Page 153 - 2014 Printable Abstract Book
P. 153
knockdown of Apollo, but not the inhibition of MRE11 with Mirin, also results in a decrease in the
frequency of small deletions at both interstitial and telomeric DSBs. These results show for the first time
that Apollo is involved in the formation of mutations at DSBs, causing small deletions during C-NHEJ at
both interstitial and telomeric DSBs, and along with MRE11, large deletions and GCRs involving A-NJEJ at
DSBs near telomeres. DSB-induced mutations can therefore occur through two separate pathways, the
KU70/86 protein pathway involved in C-NHEJ, and by processing in the MRN/ATM protein pathway
followed by A-NHEJ. Importantly, our results show that the decrease in distal NHEJ near telomeres is due
to excessive processing and not a defect in C-NHEJ, which explains the fact that the frequency of small
deletions is similar at interstitial and telomeric DSBs. Based on our results, we propose that the sensitivity
of DSBs near telomeres is a result of the inappropriate processing of DSBs near telomeres through the
MRN/ATM pathway because they are mistaken for telomeres, leading to large deletions, GCRs, and
chromosome instability.
(PS2-30) Induction of chromosomal aberrations at fluences of less than one HZE particle per cell nucleus.
1
2
3
1
1
Megumi Hada ; Lori J. Chappell ; Minli Wang ; Kerry A. George ; and Francis Cucinotta,
1
2
Universities Space Research Association, Houston, TX ; Wyle, Houston, TX ; and University of Nevada, Las
Vegas, NV
3
The assumption of a linear dose response used to describe the biological effects of high LET
radiation is fundamental in radiation protection methodologies. We investigated the dose response for
chromosomal aberrations for exposures corresponding to less than one particle traversal per cell nucleus
by high energy and charge (HZE) nuclei. Human fibroblast and lymphocyte cells where irradiated with
several low doses of <0.1 Gy, and several higher doses of up to 1 Gy with O (77 keV/µm), Si (99 keV/µm),
Fe (175 keV/µm), Fe (195 keV/µm) or Fe (240 keV/µm) particles. Chromosomal aberrations at first mitosis
were scored using fluorescence in situ hybridization (FISH) with chromosome specific paints for
chromosomes 1, 2 and 4 and DAPI staining of background chromosomes. Non-linear regression models
were used to evaluate possible linear and non-linear dose response models based on these data. Dose
responses for simple exchanges for human fibroblast irradiated under confluent culture conditions were
best fit by non-linear models motivated by a non-targeted effect (NTE). Best fits for the dose response
data for human lymphocytes irradiated in blood tubes were a NTE model for O and a linear response
model fit best for Si and Fe particles. Additional evidence for NTE were found in low dose experiments
measuring γH2AX foci, a marker of double strand breaks (DSB), and split-dose experiments with human
fibroblasts. Our results suggest that simple exchanges in normal human fibroblasts have an important NTE
contribution at low particle fluence. The current and prior experimental studies provide important
evidence against the linear dose response assumption used in radiation protection for HZE particles and
other high LET radiation at the relevant range of low doses.
151 | P a g e
frequency of small deletions at both interstitial and telomeric DSBs. These results show for the first time
that Apollo is involved in the formation of mutations at DSBs, causing small deletions during C-NHEJ at
both interstitial and telomeric DSBs, and along with MRE11, large deletions and GCRs involving A-NJEJ at
DSBs near telomeres. DSB-induced mutations can therefore occur through two separate pathways, the
KU70/86 protein pathway involved in C-NHEJ, and by processing in the MRN/ATM protein pathway
followed by A-NHEJ. Importantly, our results show that the decrease in distal NHEJ near telomeres is due
to excessive processing and not a defect in C-NHEJ, which explains the fact that the frequency of small
deletions is similar at interstitial and telomeric DSBs. Based on our results, we propose that the sensitivity
of DSBs near telomeres is a result of the inappropriate processing of DSBs near telomeres through the
MRN/ATM pathway because they are mistaken for telomeres, leading to large deletions, GCRs, and
chromosome instability.
(PS2-30) Induction of chromosomal aberrations at fluences of less than one HZE particle per cell nucleus.
1
2
3
1
1
Megumi Hada ; Lori J. Chappell ; Minli Wang ; Kerry A. George ; and Francis Cucinotta,
1
2
Universities Space Research Association, Houston, TX ; Wyle, Houston, TX ; and University of Nevada, Las
Vegas, NV
3
The assumption of a linear dose response used to describe the biological effects of high LET
radiation is fundamental in radiation protection methodologies. We investigated the dose response for
chromosomal aberrations for exposures corresponding to less than one particle traversal per cell nucleus
by high energy and charge (HZE) nuclei. Human fibroblast and lymphocyte cells where irradiated with
several low doses of <0.1 Gy, and several higher doses of up to 1 Gy with O (77 keV/µm), Si (99 keV/µm),
Fe (175 keV/µm), Fe (195 keV/µm) or Fe (240 keV/µm) particles. Chromosomal aberrations at first mitosis
were scored using fluorescence in situ hybridization (FISH) with chromosome specific paints for
chromosomes 1, 2 and 4 and DAPI staining of background chromosomes. Non-linear regression models
were used to evaluate possible linear and non-linear dose response models based on these data. Dose
responses for simple exchanges for human fibroblast irradiated under confluent culture conditions were
best fit by non-linear models motivated by a non-targeted effect (NTE). Best fits for the dose response
data for human lymphocytes irradiated in blood tubes were a NTE model for O and a linear response
model fit best for Si and Fe particles. Additional evidence for NTE were found in low dose experiments
measuring γH2AX foci, a marker of double strand breaks (DSB), and split-dose experiments with human
fibroblasts. Our results suggest that simple exchanges in normal human fibroblasts have an important NTE
contribution at low particle fluence. The current and prior experimental studies provide important
evidence against the linear dose response assumption used in radiation protection for HZE particles and
other high LET radiation at the relevant range of low doses.
151 | P a g e
