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New methods for finding common insertion sites and co-occurring common insertion sites in transposon- and virus-based genetic screens.

Bergemann TL, Starr TK, Yu H, Steinbach M, Erdmann J, Chen Y, Cormier RT, Largaespada DA, Silverstein KA - Nucleic Acids Res. (2012)

Bottom Line: Insertional mutagenesis screens in mice are used to identify individual genes that drive tumor formation.We show that the new methods identify more relevant candidate genes and candidate gene pairs than found using previous methods.Identification of the biologically relevant set of mutations that occur in a single cell and cause tumor progression will aid in the rational design of single and combinatorial therapies in the upcoming age of personalized cancer therapy.

View Article: PubMed Central - PubMed

Affiliation: Division of Biostatistics, School of Public Health, University of Minnesota, Minneapolis, MN 55455, USA. tracy.l.bergemann@medtronic.com

ABSTRACT
Insertional mutagenesis screens in mice are used to identify individual genes that drive tumor formation. In these screens, candidate cancer genes are identified if their genomic location is proximal to a common insertion site (CIS) defined by high rates of transposon or retroviral insertions in a given genomic window. In this article, we describe a new method for defining CISs based on a Poisson distribution, the Poisson Regression Insertion Model, and show that this new method is an improvement over previously described methods. We also describe a modification of the method that can identify pairs and higher orders of co-occurring common insertion sites. We apply these methods to two data sets, one generated in a transposon-based screen for gastrointestinal tract cancer genes and another based on the set of retroviral insertions in the Retroviral Tagged Cancer Gene Database. We show that the new methods identify more relevant candidate genes and candidate gene pairs than found using previous methods. Identification of the biologically relevant set of mutations that occur in a single cell and cause tumor progression will aid in the rational design of single and combinatorial therapies in the upcoming age of personalized cancer therapy.

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Location of transposon insertions in CCIs. (A) Seven tumors had insertions in both the 3′ and 5′ regions of Apc. (B) Four tumors had insertions in both the upstream promoter of Rspo2 and in intron 1 of Ppmh1. All four insertions in the Rspo2 promoter inserted with the transposon viral promoter in the same orientation as the gene. (C) Four tumors had insertions in Cltc, two of which had insertions in Flt1 and the other two in Pan3. Insertions are depicted by a bent arrow, which points in the direction of the transposon promoter. Insertion numbers indicate tumors. Red arrow indicates direction of transcription. Solid black lines depict introns while dashed black lines depict intergenic DNA. Black boxes depict exons (A and B) while blue boxes depict genes (C). Arrow on bottom indicates length of DNA.
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gkr1295-F2: Location of transposon insertions in CCIs. (A) Seven tumors had insertions in both the 3′ and 5′ regions of Apc. (B) Four tumors had insertions in both the upstream promoter of Rspo2 and in intron 1 of Ppmh1. All four insertions in the Rspo2 promoter inserted with the transposon viral promoter in the same orientation as the gene. (C) Four tumors had insertions in Cltc, two of which had insertions in Flt1 and the other two in Pan3. Insertions are depicted by a bent arrow, which points in the direction of the transposon promoter. Insertion numbers indicate tumors. Red arrow indicates direction of transcription. Solid black lines depict introns while dashed black lines depict intergenic DNA. Black boxes depict exons (A and B) while blue boxes depict genes (C). Arrow on bottom indicates length of DNA.

Mentions: The first CCI identified cooperating mutations within the Apc gene. The biology of Apc mutations has been extensively studied in colon cancer and the current hypothesis is that an initial truncating mutation resulting in a hypomorphic protein product is normally followed by loss of heterozygosity of the remaining wild-type allele (30). This hypothesis fits well with the Apc-Apc CCI identified in the set of intestinal tumors analyzed in this article. The seven tumors that constitute this CCI show multiple mutations in the first intron and 3′ upstream region of Apc accompanied by paired mutations in downstream introns (Figure 2A). It is possible that the first mutation creates a product because it occurs in the first intron or promoter region of Apc, while the second mutation creates a truncated product in the second allele.Figure 2.


New methods for finding common insertion sites and co-occurring common insertion sites in transposon- and virus-based genetic screens.

Bergemann TL, Starr TK, Yu H, Steinbach M, Erdmann J, Chen Y, Cormier RT, Largaespada DA, Silverstein KA - Nucleic Acids Res. (2012)

Location of transposon insertions in CCIs. (A) Seven tumors had insertions in both the 3′ and 5′ regions of Apc. (B) Four tumors had insertions in both the upstream promoter of Rspo2 and in intron 1 of Ppmh1. All four insertions in the Rspo2 promoter inserted with the transposon viral promoter in the same orientation as the gene. (C) Four tumors had insertions in Cltc, two of which had insertions in Flt1 and the other two in Pan3. Insertions are depicted by a bent arrow, which points in the direction of the transposon promoter. Insertion numbers indicate tumors. Red arrow indicates direction of transcription. Solid black lines depict introns while dashed black lines depict intergenic DNA. Black boxes depict exons (A and B) while blue boxes depict genes (C). Arrow on bottom indicates length of DNA.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3351147&req=5

gkr1295-F2: Location of transposon insertions in CCIs. (A) Seven tumors had insertions in both the 3′ and 5′ regions of Apc. (B) Four tumors had insertions in both the upstream promoter of Rspo2 and in intron 1 of Ppmh1. All four insertions in the Rspo2 promoter inserted with the transposon viral promoter in the same orientation as the gene. (C) Four tumors had insertions in Cltc, two of which had insertions in Flt1 and the other two in Pan3. Insertions are depicted by a bent arrow, which points in the direction of the transposon promoter. Insertion numbers indicate tumors. Red arrow indicates direction of transcription. Solid black lines depict introns while dashed black lines depict intergenic DNA. Black boxes depict exons (A and B) while blue boxes depict genes (C). Arrow on bottom indicates length of DNA.
Mentions: The first CCI identified cooperating mutations within the Apc gene. The biology of Apc mutations has been extensively studied in colon cancer and the current hypothesis is that an initial truncating mutation resulting in a hypomorphic protein product is normally followed by loss of heterozygosity of the remaining wild-type allele (30). This hypothesis fits well with the Apc-Apc CCI identified in the set of intestinal tumors analyzed in this article. The seven tumors that constitute this CCI show multiple mutations in the first intron and 3′ upstream region of Apc accompanied by paired mutations in downstream introns (Figure 2A). It is possible that the first mutation creates a product because it occurs in the first intron or promoter region of Apc, while the second mutation creates a truncated product in the second allele.Figure 2.

Bottom Line: Insertional mutagenesis screens in mice are used to identify individual genes that drive tumor formation.We show that the new methods identify more relevant candidate genes and candidate gene pairs than found using previous methods.Identification of the biologically relevant set of mutations that occur in a single cell and cause tumor progression will aid in the rational design of single and combinatorial therapies in the upcoming age of personalized cancer therapy.

View Article: PubMed Central - PubMed

Affiliation: Division of Biostatistics, School of Public Health, University of Minnesota, Minneapolis, MN 55455, USA. tracy.l.bergemann@medtronic.com

ABSTRACT
Insertional mutagenesis screens in mice are used to identify individual genes that drive tumor formation. In these screens, candidate cancer genes are identified if their genomic location is proximal to a common insertion site (CIS) defined by high rates of transposon or retroviral insertions in a given genomic window. In this article, we describe a new method for defining CISs based on a Poisson distribution, the Poisson Regression Insertion Model, and show that this new method is an improvement over previously described methods. We also describe a modification of the method that can identify pairs and higher orders of co-occurring common insertion sites. We apply these methods to two data sets, one generated in a transposon-based screen for gastrointestinal tract cancer genes and another based on the set of retroviral insertions in the Retroviral Tagged Cancer Gene Database. We show that the new methods identify more relevant candidate genes and candidate gene pairs than found using previous methods. Identification of the biologically relevant set of mutations that occur in a single cell and cause tumor progression will aid in the rational design of single and combinatorial therapies in the upcoming age of personalized cancer therapy.

Show MeSH
Related in: MedlinePlus