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Targeted Deletion of Btg1 and Btg2 Results in Homeotic Transformation of the Axial Skeleton.

Tijchon E, van Ingen Schenau D, van Opzeeland F, Tirone F, Hoogerbrugge PM, Van Leeuwen FN, Scheijen B - PLoS ONE (2015)

Bottom Line: Loss of Btg2, but not Btg1, was sufficient for complete posterior transformation of the thirteenth thoracic vertebra to the first lumbar vertebra.The Btg1-/-;Btg2-/- animals showed an even stronger phenotype, with L5 to S1 transformation.Together, these data show that both Btg1 and Btg2 are required for normal vertebral patterning of the axial skeleton, but each gene contributes differently in specifying the identity along the anterior-posterior axis of the skeleton.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Pediatric Oncology, Radboud university medical center, Nijmegen, The Netherlands.

ABSTRACT
Btg1 and Btg2 encode highly homologous proteins that are broadly expressed in different cell lineages, and have been implicated in different types of cancer. Btg1 and Btg2 have been shown to modulate the function of different transcriptional regulators, including Hox and Smad transcription factors. In this study, we examined the in vivo role of the mouse Btg1 and Btg2 genes in specifying the regional identity of the axial skeleton. Therefore, we examined the phenotype of Btg1 and Btg2 single knockout mice, as well as novel generated Btg1-/-;Btg2-/- double knockout mice, which were viable, but displayed a non-mendelian inheritance and smaller litter size. We observed both unique and overlapping phenotypes reminiscent of homeotic transformation along the anterior-posterior axis in the single and combined Btg1 and Btg2 knockout animals. Both Btg1-/- and Btg2-/- mice displayed partial posterior transformation of the seventh cervical vertebra, which was more pronounced in Btg1-/-;Btg2-/- mice, demonstrating that Btg1 and Btg2 act in synergy. Loss of Btg2, but not Btg1, was sufficient for complete posterior transformation of the thirteenth thoracic vertebra to the first lumbar vertebra. Moreover, Btg2-/- animals displayed complete posterior transformation of the sixth lumbar vertebra to the first sacral vertebra, which was only partially present at a low frequency in Btg1-/- mice. The Btg1-/-;Btg2-/- animals showed an even stronger phenotype, with L5 to S1 transformation. Together, these data show that both Btg1 and Btg2 are required for normal vertebral patterning of the axial skeleton, but each gene contributes differently in specifying the identity along the anterior-posterior axis of the skeleton.

No MeSH data available.


Related in: MedlinePlus

Characterization of Btg1-/-, Btg2-/- and Btg1-/-;Btg2-/- mice.(A) The mouse Btg1 gene is disrupted by insertion of a neomycin resistance cassette via SacII restriction sites in the first exon. The second exon of the Btg2 gene is replaced by a neomycin cassette in the antisense direction. The arrows indicate the position of primers used for genotyping. (B) Genotyping of mice was performed by PCR on genomic DNA using primers specific for the Btg1 and Btg2 wild-type (WT) and knockout (KO) allele. (C) Number of pups obtained from wild-type, heterozygous and homozygous Btg1, Btg2 and Btg1xBtg2 breedings. Data are from at least 4 independent crossings. *, P< 0.05, **, P< 0.01, ***, P< 0.001.
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pone.0131481.g001: Characterization of Btg1-/-, Btg2-/- and Btg1-/-;Btg2-/- mice.(A) The mouse Btg1 gene is disrupted by insertion of a neomycin resistance cassette via SacII restriction sites in the first exon. The second exon of the Btg2 gene is replaced by a neomycin cassette in the antisense direction. The arrows indicate the position of primers used for genotyping. (B) Genotyping of mice was performed by PCR on genomic DNA using primers specific for the Btg1 and Btg2 wild-type (WT) and knockout (KO) allele. (C) Number of pups obtained from wild-type, heterozygous and homozygous Btg1, Btg2 and Btg1xBtg2 breedings. Data are from at least 4 independent crossings. *, P< 0.05, **, P< 0.01, ***, P< 0.001.

Mentions: To investigate the function of Btg1 and Btg2 during normal development we obtained Btg1-/- and Btg2-/- single knockout [23, 28], and generated Btg1-/-;Btg2-/- double knockout mice on a C57Bl/6J background and examined their appearance for gross abnormalities (Fig 1A and 1B). Genotyping the offspring showed a non-mendelian inheritance pattern for the Btg1 knockout allele in the single knockout crosses (Table 1), while the Btg2 knockout allele was strongly underrepresented in the compound crosses (Table 2). The different homozygous knockout animals showed no obvious developmental defects, although the litter size upon interbreeding of the Btg1, Btg2 or Btg1;Btg2 homozygous knockout lines appeared to be smaller compared to WT animals (Fig 1C).


Targeted Deletion of Btg1 and Btg2 Results in Homeotic Transformation of the Axial Skeleton.

Tijchon E, van Ingen Schenau D, van Opzeeland F, Tirone F, Hoogerbrugge PM, Van Leeuwen FN, Scheijen B - PLoS ONE (2015)

Characterization of Btg1-/-, Btg2-/- and Btg1-/-;Btg2-/- mice.(A) The mouse Btg1 gene is disrupted by insertion of a neomycin resistance cassette via SacII restriction sites in the first exon. The second exon of the Btg2 gene is replaced by a neomycin cassette in the antisense direction. The arrows indicate the position of primers used for genotyping. (B) Genotyping of mice was performed by PCR on genomic DNA using primers specific for the Btg1 and Btg2 wild-type (WT) and knockout (KO) allele. (C) Number of pups obtained from wild-type, heterozygous and homozygous Btg1, Btg2 and Btg1xBtg2 breedings. Data are from at least 4 independent crossings. *, P< 0.05, **, P< 0.01, ***, P< 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4517811&req=5

pone.0131481.g001: Characterization of Btg1-/-, Btg2-/- and Btg1-/-;Btg2-/- mice.(A) The mouse Btg1 gene is disrupted by insertion of a neomycin resistance cassette via SacII restriction sites in the first exon. The second exon of the Btg2 gene is replaced by a neomycin cassette in the antisense direction. The arrows indicate the position of primers used for genotyping. (B) Genotyping of mice was performed by PCR on genomic DNA using primers specific for the Btg1 and Btg2 wild-type (WT) and knockout (KO) allele. (C) Number of pups obtained from wild-type, heterozygous and homozygous Btg1, Btg2 and Btg1xBtg2 breedings. Data are from at least 4 independent crossings. *, P< 0.05, **, P< 0.01, ***, P< 0.001.
Mentions: To investigate the function of Btg1 and Btg2 during normal development we obtained Btg1-/- and Btg2-/- single knockout [23, 28], and generated Btg1-/-;Btg2-/- double knockout mice on a C57Bl/6J background and examined their appearance for gross abnormalities (Fig 1A and 1B). Genotyping the offspring showed a non-mendelian inheritance pattern for the Btg1 knockout allele in the single knockout crosses (Table 1), while the Btg2 knockout allele was strongly underrepresented in the compound crosses (Table 2). The different homozygous knockout animals showed no obvious developmental defects, although the litter size upon interbreeding of the Btg1, Btg2 or Btg1;Btg2 homozygous knockout lines appeared to be smaller compared to WT animals (Fig 1C).

Bottom Line: Loss of Btg2, but not Btg1, was sufficient for complete posterior transformation of the thirteenth thoracic vertebra to the first lumbar vertebra.The Btg1-/-;Btg2-/- animals showed an even stronger phenotype, with L5 to S1 transformation.Together, these data show that both Btg1 and Btg2 are required for normal vertebral patterning of the axial skeleton, but each gene contributes differently in specifying the identity along the anterior-posterior axis of the skeleton.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Pediatric Oncology, Radboud university medical center, Nijmegen, The Netherlands.

ABSTRACT
Btg1 and Btg2 encode highly homologous proteins that are broadly expressed in different cell lineages, and have been implicated in different types of cancer. Btg1 and Btg2 have been shown to modulate the function of different transcriptional regulators, including Hox and Smad transcription factors. In this study, we examined the in vivo role of the mouse Btg1 and Btg2 genes in specifying the regional identity of the axial skeleton. Therefore, we examined the phenotype of Btg1 and Btg2 single knockout mice, as well as novel generated Btg1-/-;Btg2-/- double knockout mice, which were viable, but displayed a non-mendelian inheritance and smaller litter size. We observed both unique and overlapping phenotypes reminiscent of homeotic transformation along the anterior-posterior axis in the single and combined Btg1 and Btg2 knockout animals. Both Btg1-/- and Btg2-/- mice displayed partial posterior transformation of the seventh cervical vertebra, which was more pronounced in Btg1-/-;Btg2-/- mice, demonstrating that Btg1 and Btg2 act in synergy. Loss of Btg2, but not Btg1, was sufficient for complete posterior transformation of the thirteenth thoracic vertebra to the first lumbar vertebra. Moreover, Btg2-/- animals displayed complete posterior transformation of the sixth lumbar vertebra to the first sacral vertebra, which was only partially present at a low frequency in Btg1-/- mice. The Btg1-/-;Btg2-/- animals showed an even stronger phenotype, with L5 to S1 transformation. Together, these data show that both Btg1 and Btg2 are required for normal vertebral patterning of the axial skeleton, but each gene contributes differently in specifying the identity along the anterior-posterior axis of the skeleton.

No MeSH data available.


Related in: MedlinePlus