Analysis of mutation in the rat Pig-a assay: I) studies with bone marrow erythroid cells.
Environ Mol Mutagen
; 59(8): 722-732, 2018 10.
Article
en En
| MEDLINE
| ID: mdl-30091272
ABSTRACT
We have established a flow cytometry-based Pig-a assay for rat bone marrow erythroid cells (BMEs). The BME Pig-a assay uses a DNA-specific stain and two antibodies one against the transmembrane transferrin receptor (CD71 marker) and the other against the GPI-anchored complement inhibitory protein (CD59 marker). In F344 male rats treated acutely with a total of 120 mg/kg of N-ethyl-N-nitrosourea (ENU) the frequency of CD59-deficient phenotypically mutant BMEs increased approximately 24-fold compared to the rats concurrently treated with the vehicle. Such an increase of mutant BMEs coincides with increases of CD59-deficient reticulocytes measured in rats treated with similar doses of ENU. Sequence analysis of the endogenous X-linked Pig-a gene of CD59-deficient BMEs revealed that they are Pig-a mutants. The spectrum of ENU-induced Pig-a mutations in these BMEs was consistent with the in vivo mutagenic signature of ENU 73% of mutations occurred at AT basepairs, with the mutated T on the nontranscribed strand of the gene. TâA transversion was the most frequent mutation followed by TâC transition; no deletion or insertion mutations were present in the spectrum. Since BMEs are precursors of peripheral red blood cells, our findings suggest that CD59-deficient erythrocytes measured in the flow cytometric erythrocyte Pig-a assay develop from BMEs containing mutations in the Pig-a gene. Thus, the erythrocyte Pig-a assay detects mutation in the Pig-a gene. Environ. Mol. Mutagen. 59722-732, 2018. © 2018 Wiley Periodicals, Inc.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Receptores de Transferrina
/
Antígenos CD
/
Glicosilfosfatidilinositoles
/
Antígenos CD59
/
Células Eritroides
/
Citometría de Flujo
/
Proteínas de la Membrana
Límite:
Animals
Idioma:
En
Revista:
Environ Mol Mutagen
Asunto de la revista:
BIOLOGIA MOLECULAR
/
SAUDE AMBIENTAL
Año:
2018
Tipo del documento:
Article