Extensive microRNA-mediated crosstalk between lncRNAs and ...
Contribution of non-coding genome to susceptibility at low doses · 2013. 11. 12. · Long...
1
Project N° 323216 Project partners www.helmholtz-muenchen.de/isb/darkrisk The photos are original scans by the Belgrade Historical Archives. A young patient receiving a dose of radiation during Tinea Capitis treatment (a); manual epilation of remaining hair after x-ray treatment (b); children after epilation and irradiation completed (c); children with the zinc hat (d). Initial position Radiation epilation (RE) was the mainstay of therapy for Tinea capitis in the first half of the 20 th century. RE continues to have repercussions today due to the large number of children treated by this technique over 60 years ago and who may still develop radiation induced brain tumours. Radiation epilation was rapidly adopted and became the treatment of choice wherever it was available. Tinea capitis was endemic and considered a major health hazard in several countries. A child affected by ringworm was isolated and socially stigmatised and would often miss school for peri- ods of a year or more. The epidemiological study of Tinea capitis patients has proven useful for the study of carcinogenic effects of low dose radia- tion. In Dark.Risk we will develop a new epidemiological cohort based upon the ~ 25.000 cases irradiated in childhood that we will document to create the Serbian Registry of Tinaea Capitis Children (SRTCC). Unlike many other epi- demiological cohorts SRTCC can be used to collect biological materials for molecular epidemiological studies. Summary Dark.Risk will prepare the way to use SRTCC biomaterials to quantify the contribution of individual differences in sensitivity to risk from low doses of ionizing radiation. The focus will be on a novel and unique process to identify individual differences. Until recently 95% of the human genome was considered to be non-translated “junk" DNA. However, almost all of this “dark matter” is actively transcribed into a vareity of non-coding RNA species (ncRNAs) that regulate interactions with the external environment, such as repressing retrotransposons, regulating gene transcription responses to stress, governing chromatic remodeling and coordinating the translation of mRNAs. The ncRNAs play an essential role in coordinating critical cellular responses to radiation such as maintenance of genomic stability, senescence, apoptosis and survival. If you are one of the 50 000 children The newspaper advertisement calling for patients treated during the Yugoslavian Tinea Capitis campaign to contact the research team. [email protected] Ako ste jedno od 50,000 dece koja su tokom 1950-ih godina bila lečena od mikoze glave postupkom stavljanja Lajmove ("gipsane") kape ŽELELI BISMO DA STUPIMO U KONTAKT SA VAMA!! Pokušavamo da obnovimo uspomene na ovo važno poglavlje iz istorije naše medicine. Vaša iskustva bi nam bila dragocena. Ukoliko ste u mogu ćnosti, molimo Vas da nam se javite. tel: (020) 31-47-22/120 mob: (064) 20-58-799 tel: (011) 20-67-812, 25-63-269 mob: (064) 16 39 702 [email protected] Kontakt Beograd: dr Goran Ševo Kontakt Novi Pazar: dr Safet Ganić c a b d Epidemiological study and clinical cooperation Long non-coding RNAs (lncRNAs) screening miRNA target analysis New targets in radiation response Non-coding RNAs in radiation response Irradiation - or + 2D / 3D cellular model response analysis Mouse models miRNAs as indicator of irradiation Contribution of non-coding genome to susceptibility at low doses Dark.Risk consortium First Results Non-coding RNA expression profiling in human samples following low dose irradiation. A: Quality control through probe hybridization verification at all corners of an array; B: Volcano plot representing differentially expressed non-coding RNAs (red) identified through fold change filtering between two samples (x-axis: non-irradiated cells versus y-axis: 2.5 Gy irradiation); C: Heat-Map and hierarchical clustering of differentially expressed non-coding RNA in human samples (3 cell lines as indicated, 4 hours following non-irradiation or 2.5Gy irradiation). High and low relative ncRNA expression indicated in red and blue respectively with additional colour levels reflective of intermediate gene expression levels; D: Scatter-plot demonstrating ncRNA expression variation between comparison groups. Green lines represent fold change (1.5 and 2 fold). Quantile normalisation and data filtering performed using GeneSpring GX v11.5.1 (Agilent Technologies). Differentially expressed lncRNAs were identified following quantile normalization, fold change filter processing (GeneSpring) with sample and control (sham irradiated) comparison. Of the seven regulated lncRNAs that we subsequently selected for validation, all exhibited the same up- or down-regulation following irradiation in each of the three cell lines. Consequently an lncRNA that shows strong up-regulation in HUVEC cells was also found up-regulated in T47D and U2OS cells, and vice versa. This is the first indication that a given radiation- regulated lncRNA has a defined role across different cell lines during the radiation response. Such a result is a strong contrast to the situation observed for miRNAs, which exhibit highly cell-type specific changes in expression. Mean regulations of more than 2-fold upregulation were found for the lncRNAs PANDA and linc-p21. Significantly altered lncRNAs as- sociated with genes involved in cancer were selected for Taqman real-time RT-PCR verification in T47D and MDA-MB-361 breast cancer cell lines 4hr and 24hr after 0.25Gy and 2.5Gy irradiation. Although many long non-coding RNAs (lncRNAs) are deregulated, the characterisation of the majority still remains unresolved. The genomic context of lncRNAs can assist in predicting their functional role. One previously uncharacterised lncRNA (abbreviated rad-lncRNA1) was highly expressed in T47D and MDA-MB-361 cells 24hr after low dose (0.25Gy) irradiation exposure. These results suggest a potential role of lncRNAs in the regulation of specific gene ex- pression with implications for epigenetic control of low dose radiation effects.
Transcript of Contribution of non-coding genome to susceptibility at low doses · 2013. 11. 12. · Long...
Project N° 323216
Project partners www.helmholtz-muenchen.de/isb/darkrisk
The photos are original scans by the Belgrade Historical Archives.
A young patient receiving a dose of radiation during Tinea Capitis treatment (a);
manual epilation of remaining hair after x-ray treatment (b); children after epilation and
irradiation completed (c); children with the zinc hat (d).
Initial positionRadiation epilation (RE) was the mainstay of therapy for Tinea capitis in the first half of the 20th century. RE continues to have repercussions today due to the large number of children treated by this technique over 60 years ago and who may still develop radiation induced brain tumours. Radiation epilation was rapidly adopted and became the treatment of choice wherever it was available. Tinea capitis was endemic and considered a major health hazard in several countries. A child affected by ringworm was isolated and socially stigmatised and would often miss school for peri-ods of a year or more. The epidemiological study of Tinea capitis patients has proven useful for the study of carcinogenic effects of low dose radia-tion.
In Dark.Risk we will develop a new epidemiological cohort based upon the ~ 25.000 cases irradiated in childhood that we will document to create the Serbian Registry of Tinaea Capitis Children (SRTCC). Unlike many other epi-demiological cohorts SRTCC can be used to collect biological materials for molecular epidemiological studies.
SummaryDark.Risk will prepare the way to use SRTCC biomaterials to quantify the contribution of individual differences in sensitivity to risk from low doses of ionizing radiation. The focus will be on a novel and unique process to identify individual differences. Until recently 95% of the human genome was considered to be non-translated “junk" DNA. However, almost all of this “dark matter” is actively transcribed into a vareity of non-coding RNA species (ncRNAs) that regulate interactions with the external environment, such as repressing retrotransposons, regulating gene transcription responses to stress, governing chromatic remodeling and coordinating the translation of mRNAs. The ncRNAs play an essential role in coordinating critical cellular responses to radiation such as maintenance of genomic stability, senescence, apoptosis and survival.
If you are one of the 50 000 childrenThe newspaper advertisement calling for patients
treated during the Yugoslavian Tinea Capitis campaign to contact the
research team.
Ako ste jedno od 50,000 dece
koja su tokom 1950-ih godina bila lečena od mikoze glavepostupkom stavljanja Lajmove ("gipsane") kape
ŽELELI BISMO DA STUPIMO U KONTAKT SA VAMA!!Pokušavamo da obnovimo uspomene na ovo važno poglavlje iz istorije naše
medicine. Vaša iskustva bi nam bila dragocena.Ukoliko ste u mogu ćnosti, molimo Vas da nam se javite.
tel: (020) 31-47-22/120mob: (064) 20-58-799
tel: (011) 20-67-812, 25-63-269 mob: (064) 16 39 [email protected]
Kontakt Beograd: dr Goran Ševo Kontakt Novi Pazar: dr Safet Ganić
c
a b
d
Epidemiological study and clinical cooperation
Long non-coding RNAs(lncRNAs) screening
miRNA target analysisNew targets in radiation response
Non-coding RNAsin radiation response
Irradiation- or + 2D / 3D cellular model
response analysis
Mouse models
miRNAs as indicatorof irradiation
Contribution of non-coding genome to susceptibility at low dosesDark.Risk consortium
First ResultsNon-coding RNA expression profiling in
human samples following low dose irradiation.
A: Quality control through probe hybridization
verification at all corners of an array; B:
Volcano plot representing differentially expressed non-coding RNAs (red) identified through fold change filtering between two
samples (x-axis: non-irradiated cells versus y-axis: 2.5 Gy irradiation);
C: Heat-Map and hierarchical clustering of
differentially expressed non-coding RNA in human samples (3 cell lines as indicated, 4 hours
following non-irradiation or 2.5Gy irradiation). High and low relative ncRNA expression
indicated in red and blue respectively with additional colour levels reflective of intermediate gene expression levels;
D: Scatter-plot demonstrating ncRNA expression variation between comparison groups. Green
lines represent fold change (1.5 and 2 fold). Quantile normalisation and data filtering
performed using GeneSpring GX v11.5.1 (Agilent Technologies).
Differentially expressed lncRNAs were identified following quantile normalization, fold change filter processing (GeneSpring) with sample and control (sham irradiated) comparison. Of the seven regulated lncRNAs that we subsequently selected for validation, all exhibited the same up- or down-regulation following irradiation in each of the three cell lines.
Consequently an lncRNA that shows strong up-regulation in HUVEC cells was also found up-regulated in T47D and U2OS cells, and vice versa. This is the first indication that a given radiation-regulated lncRNA has a defined role across different cell lines during the radiation response. Such a result is a strong contrast to the situation observed for miRNAs, which exhibit highly cell-type specific changes in expression.
Mean regulations of more than 2-fold upregulation were found for the lncRNAs PANDA and linc-p21. Significantly altered lncRNAs as-sociated with genes involved in cancer were selected for Taqman real-time RT-PCR verification in T47D and MDA-MB-361 breast cancer cell lines 4hr and 24hr after 0.25Gy and 2.5Gy irradiation.
Although many long non-coding RNAs (lncRNAs) are deregulated, the characterisation of the majority still remains unresolved. The genomic context of lncRNAs can assist in predicting their functional role. One previously uncharacterised lncRNA (abbreviated rad-lncRNA1) was highly expressed in T47D and MDA-MB-361 cells 24hr after low dose (0.25Gy) irradiation exposure. These results suggest a potential role of lncRNAs in the regulation of specific gene ex-pression with implications for epigenetic control of low dose radiation effects.
