze the alternative lengthening of telomeres (ALT) pathway. Via this pathway telomerase‐negative cancer cells can maintain their telomeres and with it their unlimited proliferative potential. A hallmark of ALT cells is the colocalization of promyelocytic leukemia (PML) nuclear bodies with telomeres to form ALT‐associated PML nuclear bodies (APBs). In our screen, the presence of APBs was used as a marker to identify proteins required for the ALT mechanism. A cell‐based assay and an automatic confocal image acquisition procedure were established. Using automatic image analysis based on 3D parametric intensity models to identify APBs, we conducted an unbiased and quantitative analysis of nine different candidate genes. A comparison with the literature and manual analysis of the gene knockdown demonstrates the reliability of our approach. It extends the available repertoire of high‐content screening to studies of cellular colocalizations and allows the identification of candidate genes for the ALT mechanism that represent possible targets for cancer therapy.

Journal

Biotechnology JournalWiley

Published: Jan 1, 2012

Keywords: ; ; ; ;

There are no references for this article.