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Methylated DNA-binding protein is present in various mammalian cell types
P C Supakar1, D Weist, D L Zhang
1Department of Biochemistry, Tulane Medical School, New Orleans, LA 70112.
Nucleic Acids Research
|August 25, 1988
Summary
Methylated DNA-binding protein (MDBP) from human placenta recognizes specific DNA sequences containing 5-methylcytosine. Similar MDBP activity was found across various mammalian cells but not in mosquito cells, suggesting a link to vertebrate DNA methylation.
Area of Science:
- Molecular Biology
- Epigenetics
- Biochemistry
Background:
- DNA methylation is a crucial epigenetic modification regulating gene expression in eukaryotes.
- Methylated DNA-binding proteins (MDBPs) play a role in recognizing methylated DNA.
- Understanding the specificity and distribution of MDBPs is key to deciphering epigenetic regulation.
Purpose of the Study:
- To investigate the presence and characteristics of DNA-binding proteins similar to human placental MDBP in various mammalian cell types.
- To determine if these DNA-binding activities exhibit similar sequence and methylation specificity as human MDBP.
- To explore the phylogenetic distribution of MDBP activity.
Main Methods:
- Nuclear extracts were prepared from various mammalian tissues and cell lines (rat, calf, human, mouse).
- Electrophoretic mobility shift assays (EMSAs) were used to detect and characterize DNA-binding proteins.
- A 14-bp MDBP-specific oligonucleotide duplex was employed to assess binding specificity.
Main Results:
- DNA-binding activity analogous to human placental MDBP was detected in nuclear extracts of rat tissues, calf thymus, human embryonal carcinoma cells, HeLa cells, and mouse LTK cells.
- These analogous proteins formed complexes with the MDBP-specific oligonucleotide, exhibiting identical DNA methylation and sequence specificity.
- While complexes from rat and calf tissues showed similar specificity, they had higher electrophoretic mobility, likely due to protease activity.
- MDBP activity was absent in cultured mosquito cells, contrasting with its presence in all tested mammalian cell types.
Conclusions:
- A conserved DNA-binding protein activity, similar to human placental MDBP, is present across diverse mammalian cell types.
- This activity is specific for methylated DNA sequences and shows conservation in binding characteristics.
- The absence of MDBP activity in mosquito cells suggests an association with vertebrate-type DNA methylation mechanisms.