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Histone acetyl transferase 1 is essential for mammalian development, genome stability, and the processing of newly synthesized histones H3 and H4.

Nagarajan P, Ge Z, Sirbu B, Doughty C, Agudelo Garcia PA, Schlederer M, Annunziato AT, Cortez D, Kenner L, Parthun MR - PLoS Genet. (2013)

Bottom Line: Murine Hat1 is essential for viability, as homozygous deletion of Hat1 results in neonatal lethality.The neonatal lethality is due to severe defects in lung development that result in less aeration and respiratory distress.Many of the Hat1(-/-) neonates also display significant craniofacial defects with abnormalities in the bones of the skull and jaw.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biochemistry, The Ohio State University, Columbus, OH, USA.

ABSTRACT
Histone acetyltransferase 1 is an evolutionarily conserved type B histone acetyltransferase that is thought to be responsible for the diacetylation of newly synthesized histone H4 on lysines 5 and 12 during chromatin assembly. To understand the function of this enzyme in a complex organism, we have constructed a conditional mouse knockout model of Hat1. Murine Hat1 is essential for viability, as homozygous deletion of Hat1 results in neonatal lethality. The lungs of embryos and pups genetically deficient in Hat1 were much less mature upon histological evaluation. The neonatal lethality is due to severe defects in lung development that result in less aeration and respiratory distress. Many of the Hat1(-/-) neonates also display significant craniofacial defects with abnormalities in the bones of the skull and jaw. Hat1(-/-) mouse embryonic fibroblasts (MEFs) are defective in cell proliferation and are sensitive to DNA damaging agents. In addition, the Hat1(-/-) MEFs display a marked increase in genome instability. Analysis of histone dynamics at sites of replication-coupled chromatin assembly demonstrates that Hat1 is not only responsible for the acetylation of newly synthesized histone H4 but is also required to maintain the acetylation of histone H3 on lysines 9, 18, and 27 during replication-coupled chromatin assembly.

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Histone acetyltransferase 1 is essential for viability in mice.A) Schematic diagram of the wild type mouse Hat1 locus (top), the Hat1 locus following integration of loxP sequences flanking intron three (middle) and the Hat1 locus following Cre mediated deletion of exon 3. Exons are represented by purple rectangles. Locations of probes and PCR primers are indicated. (B) Genomic DNA isolated from a parental (C57/bl6) mouse and mice generated from a cross between a chimeric mouse and a wild type mouse was digested with EcoRV and analyzed by Southern blot using the indicated probe. Mice 1, 4 and 6 are Hat1flox/WT. C) A Hat1flox/WT mouse was crossed with a mouse that ubiquitously expresses the cre recombinase. Genomic DNA was isolated from mice generated by this cross, digested with EcoRV and analyzed by Southern blot with the indicated probe. Mice 3, 5 and 9 are WT/KO/Cre. D) Table lists the number of expected, obtained and viable pups of the indicated genomes derived from matings of Hat1+/− mice. E) PCR genotyping of a representative litter from a Hat1+/− X Hat1+/− mating. Primers P1 and P2 (see above) were used for amplification. Arrows indicate specific PCR products. F) Representative neonatal pups from Hat1+/− X Hat1+/− matings with the indicated genotype. F) Body weight of pups was measured immediately following birth. Data are derived from 10 pups of each genotype.
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pgen-1003518-g001: Histone acetyltransferase 1 is essential for viability in mice.A) Schematic diagram of the wild type mouse Hat1 locus (top), the Hat1 locus following integration of loxP sequences flanking intron three (middle) and the Hat1 locus following Cre mediated deletion of exon 3. Exons are represented by purple rectangles. Locations of probes and PCR primers are indicated. (B) Genomic DNA isolated from a parental (C57/bl6) mouse and mice generated from a cross between a chimeric mouse and a wild type mouse was digested with EcoRV and analyzed by Southern blot using the indicated probe. Mice 1, 4 and 6 are Hat1flox/WT. C) A Hat1flox/WT mouse was crossed with a mouse that ubiquitously expresses the cre recombinase. Genomic DNA was isolated from mice generated by this cross, digested with EcoRV and analyzed by Southern blot with the indicated probe. Mice 3, 5 and 9 are WT/KO/Cre. D) Table lists the number of expected, obtained and viable pups of the indicated genomes derived from matings of Hat1+/− mice. E) PCR genotyping of a representative litter from a Hat1+/− X Hat1+/− mating. Primers P1 and P2 (see above) were used for amplification. Arrows indicate specific PCR products. F) Representative neonatal pups from Hat1+/− X Hat1+/− matings with the indicated genotype. F) Body weight of pups was measured immediately following birth. Data are derived from 10 pups of each genotype.

Mentions: There is a single homolog of Hat1 in the murine genome that is highly similar to human and yeast Hat1. The murine Hat1 gene consists of 11 exons (Figure 1A). A construct was generated to target the integration of loxP sites to flank Hat1 exon 3. In the presence of cre recombinase, exon 3 can be deleted with the subsequent introduction of a stop codon (Figure 1A). This will create a truncation mutant of Hat1 that produces a product less 50 amino acids long. In the event that alternate splicing occurs in the Hat1 gene that could skip exon 3, only exon 9 can be spliced to exon 2 and retain the proper reading frame. In this case, the protein that would be produced would lack the entire Hat1 active site.


Histone acetyl transferase 1 is essential for mammalian development, genome stability, and the processing of newly synthesized histones H3 and H4.

Nagarajan P, Ge Z, Sirbu B, Doughty C, Agudelo Garcia PA, Schlederer M, Annunziato AT, Cortez D, Kenner L, Parthun MR - PLoS Genet. (2013)

Histone acetyltransferase 1 is essential for viability in mice.A) Schematic diagram of the wild type mouse Hat1 locus (top), the Hat1 locus following integration of loxP sequences flanking intron three (middle) and the Hat1 locus following Cre mediated deletion of exon 3. Exons are represented by purple rectangles. Locations of probes and PCR primers are indicated. (B) Genomic DNA isolated from a parental (C57/bl6) mouse and mice generated from a cross between a chimeric mouse and a wild type mouse was digested with EcoRV and analyzed by Southern blot using the indicated probe. Mice 1, 4 and 6 are Hat1flox/WT. C) A Hat1flox/WT mouse was crossed with a mouse that ubiquitously expresses the cre recombinase. Genomic DNA was isolated from mice generated by this cross, digested with EcoRV and analyzed by Southern blot with the indicated probe. Mice 3, 5 and 9 are WT/KO/Cre. D) Table lists the number of expected, obtained and viable pups of the indicated genomes derived from matings of Hat1+/− mice. E) PCR genotyping of a representative litter from a Hat1+/− X Hat1+/− mating. Primers P1 and P2 (see above) were used for amplification. Arrows indicate specific PCR products. F) Representative neonatal pups from Hat1+/− X Hat1+/− matings with the indicated genotype. F) Body weight of pups was measured immediately following birth. Data are derived from 10 pups of each genotype.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3675013&req=5

pgen-1003518-g001: Histone acetyltransferase 1 is essential for viability in mice.A) Schematic diagram of the wild type mouse Hat1 locus (top), the Hat1 locus following integration of loxP sequences flanking intron three (middle) and the Hat1 locus following Cre mediated deletion of exon 3. Exons are represented by purple rectangles. Locations of probes and PCR primers are indicated. (B) Genomic DNA isolated from a parental (C57/bl6) mouse and mice generated from a cross between a chimeric mouse and a wild type mouse was digested with EcoRV and analyzed by Southern blot using the indicated probe. Mice 1, 4 and 6 are Hat1flox/WT. C) A Hat1flox/WT mouse was crossed with a mouse that ubiquitously expresses the cre recombinase. Genomic DNA was isolated from mice generated by this cross, digested with EcoRV and analyzed by Southern blot with the indicated probe. Mice 3, 5 and 9 are WT/KO/Cre. D) Table lists the number of expected, obtained and viable pups of the indicated genomes derived from matings of Hat1+/− mice. E) PCR genotyping of a representative litter from a Hat1+/− X Hat1+/− mating. Primers P1 and P2 (see above) were used for amplification. Arrows indicate specific PCR products. F) Representative neonatal pups from Hat1+/− X Hat1+/− matings with the indicated genotype. F) Body weight of pups was measured immediately following birth. Data are derived from 10 pups of each genotype.
Mentions: There is a single homolog of Hat1 in the murine genome that is highly similar to human and yeast Hat1. The murine Hat1 gene consists of 11 exons (Figure 1A). A construct was generated to target the integration of loxP sites to flank Hat1 exon 3. In the presence of cre recombinase, exon 3 can be deleted with the subsequent introduction of a stop codon (Figure 1A). This will create a truncation mutant of Hat1 that produces a product less 50 amino acids long. In the event that alternate splicing occurs in the Hat1 gene that could skip exon 3, only exon 9 can be spliced to exon 2 and retain the proper reading frame. In this case, the protein that would be produced would lack the entire Hat1 active site.

Bottom Line: Murine Hat1 is essential for viability, as homozygous deletion of Hat1 results in neonatal lethality.The neonatal lethality is due to severe defects in lung development that result in less aeration and respiratory distress.Many of the Hat1(-/-) neonates also display significant craniofacial defects with abnormalities in the bones of the skull and jaw.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biochemistry, The Ohio State University, Columbus, OH, USA.

ABSTRACT
Histone acetyltransferase 1 is an evolutionarily conserved type B histone acetyltransferase that is thought to be responsible for the diacetylation of newly synthesized histone H4 on lysines 5 and 12 during chromatin assembly. To understand the function of this enzyme in a complex organism, we have constructed a conditional mouse knockout model of Hat1. Murine Hat1 is essential for viability, as homozygous deletion of Hat1 results in neonatal lethality. The lungs of embryos and pups genetically deficient in Hat1 were much less mature upon histological evaluation. The neonatal lethality is due to severe defects in lung development that result in less aeration and respiratory distress. Many of the Hat1(-/-) neonates also display significant craniofacial defects with abnormalities in the bones of the skull and jaw. Hat1(-/-) mouse embryonic fibroblasts (MEFs) are defective in cell proliferation and are sensitive to DNA damaging agents. In addition, the Hat1(-/-) MEFs display a marked increase in genome instability. Analysis of histone dynamics at sites of replication-coupled chromatin assembly demonstrates that Hat1 is not only responsible for the acetylation of newly synthesized histone H4 but is also required to maintain the acetylation of histone H3 on lysines 9, 18, and 27 during replication-coupled chromatin assembly.

Show MeSH
Related in: MedlinePlus