Characterization of a partial cDNA for lysyl hydroxylase from human skin fibroblasts; lysyl hydroxylase mRNAs are regulated differently by minoxidil derivatives and hydralazine.
Lysyl hydroxylase (LH) is an essential enzyme in collagen biosynthesis that catalyzes the formation of hydroxylysine required for intermolecular crosslinking of collagen. We have isolated a partial (2.2-kb) cDNA for LH from human skin fibroblasts using PCR. DNA sequencing revealed 72% homology of the human coding sequence with the chick LH sequence at the nucleotide level and 76% homology predicted at the amino acid level. The LH cDNA hybridized strongly with two mRNA species of 2.4 and 3.4 kb on Northern blots of normal fibroblast RNA. Administration of minoxidil decreased both mRNA species without affecting levels of the mRNAs for the beta subunit of prolyl 4-hydroxylase (PH) or alpha 1(I) collagen. Two derivatives of minoxidil (3' hydroxyminoxidil and 4' hydroxyminoxidil) produced similar decreases in LH mRNAs. In contrast hydralazine increased the mRNAs for LH in parallel with its previously reported effect on the mRNA for the beta subunit of PH. This effect is accompanied by virtual elimination of the alpha 1(I) collagen mRNAs. These results on the action of minoxidil and hydralazine at the pretranslational level correlate well with their previously reported effect on enzyme activity and collagen biosynthesis and indicate that changes in steady-state mRNA levels can account directly for changes at the protein level. Moreover, the unique action of minoxidil in specifically decreasing LH mRNAs contrasts with the less specific stimulatory effects of hydralazine and suggests that these pharmaceuticals are regulating expression of LH at a pretranslational level by different mechanisms.
Yeowell
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HN
Duke University Medical Center, Division of Dermatology, Durham, NC 27710.
Ha
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Walker
L C
LC
Murad
S
S
Pinnell
S R
SR
D001483:Base Sequence; D015152:Blotting, Northern; D003094:Collagen / Q000235:genetics; D004247:DNA / Q000032:analysis; D005347:Fibroblasts / Q000201:enzymology*; D005784:Gene Amplification; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D006830:Hydralazine / Q000494:pharmacology*; D046911:Macromolecular Substances; D008914:Minoxidil / Q000031:analogs & derivatives* / Q000494:pharmacology; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D008240:Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase / Q000235:genetics*; D011393:Procollagen-Proline Dioxygenase / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D012867:Skin / Q000166:cytology* / Q000201:enzymology
D046911:Macromolecular Substances; D012333:RNA, Messenger; D006830:Hydralazine; D008914:Minoxidil; D003094:Collagen; D004247:DNA; D011393:Procollagen-Proline Dioxygenase; D008240:Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase