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S-Nitrosylation in Organs of Mice Exposed to Low or High Doses of γ-Rays: The Modulating Effect of Iodine Contrast Agent at a Low Radiation Dose.

Nicolas F, Wu C, Bukhari S, de Toledo SM, Li H, Shibata M, Azzam EI - Proteomes (2015)

Bottom Line: The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins.To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays.The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Health Informatics, Rutgers School of Health Related Professions, Newark, NJ 07107, USA; nicolafa@shrp.rutgers.edu.

ABSTRACT

The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins. The dysregulation of normal S-nitrosylation contributes to degenerative conditions and to cancer. To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays. The dose of radiation was delivered to the whole body in the presence or absence of iopamidol, an iodinated contrast agent used during radiological examinations. To investigate whether similar or distinct nitrosylation patterns are induced following high-dose irradiation, mice were exposed in parallel to acute 4 Gy of (137)Cs γ rays. Analysis of modulated S-nitrosothiols (SNO-proteins) in freshly-harvested organs of animals sacrificed 13 days after irradiation revealed radiation dose- and contrast agent-dependent changes. The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

No MeSH data available.


Related in: MedlinePlus

Modulation of the S-nitrosoproteome in brain of mice exposed to low- or high-dose 137Cs γ-rays. Changes in the levels of nitrosylated proteins detected by mass spectrometry analyses in brains of C57Bl/6J mice exposed 13 days earlier to acute doses of 0.1 Gy or 4 Gy of 137Cs γ rays. The X-axis is the relative level of nitrosylated proteins after a given radiation dose compared to the control (numbers shown represent the fold-change of nitrosylated proteins detected and expressed in the log2 ratio, where x indicates the interval between x − 0.1 and x + 0.1). The Y-axis indicates the frequency or number of occurrences in terms of the number of proteins. Blue and red bars indicate observations at 0.1 Gy and 4 Gy, respectively, compared to the control.
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Figure 2: Modulation of the S-nitrosoproteome in brain of mice exposed to low- or high-dose 137Cs γ-rays. Changes in the levels of nitrosylated proteins detected by mass spectrometry analyses in brains of C57Bl/6J mice exposed 13 days earlier to acute doses of 0.1 Gy or 4 Gy of 137Cs γ rays. The X-axis is the relative level of nitrosylated proteins after a given radiation dose compared to the control (numbers shown represent the fold-change of nitrosylated proteins detected and expressed in the log2 ratio, where x indicates the interval between x − 0.1 and x + 0.1). The Y-axis indicates the frequency or number of occurrences in terms of the number of proteins. Blue and red bars indicate observations at 0.1 Gy and 4 Gy, respectively, compared to the control.

Mentions: Exposures to ionizing radiation even at doses similar to those received during CT scans have been suggested to increase the risk of developing brain tumors [39]. Interestingly, the data in Figure 1 revealed differential changes in levels of SNO proteins (e.g., in the 40–60-kDa range) in brains of mice exposed to 0.1 or 4 Gy. To gain insight into the modulated proteins, we performed mass spectrometry (MS) measurements of S-nitrosylated proteins and peptides in lysates of whole brain of mice exposed 13 days earlier to 0, 0.1 or 4 Gy. To this end, equal mass aliquots of protein lysates from five different mice in each group were combined and submitted for the biotin switch assay and MS analyses. Spectral counts (Supplementary Table S1) revealed that 377 SNO proteins were detectable in at least one condition among the three conditions examined (0 Gy, 0.1 Gy and 4 Gy). Relative to the control, the majority of proteins from irradiated samples exhibited very small changes in nitrosylation, with most belonging to less than a two-fold increase or decrease (between −1 and +1 in the log2 ratio) (Figure 2). It is noteworthy that in the 4 Gy group, a large number of proteins (77 proteins) showed almost no change in the nitrosylation level from the control, with ~86% of the proteins showing less than a ±2-fold change (i.e., within ±1 in the log2 ratio). Interestingly, a greater number of proteins showed decreased nitrosylation after low-dose radiation exposure, which is similar to what has been found in the immunoprecipitation experiments (Figure 1). Interestingly, the Western blot analyses in Figure 3, showing expression levels of both nitrosylated and total ATPG (ATP synthase subunit γ, mitochondrial), indicate that at least for some proteins, the change in level of nitrosylation is not due to the modulation of total protein expression level, but rather to regulation of S-nitrosylation.


S-Nitrosylation in Organs of Mice Exposed to Low or High Doses of γ-Rays: The Modulating Effect of Iodine Contrast Agent at a Low Radiation Dose.

Nicolas F, Wu C, Bukhari S, de Toledo SM, Li H, Shibata M, Azzam EI - Proteomes (2015)

Modulation of the S-nitrosoproteome in brain of mice exposed to low- or high-dose 137Cs γ-rays. Changes in the levels of nitrosylated proteins detected by mass spectrometry analyses in brains of C57Bl/6J mice exposed 13 days earlier to acute doses of 0.1 Gy or 4 Gy of 137Cs γ rays. The X-axis is the relative level of nitrosylated proteins after a given radiation dose compared to the control (numbers shown represent the fold-change of nitrosylated proteins detected and expressed in the log2 ratio, where x indicates the interval between x − 0.1 and x + 0.1). The Y-axis indicates the frequency or number of occurrences in terms of the number of proteins. Blue and red bars indicate observations at 0.1 Gy and 4 Gy, respectively, compared to the control.
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Related In: Results  -  Collection

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Figure 2: Modulation of the S-nitrosoproteome in brain of mice exposed to low- or high-dose 137Cs γ-rays. Changes in the levels of nitrosylated proteins detected by mass spectrometry analyses in brains of C57Bl/6J mice exposed 13 days earlier to acute doses of 0.1 Gy or 4 Gy of 137Cs γ rays. The X-axis is the relative level of nitrosylated proteins after a given radiation dose compared to the control (numbers shown represent the fold-change of nitrosylated proteins detected and expressed in the log2 ratio, where x indicates the interval between x − 0.1 and x + 0.1). The Y-axis indicates the frequency or number of occurrences in terms of the number of proteins. Blue and red bars indicate observations at 0.1 Gy and 4 Gy, respectively, compared to the control.
Mentions: Exposures to ionizing radiation even at doses similar to those received during CT scans have been suggested to increase the risk of developing brain tumors [39]. Interestingly, the data in Figure 1 revealed differential changes in levels of SNO proteins (e.g., in the 40–60-kDa range) in brains of mice exposed to 0.1 or 4 Gy. To gain insight into the modulated proteins, we performed mass spectrometry (MS) measurements of S-nitrosylated proteins and peptides in lysates of whole brain of mice exposed 13 days earlier to 0, 0.1 or 4 Gy. To this end, equal mass aliquots of protein lysates from five different mice in each group were combined and submitted for the biotin switch assay and MS analyses. Spectral counts (Supplementary Table S1) revealed that 377 SNO proteins were detectable in at least one condition among the three conditions examined (0 Gy, 0.1 Gy and 4 Gy). Relative to the control, the majority of proteins from irradiated samples exhibited very small changes in nitrosylation, with most belonging to less than a two-fold increase or decrease (between −1 and +1 in the log2 ratio) (Figure 2). It is noteworthy that in the 4 Gy group, a large number of proteins (77 proteins) showed almost no change in the nitrosylation level from the control, with ~86% of the proteins showing less than a ±2-fold change (i.e., within ±1 in the log2 ratio). Interestingly, a greater number of proteins showed decreased nitrosylation after low-dose radiation exposure, which is similar to what has been found in the immunoprecipitation experiments (Figure 1). Interestingly, the Western blot analyses in Figure 3, showing expression levels of both nitrosylated and total ATPG (ATP synthase subunit γ, mitochondrial), indicate that at least for some proteins, the change in level of nitrosylation is not due to the modulation of total protein expression level, but rather to regulation of S-nitrosylation.

Bottom Line: The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins.To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays.The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Health Informatics, Rutgers School of Health Related Professions, Newark, NJ 07107, USA; nicolafa@shrp.rutgers.edu.

ABSTRACT

The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins. The dysregulation of normal S-nitrosylation contributes to degenerative conditions and to cancer. To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays. The dose of radiation was delivered to the whole body in the presence or absence of iopamidol, an iodinated contrast agent used during radiological examinations. To investigate whether similar or distinct nitrosylation patterns are induced following high-dose irradiation, mice were exposed in parallel to acute 4 Gy of (137)Cs γ rays. Analysis of modulated S-nitrosothiols (SNO-proteins) in freshly-harvested organs of animals sacrificed 13 days after irradiation revealed radiation dose- and contrast agent-dependent changes. The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

No MeSH data available.


Related in: MedlinePlus