Towards a complete picture of splice variants of the gene for 25-hydroxyvitamin D31alpha-hydroxylase in brain and skin cancer.
Recently, we reported amplification of the gene encoding the P450 Cytochrome 25-hydroxyvitamin D(3)1alpha-hydroxylase (CYP27B1) in 25% of human malignant glioma. Additionally, we reported the first alternative splice variants of CYP27B1. Here, we developed and employed a highly specific approach that combined nested and touchdown PCR to clone full length CYP27B1. In addition, we identified several new splice variants in human melanoma, glioblastoma multiforme (GBM), cervix carcinoma and kidney cell lines. All of the examined cell lines showed a similar expression pattern of the CYP27B1 variants. The new splice variants that were termed Hyd-V5, -V6, -V7, and -V8 were cloned and sequenced. All but one of the new variants showed an insertion of intron 1 leading to a premature termination signal and to truncated proteins without ferredoxin and haem-binding site of the P450 protein. There was no influence of 1alpha,25(OH)(2)D(3) on the expression pattern of the splice variants in melanoma cell line SkMel28.
Diesel
Britta
B
Institut für Humangenetik, Theoretische Medizin, Bldg. 60, Universität des Saarlandes, 66421 Homburg/Saar, Germany.
Seifert
Markus
M
Radermacher
Jens
J
Fischer
Ulrike
U
Tilgen
Wolfgang
W
Reichrath
Jörg
J
Meese
Eckart
E
D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000235:genetics*; D017398:Alternative Splicing; D001483:Base Sequence; D001932:Brain Neoplasms / Q000201:enzymology / Q000235:genetics*; D002460:Cell Line; D017931:DNA Primers; D006801:Humans; D008545:Melanoma / Q000201:enzymology / Q000235:genetics; D012878:Skin Neoplasms / Q000201:enzymology / Q000235:genetics*
D017931:DNA Primers; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase