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Junction site analysis of chimeric CYP21A1P/CYP21A2 genes in 21-hydroxylase deficiency.
Clin Chem. 2012 Feb; 58(2):421-30.CC

Abstract

BACKGROUND

Chimeric CYP21A1P/CYP21A2 genes, caused by homologous recombination between CYP21A2 (cytochrome P450, family 21, subfamily A, polypeptide 2) and its highly homologous pseudogene CYP21A1P (cytochrome P450, family 21, subfamily A, polypeptide 1 pseudogene), are common in patients with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD). A comprehensive junction site analysis of chimeric CYP21A1P/CYP21A2 genes is needed for optimizing genetic analysis strategy and determining clinical relevance.

METHODS

We conducted a comprehensive genetic analysis of chimeric CYP21A1P/CYP21A2 genes in a cohort of 202 unrelated 21-OHD patients. Targeted CYP21A2 mutation analysis was performed, and genotyping of chimeric CYP21A1P/CYP21A2 genes was cross-confirmed with Southern blot, RFLP, and multiplex ligation-dependent probe amplification analyses. Junction sites of chimera genes were determined by sequencing the long-PCR products amplified with primers CYP779f and Tena32F. An updated bioinformatics survey of Chi-like sequences was also performed.

RESULTS

Of 100 probands with a chimeric allele, 96 had a chimera associated with the severe classic salt-wasting form of CAH, and the remaining 4 carried an uncommon attenuated chimera with junction sites upstream of In2G (c.293-13A/C>G), which is associated with a milder phenotype. In addition to 6 of 7 reported chimeras, we identified a novel classic chimera (CH-8) and a novel attenuated chimera (CH-9). Attenuated chimeras explained prior genotype-phenotype discrepancies in 3 of the patients. Sequencing the CYP779f/Tena32F amplicons accurately differentiated between classic and attenuated chimeras. The bioinformatics survey revealed enrichment of Chi-like sequences within or in the vicinity of intron 2.

CONCLUSIONS

Junction site analysis can explain some genotype-phenotype discrepancies. Sequencing the well-established CYP779f/Tena32F amplicons is an unequivocal strategy for detecting attenuated chimeric CYP21A1P/CYP21A2 genes, which are clinically relevant.

Authors+Show Affiliations

Laboratory of Clinical Investigation, National Institute on Aging, Baltimore, MD 21224, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

22156666

Citation

Chen, Wuyan, et al. "Junction Site Analysis of Chimeric CYP21A1P/CYP21A2 Genes in 21-hydroxylase Deficiency." Clinical Chemistry, vol. 58, no. 2, 2012, pp. 421-30.
Chen W, Xu Z, Sullivan A, et al. Junction site analysis of chimeric CYP21A1P/CYP21A2 genes in 21-hydroxylase deficiency. Clin Chem. 2012;58(2):421-30.
Chen, W., Xu, Z., Sullivan, A., Finkielstain, G. P., Van Ryzin, C., Merke, D. P., & McDonnell, N. B. (2012). Junction site analysis of chimeric CYP21A1P/CYP21A2 genes in 21-hydroxylase deficiency. Clinical Chemistry, 58(2), 421-30. https://doi.org/10.1373/clinchem.2011.174037
Chen W, et al. Junction Site Analysis of Chimeric CYP21A1P/CYP21A2 Genes in 21-hydroxylase Deficiency. Clin Chem. 2012;58(2):421-30. PubMed PMID: 22156666.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Junction site analysis of chimeric CYP21A1P/CYP21A2 genes in 21-hydroxylase deficiency. AU - Chen,Wuyan, AU - Xu,Zhi, AU - Sullivan,Annie, AU - Finkielstain,Gabriela P, AU - Van Ryzin,Carol, AU - Merke,Deborah P, AU - McDonnell,Nazli B, Y1 - 2011/12/07/ PY - 2011/12/14/entrez PY - 2011/12/14/pubmed PY - 2012/3/16/medline SP - 421 EP - 30 JF - Clinical chemistry JO - Clin Chem VL - 58 IS - 2 N2 - BACKGROUND: Chimeric CYP21A1P/CYP21A2 genes, caused by homologous recombination between CYP21A2 (cytochrome P450, family 21, subfamily A, polypeptide 2) and its highly homologous pseudogene CYP21A1P (cytochrome P450, family 21, subfamily A, polypeptide 1 pseudogene), are common in patients with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD). A comprehensive junction site analysis of chimeric CYP21A1P/CYP21A2 genes is needed for optimizing genetic analysis strategy and determining clinical relevance. METHODS: We conducted a comprehensive genetic analysis of chimeric CYP21A1P/CYP21A2 genes in a cohort of 202 unrelated 21-OHD patients. Targeted CYP21A2 mutation analysis was performed, and genotyping of chimeric CYP21A1P/CYP21A2 genes was cross-confirmed with Southern blot, RFLP, and multiplex ligation-dependent probe amplification analyses. Junction sites of chimera genes were determined by sequencing the long-PCR products amplified with primers CYP779f and Tena32F. An updated bioinformatics survey of Chi-like sequences was also performed. RESULTS: Of 100 probands with a chimeric allele, 96 had a chimera associated with the severe classic salt-wasting form of CAH, and the remaining 4 carried an uncommon attenuated chimera with junction sites upstream of In2G (c.293-13A/C>G), which is associated with a milder phenotype. In addition to 6 of 7 reported chimeras, we identified a novel classic chimera (CH-8) and a novel attenuated chimera (CH-9). Attenuated chimeras explained prior genotype-phenotype discrepancies in 3 of the patients. Sequencing the CYP779f/Tena32F amplicons accurately differentiated between classic and attenuated chimeras. The bioinformatics survey revealed enrichment of Chi-like sequences within or in the vicinity of intron 2. CONCLUSIONS: Junction site analysis can explain some genotype-phenotype discrepancies. Sequencing the well-established CYP779f/Tena32F amplicons is an unequivocal strategy for detecting attenuated chimeric CYP21A1P/CYP21A2 genes, which are clinically relevant. SN - 1530-8561 UR - https://www.unboundmedicine.com/medline/citation/22156666/Junction_site_analysis_of_chimeric_CYP21A1P/CYP21A2_genes_in_21_hydroxylase_deficiency_ L2 - https://academic.oup.com/clinchem/article-lookup/doi/10.1373/clinchem.2011.174037 DB - PRIME DP - Unbound Medicine ER -