Alternative splicing of vitamin D-24-hydroxylase: a novel mechanism for the regulation of extrarenal 1,25-dihydroxyvitamin D synthesis.
Synthesis of the active form of vitamin D, 1,25-dihydroxyvitamin D (1,25-(OH)(2)D), by renal epithelial cells is tightly controlled during normal calcium homeostasis. By contrast, macrophage production of 1,25-(OH)(2)D is often dysregulated with potential hypercalcemic complications. We have postulated that this is due to abnormal catabolism of 1,25-(OH)(2)D by the feedback control enzyme, vitamin D-24-hydroxylase (CYP24). Using chick HD-11 and human THP-1 myelomonocytic cell lines, we have shown that macrophage-like cells express a splice variant of the CYP24 gene (CYP24-SV), which encodes a truncated protein. Compared with the holo-CYP24 gene product in chick and human cells (508 and 513 amino acids, respectively), the truncated CYP24-SV versions consisted of 351 and 372 amino acids. These CYP24-SV proteins retained intact substrate-binding domains but lacked mitochondrial targeting sequences and were therefore catalytically inactive. In common with CYP24, expression of the CYP24 variants was induced by 1,25-(OH)(2)D but without a concomitant rise in 24-hydroxylase activity. However, overexpression of CYP24-SV in HD-11 and THP-1 cells reduced synthesis of 1,25-(OH)(2) D (40-50%), whereas antisense CYP24-SV expression increased 1,25-(OH)(2)D production by 2-7-fold. These data suggest that alternative splicing of CYP24 leads to the generation of a dominant negative-acting protein that is catalytically dysfunctional. We theorize that expression of the CYP24-SV may contribute to the extracellular accumulation of 1,25(OH)(2)D in human health and disease.
Ren
Songyang
S
Department of Medicine, Division of Endocrinology, Burns and Allen Research Institute, Cedars-Sinai Medical Center, Los Angeles, California 90048, USA.
Nguyen
Lisa
L
Wu
Shaoxing
S
Encinas
Carlos
C
Adams
John S
JS
Hewison
Martin
M
D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001921:Brain / Q000201:enzymology; D002117:Calcitriol / Q000096:biosynthesis*; D002384:Catalysis; D002460:Cell Line; D002645:Chickens; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000235:genetics*; D016373:DNA, Antisense / Q000235:genetics; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D005260:Female; D006801:Humans; D007668:Kidney / Q000201:enzymology; D008264:Macrophages / Q000201:enzymology / Q000378:metabolism; D008969:Molecular Sequence Data; D009928:Organ Specificity; D010920:Placenta / Q000201:enzymology; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012867:Skin / Q000201:enzymology; D013250:Steroid Hydroxylases / Q000737:chemistry / Q000235:genetics*; D013329:Structure-Activity Relationship; D014162:Transfection; D065668:Vitamin D3 24-Hydroxylase
D016373:DNA, Antisense; D018076:DNA, Complementary; D012333:RNA, Messenger; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D065668:Vitamin D3 24-Hydroxylase; D002117:Calcitriol