TY - JOUR
T1 - A prototypical non-malignant epithelial model to study genome dynamics and concurrently monitor micro-RNAs and proteins in situ during oncogene-induced senescence
AU - Komseli, Eirini Stavroula
AU - Pateras, Ioannis S.
AU - Krejsgaard, Thorbjørn
AU - Stawiski, Konrad
AU - Rizou, Sophia V.
AU - Polyzos, Alexander
AU - Roumelioti, Fani Marlen
AU - Chiourea, Maria
AU - Mourkioti, Ioanna
AU - Paparouna, Eleni
AU - Zampetidis, Christos P.
AU - Gumeni, Sentiljana
AU - Trougakos, Ioannis P.
AU - Pefani, Dafni Eleftheria
AU - O'Neill, Eric
AU - Gagos, Sarantis
AU - Eliopoulos, Aristides G.
AU - Fendler, Wojciech
AU - Chowdhury, Dipanjan
AU - Bartek, Jiri
AU - Gorgoulis, Vassilis G.
PY - 2018/1
Y1 - 2018/1
N2 - Background: Senescence is a fundamental biological process implicated in various pathologies, including cancer. Regarding carcinogenesis, senescence signifies, at least in its initial phases, an anti-tumor response that needs to be circumvented for cancer to progress. Micro-RNAs, a subclass of regulatory, non-coding RNAs, participate in senescence regulation. At the subcellular level micro-RNAs, similar to proteins, have been shown to traffic between organelles influencing cellular behavior. The differential function of micro-RNAs relative to their subcellular localization and their role in senescence biology raises concurrent in situ analysis of coding and non-coding gene products in senescent cells as a necessity. However, technical challenges have rendered in situ co-detection unfeasible until now. Methods: In the present report we describe a methodology that bypasses these technical limitations achieving for the first time simultaneous detection of both a micro-RNA and a protein in the biological context of cellular senescence, utilizing the new commercially available SenTraGorTM compound. The method was applied in a prototypical human non-malignant epithelial model of oncogene-induced senescence that we generated for the purposes of the study. For the characterization of this novel system, we applied a wide range of cellular and molecular techniques, as well as high-throughput analysis of the transcriptome and micro-RNAs. Results: This experimental setting has three advantages that are presented and discussed: i) it covers a "gap" in the molecular carcinogenesis field, as almost all corresponding in vitro models are fibroblast-based, even though the majority of neoplasms have epithelial origin, ii) it recapitulates the precancerous and cancerous phases of epithelial tumorigenesis within a short time frame under the light of natural selection and iii) it uses as an oncogenic signal, the replication licensing factor CDC6, implicated in both DNA replication and transcription when over-expressed, a characteristic that can be exploited to monitor RNA dynamics. Conclusions: Consequently, we demonstrate that our model is optimal for studying the molecular basis of epithelial carcinogenesis shedding light on the tumor-initiating events. The latter may reveal novel molecular targets with clinical benefit. Besides, since this method can be incorporated in a wide range of low, medium or high-throughput image-based approaches, we expect it to be broadly applicable.
AB - Background: Senescence is a fundamental biological process implicated in various pathologies, including cancer. Regarding carcinogenesis, senescence signifies, at least in its initial phases, an anti-tumor response that needs to be circumvented for cancer to progress. Micro-RNAs, a subclass of regulatory, non-coding RNAs, participate in senescence regulation. At the subcellular level micro-RNAs, similar to proteins, have been shown to traffic between organelles influencing cellular behavior. The differential function of micro-RNAs relative to their subcellular localization and their role in senescence biology raises concurrent in situ analysis of coding and non-coding gene products in senescent cells as a necessity. However, technical challenges have rendered in situ co-detection unfeasible until now. Methods: In the present report we describe a methodology that bypasses these technical limitations achieving for the first time simultaneous detection of both a micro-RNA and a protein in the biological context of cellular senescence, utilizing the new commercially available SenTraGorTM compound. The method was applied in a prototypical human non-malignant epithelial model of oncogene-induced senescence that we generated for the purposes of the study. For the characterization of this novel system, we applied a wide range of cellular and molecular techniques, as well as high-throughput analysis of the transcriptome and micro-RNAs. Results: This experimental setting has three advantages that are presented and discussed: i) it covers a "gap" in the molecular carcinogenesis field, as almost all corresponding in vitro models are fibroblast-based, even though the majority of neoplasms have epithelial origin, ii) it recapitulates the precancerous and cancerous phases of epithelial tumorigenesis within a short time frame under the light of natural selection and iii) it uses as an oncogenic signal, the replication licensing factor CDC6, implicated in both DNA replication and transcription when over-expressed, a characteristic that can be exploited to monitor RNA dynamics. Conclusions: Consequently, we demonstrate that our model is optimal for studying the molecular basis of epithelial carcinogenesis shedding light on the tumor-initiating events. The latter may reveal novel molecular targets with clinical benefit. Besides, since this method can be incorporated in a wide range of low, medium or high-throughput image-based approaches, we expect it to be broadly applicable.
KW - Cancer
KW - CDC6
KW - DNA damage response
KW - In situ hybridization
KW - Micro-RNAs
KW - Oncogene-induced senescence
KW - R loops
KW - RDNA
KW - Replication stress
KW - SenTraGorTM
U2 - 10.1186/s12864-017-4375-1
DO - 10.1186/s12864-017-4375-1
M3 - Journal article
C2 - 29321003
AN - SCOPUS:85040315831
SN - 1471-2164
VL - 19
JO - BMC Genomics
JF - BMC Genomics
IS - 1
M1 - 37
ER -
