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Cloning and characterization of the porcine DBC1 gene encoding deleted in bladder cancer
Knud Larsen1, Jamal Momeni, Leila Farajzadeh
1Department of Molecular Biology and Genetics, Science and Technology, Aarhus University, Blichers Alle 20, P.O. Box 50, 8830, Tjele, Denmark, Knud.Larsen@agrsci.dk.
Abstract:
Deleted in bladder cancer 1 (DBC1) is a tumour suppressor which is involved in the regulation of cell growth and programmed cell death. In this study we report the cloning and characterization of porcine DBC1 cDNA. RT-PCR cloning produced a cDNA with an open reading frame of 2,283 bp encoding a polypeptide of 761 amino acids with a predicted molecular mass of 88.6 kDa and estimated isoelectric point of 9.1. The encoded pig DBC1 protein shows a very high amino acid similarity to human (99 %) and to mouse (98 %) DBC1. The porcine DBC1 gene was mapped to chromosome 1. The nucleotide sequence of the promoter displayed a high degree of conservation of elements responsible for neuron-specific expression. The porcine DBC1 gene was found to be highly expressed in brain tissues. The methylation status of the porcine DBC1 gene was examined in brain and liver by bisulfite sequencing. Methylation percentages of 53-61 were observed for the gene body whereas significantly lower values (1-4 %) were found in exon 1 and the promoter sequence of DBC1. The sequences of the porcine DBC1 cDNA and the DBC1 promoter and exon 1 sequence have been submitted to DDBJ/EMBL/GenBank under the accession numbers KF733442 and KJ396193, respectively.
Insights
Researchers cloned and characterized the porcine Deleted in bladder cancer 1 (DBC1) gene, finding high similarity to human and mouse DBC1. This tumor suppressor gene is highly expressed in pig brains and shows specific methylation patterns.
Area of Science:
- Genetics
- Molecular Biology
- Comparative Genomics
Background:
- Deleted in bladder cancer 1 (DBC1) is a known tumor suppressor involved in cell growth and apoptosis.
- Understanding DBC1's role in different species can provide insights into its function and potential therapeutic applications.
Purpose of the Study:
- To clone and characterize the porcine Deleted in bladder cancer 1 (DBC1) cDNA.
- To analyze the expression and methylation status of the porcine DBC1 gene.
- To compare porcine DBC1 with its counterparts in other species.
Main Methods:
- RT-PCR was used to clone the porcine DBC1 cDNA.
- Bioinformatic analysis was performed to predict protein characteristics.
- Gene mapping was conducted to determine the chromosomal location.
- Bisulfite sequencing was employed to examine DNA methylation patterns.
Main Results:
- A porcine DBC1 cDNA of 2,283 bp was cloned, encoding a 761-amino acid protein.
- Porcine DBC1 exhibits high amino acid similarity (99% human, 98% mouse).
- The porcine DBC1 gene was mapped to chromosome 1 and showed high expression in brain tissues.
- Differential methylation was observed, with lower levels in the promoter and exon 1 compared to the gene body.
Conclusions:
- The study successfully cloned and characterized porcine DBC1, revealing high conservation with mammalian orthologs.
- Porcine DBC1 is highly expressed in the brain, suggesting a potential role in neuronal function.
- The distinct methylation patterns indicate epigenetic regulation of DBC1 in pigs.
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