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Molecular Mechanisms for High Hydrostatic Pressure-Induced Wing Mutagenesis in Drosophila melanogaster.

Wang H, Wang K, Xiao G, Ma J, Wang B, Shen S, Fu X, Zou G, Zou B - Sci Rep (2015)

Bottom Line: Abnormal wing phenotypes of D. melanogaster induced by HHP were used to investigate the mutagenic mechanisms of HHP on organism.Thus 285 differentially expressed genes associated with wing mutations were identified using Affymetrix Drosophila Genome Array 2.0 and verified with RT-PCR.This study revealed the mutagenic mechanisms of HHP-induced mutagenesis in D. melanogaster and provided a new model for the study of evolution on organisms.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Superhard Materials, Jilin University, Changchun, 130012, P. R. China.

ABSTRACT
Although High hydrostatic pressure (HHP) as an important physical and chemical tool has been increasingly applied to research of organism, the response mechanisms of organism to HHP have not been elucidated clearly thus far. To identify mutagenic mechanisms of HHP on organisms, here, we treated Drosophila melanogaster (D. melanogaster) eggs with HHP. Approximately 75% of the surviving flies showed significant morphological abnormalities from the egg to the adult stages compared with control flies (p < 0.05). Some eggs displayed abnormal chorionic appendages, some larvae were large and red, and some adult flies showed wing abnormalities. Abnormal wing phenotypes of D. melanogaster induced by HHP were used to investigate the mutagenic mechanisms of HHP on organism. Thus 285 differentially expressed genes associated with wing mutations were identified using Affymetrix Drosophila Genome Array 2.0 and verified with RT-PCR. We also compared wing development-related central genes in the mutant flies with control flies using DNA sequencing to show two point mutations in the vestigial (vg) gene. This study revealed the mutagenic mechanisms of HHP-induced mutagenesis in D. melanogaster and provided a new model for the study of evolution on organisms.

No MeSH data available.


Related in: MedlinePlus

Model of the molecular mechanism for the wing mutagenesis induced by HHP treatment in D. melanogaster.HHP stress induces 285 differentially expressed genes in the mutants. Among them, 6 genes, including sca, hth, osa, dad, ilk, and stck have a direct relationship with wing development. In addition, Numb, tom, sca, bun, and nedd4 genes have an indirectly role in D. melanogaster wing development through Notch signaling pathways. The sca gene is common to both. The Notch signaling pathway also affects D. melanogaster wing phenotype via the vg target gene, and HHP stress induces two point mutations in the vg gene. The two mutations are synonymous mutations. Furthermore, the four bases deletion in the exon 1 of vg gene resulting in the change of reading frame and the consequent failure to generate functional protein. An insertion sequence between the second and third exon of vg indirectly affect the function of protein. The coordinated interaction of these abnormal genes is crucial for the wing mutagenesis induced by HHP treatment in D. melanogaster.
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f5: Model of the molecular mechanism for the wing mutagenesis induced by HHP treatment in D. melanogaster.HHP stress induces 285 differentially expressed genes in the mutants. Among them, 6 genes, including sca, hth, osa, dad, ilk, and stck have a direct relationship with wing development. In addition, Numb, tom, sca, bun, and nedd4 genes have an indirectly role in D. melanogaster wing development through Notch signaling pathways. The sca gene is common to both. The Notch signaling pathway also affects D. melanogaster wing phenotype via the vg target gene, and HHP stress induces two point mutations in the vg gene. The two mutations are synonymous mutations. Furthermore, the four bases deletion in the exon 1 of vg gene resulting in the change of reading frame and the consequent failure to generate functional protein. An insertion sequence between the second and third exon of vg indirectly affect the function of protein. The coordinated interaction of these abnormal genes is crucial for the wing mutagenesis induced by HHP treatment in D. melanogaster.

Mentions: D. melanogaster is a typical model organism for studying the response mechanisms of stress resistance. The present study took advantage of this model, by utilizing three molecular strategies, DNA microarray, RT-PCR and DNA sequencing, to identify potential causative genes underlying HHP-induced mutagenesis. With regard to the effect of HHP on organisms, previous experiments were performed on microorganisms and plants11. For example, Yayanos et al. reported that the synthesis of DNA, RNA, and protein was inhibited by HHP in living E. coli cells. Zhang et al. indicated that HHP impaired DNA strand integrity in rice. However, their results just limited the effect of HHP on biopolymer such as DNA, RNA, and protein and did not determine specific genes pertaining to HHP mutagenesis. By contrast, our experiments, using D. melanogaster as an animal model, provided further evidence for the mechanisms of the HHP-induced mutations in terms of the analysis of gene differential expression and DNA sequence variation. Specifically, we identified 285 differentially expressed genes in the mutants. We assigned these genes to the subcategories of molecular function, cellular component, and biological process. The 10 genes related to wing development were screened and verified using RT-PCR, with a remarkable concordance in the results. Among them, 6 genes, including sca, hth, osa, dad, Ilk, and stck, played important roles in wing development and directly induced the wing mutations in D. melanogaster. Cell signaling pathways regulate important physiological processes, such as cell proliferation, differentiation, growth, and development using various mechanisms141516. Our experimental results implicate multiple signaling pathways in the acquisition of HHP tolerance in D. melanogaster mutants. Among them, the Notch signaling pathway plays important roles in the wing development of D. melanogaster. Notch signaling is mediated by the down-regulated expression of Numb, Tom, and sca, and the up-regulated expression of bun and Nedd4 (Supplementary Table S1 and Supplementary Table S3). In general, the normal wing development in D. melanogaster requires the coordinated actions of several genes, such as vg, sd, Notch, and wg17. The vg gene in D. melanogaster not only plays a central role in wing development but is also a target of the Notch signaling pathway1819. In addition, vg expression occurs throughout the entire developing wing field20. Loss of vg results in wing development failures, and ectopic expression of vg leads to the development of ectopic wings21. For D. melanogaster, the vg gene is located on chromosome 2, and the complete sequence contains seven exons and eight introns. Our DNA sequencing results indicated that two point mutations occurred in exon 3 of vg gene, they were synonymous mutations. Synonymous mutations indirectly affect the function of proteins222324. Furthermore, the four bases deletion in the exon 1 of vg gene was done in D. melanogaster mutant, resulting in the change of reading frame and the consequent failure to generate functional protein. Generally, frameshift mutations are base deletions or additions within the coding region of a gene. As expected, because of potential problems with disturbing the reading frame, the entire set of triplets downstream of the deletion or addition is altered. In many cases, the deletion or addition results in the presence of in-frame termination sequences which stop the product. Frameshift mutations may therefore result in more severe phenotypic effects than do a number of the base changes which cause either silent or conservative mutations in protein products25. In addition, our DNA sequencing results showed an insertion sequence with 12 nucleotides (5′-CTCGCCCTGTCT-3′) between the second and third exons, namely, the junction of exon 3 and the intron 3 of vg gene. This insertion sequence was another incentive for abnormal wing development of D. melanogaster induced by HHP. The coordinated interaction of these ectopic expression genes and vg gene mutants is crucial for abnormal wing development in D. melanogaster induced by HHP treatment. However, how these genes induced by HHP regulate abnormal wing development of D. melanogaster is not well understood and needs further study. Our results suggest the model shown in Fig. 5 for the molecular mechanism of wing mutagenesis induced by HHP treatment in D. melanogaster.


Molecular Mechanisms for High Hydrostatic Pressure-Induced Wing Mutagenesis in Drosophila melanogaster.

Wang H, Wang K, Xiao G, Ma J, Wang B, Shen S, Fu X, Zou G, Zou B - Sci Rep (2015)

Model of the molecular mechanism for the wing mutagenesis induced by HHP treatment in D. melanogaster.HHP stress induces 285 differentially expressed genes in the mutants. Among them, 6 genes, including sca, hth, osa, dad, ilk, and stck have a direct relationship with wing development. In addition, Numb, tom, sca, bun, and nedd4 genes have an indirectly role in D. melanogaster wing development through Notch signaling pathways. The sca gene is common to both. The Notch signaling pathway also affects D. melanogaster wing phenotype via the vg target gene, and HHP stress induces two point mutations in the vg gene. The two mutations are synonymous mutations. Furthermore, the four bases deletion in the exon 1 of vg gene resulting in the change of reading frame and the consequent failure to generate functional protein. An insertion sequence between the second and third exon of vg indirectly affect the function of protein. The coordinated interaction of these abnormal genes is crucial for the wing mutagenesis induced by HHP treatment in D. melanogaster.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4597337&req=5

f5: Model of the molecular mechanism for the wing mutagenesis induced by HHP treatment in D. melanogaster.HHP stress induces 285 differentially expressed genes in the mutants. Among them, 6 genes, including sca, hth, osa, dad, ilk, and stck have a direct relationship with wing development. In addition, Numb, tom, sca, bun, and nedd4 genes have an indirectly role in D. melanogaster wing development through Notch signaling pathways. The sca gene is common to both. The Notch signaling pathway also affects D. melanogaster wing phenotype via the vg target gene, and HHP stress induces two point mutations in the vg gene. The two mutations are synonymous mutations. Furthermore, the four bases deletion in the exon 1 of vg gene resulting in the change of reading frame and the consequent failure to generate functional protein. An insertion sequence between the second and third exon of vg indirectly affect the function of protein. The coordinated interaction of these abnormal genes is crucial for the wing mutagenesis induced by HHP treatment in D. melanogaster.
Mentions: D. melanogaster is a typical model organism for studying the response mechanisms of stress resistance. The present study took advantage of this model, by utilizing three molecular strategies, DNA microarray, RT-PCR and DNA sequencing, to identify potential causative genes underlying HHP-induced mutagenesis. With regard to the effect of HHP on organisms, previous experiments were performed on microorganisms and plants11. For example, Yayanos et al. reported that the synthesis of DNA, RNA, and protein was inhibited by HHP in living E. coli cells. Zhang et al. indicated that HHP impaired DNA strand integrity in rice. However, their results just limited the effect of HHP on biopolymer such as DNA, RNA, and protein and did not determine specific genes pertaining to HHP mutagenesis. By contrast, our experiments, using D. melanogaster as an animal model, provided further evidence for the mechanisms of the HHP-induced mutations in terms of the analysis of gene differential expression and DNA sequence variation. Specifically, we identified 285 differentially expressed genes in the mutants. We assigned these genes to the subcategories of molecular function, cellular component, and biological process. The 10 genes related to wing development were screened and verified using RT-PCR, with a remarkable concordance in the results. Among them, 6 genes, including sca, hth, osa, dad, Ilk, and stck, played important roles in wing development and directly induced the wing mutations in D. melanogaster. Cell signaling pathways regulate important physiological processes, such as cell proliferation, differentiation, growth, and development using various mechanisms141516. Our experimental results implicate multiple signaling pathways in the acquisition of HHP tolerance in D. melanogaster mutants. Among them, the Notch signaling pathway plays important roles in the wing development of D. melanogaster. Notch signaling is mediated by the down-regulated expression of Numb, Tom, and sca, and the up-regulated expression of bun and Nedd4 (Supplementary Table S1 and Supplementary Table S3). In general, the normal wing development in D. melanogaster requires the coordinated actions of several genes, such as vg, sd, Notch, and wg17. The vg gene in D. melanogaster not only plays a central role in wing development but is also a target of the Notch signaling pathway1819. In addition, vg expression occurs throughout the entire developing wing field20. Loss of vg results in wing development failures, and ectopic expression of vg leads to the development of ectopic wings21. For D. melanogaster, the vg gene is located on chromosome 2, and the complete sequence contains seven exons and eight introns. Our DNA sequencing results indicated that two point mutations occurred in exon 3 of vg gene, they were synonymous mutations. Synonymous mutations indirectly affect the function of proteins222324. Furthermore, the four bases deletion in the exon 1 of vg gene was done in D. melanogaster mutant, resulting in the change of reading frame and the consequent failure to generate functional protein. Generally, frameshift mutations are base deletions or additions within the coding region of a gene. As expected, because of potential problems with disturbing the reading frame, the entire set of triplets downstream of the deletion or addition is altered. In many cases, the deletion or addition results in the presence of in-frame termination sequences which stop the product. Frameshift mutations may therefore result in more severe phenotypic effects than do a number of the base changes which cause either silent or conservative mutations in protein products25. In addition, our DNA sequencing results showed an insertion sequence with 12 nucleotides (5′-CTCGCCCTGTCT-3′) between the second and third exons, namely, the junction of exon 3 and the intron 3 of vg gene. This insertion sequence was another incentive for abnormal wing development of D. melanogaster induced by HHP. The coordinated interaction of these ectopic expression genes and vg gene mutants is crucial for abnormal wing development in D. melanogaster induced by HHP treatment. However, how these genes induced by HHP regulate abnormal wing development of D. melanogaster is not well understood and needs further study. Our results suggest the model shown in Fig. 5 for the molecular mechanism of wing mutagenesis induced by HHP treatment in D. melanogaster.

Bottom Line: Abnormal wing phenotypes of D. melanogaster induced by HHP were used to investigate the mutagenic mechanisms of HHP on organism.Thus 285 differentially expressed genes associated with wing mutations were identified using Affymetrix Drosophila Genome Array 2.0 and verified with RT-PCR.This study revealed the mutagenic mechanisms of HHP-induced mutagenesis in D. melanogaster and provided a new model for the study of evolution on organisms.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Superhard Materials, Jilin University, Changchun, 130012, P. R. China.

ABSTRACT
Although High hydrostatic pressure (HHP) as an important physical and chemical tool has been increasingly applied to research of organism, the response mechanisms of organism to HHP have not been elucidated clearly thus far. To identify mutagenic mechanisms of HHP on organisms, here, we treated Drosophila melanogaster (D. melanogaster) eggs with HHP. Approximately 75% of the surviving flies showed significant morphological abnormalities from the egg to the adult stages compared with control flies (p < 0.05). Some eggs displayed abnormal chorionic appendages, some larvae were large and red, and some adult flies showed wing abnormalities. Abnormal wing phenotypes of D. melanogaster induced by HHP were used to investigate the mutagenic mechanisms of HHP on organism. Thus 285 differentially expressed genes associated with wing mutations were identified using Affymetrix Drosophila Genome Array 2.0 and verified with RT-PCR. We also compared wing development-related central genes in the mutant flies with control flies using DNA sequencing to show two point mutations in the vestigial (vg) gene. This study revealed the mutagenic mechanisms of HHP-induced mutagenesis in D. melanogaster and provided a new model for the study of evolution on organisms.

No MeSH data available.


Related in: MedlinePlus