Isolation of ribonuclease P activity from human epidermis and its regulation by retinoids in vitro.
Ribonuclease P (RNase P) is a key enzyme in tRNA biogenesis that catalyses the endonucleolytic cleavage of tRNA precursors and generates their mature 5' ends. The activity of this ribozyme has never been isolated from living human tissues and data about epidermal tRNA biogenesis are not available. The purpose of the present study was to isolate and purify RNase P from human epidermis and to investigate the in vitro effects of retinoids on its activity. Enzyme isolation and purification from homogenates of keratinocytes derived after trypsinization from dispase-separated human epidermis were carried out using phosphocellulose chromatography. The optimal activity of the enzyme was found at 100 mM NH4Cl and 5 mM MgCl2 at pH 7.5 and 37 degrees C. All-trans retinoic acid and acitretin revealed a dose-dependent inhibitory effect on RNase P activity. The isolation of RNase P activity from human epidermis, reported here for the first time, will enable the investigation of the possible involvement of this ribozyme in the regulation of epidermal differentiation and proliferation and the evaluation of its significance for the pathogenesis and gene therapy of various cutaneous disorders.
Pavlidou
Despina
D
Department of Biochemistry, School of Medicine, University of Patras, GR-26504 Rio-Patras, Greece.
Vourekas
Anastassios
A
Monastirli
Alexandra
A
Kalavrizioti
Dimitra
D
Tsambaos
Dionysios
D
Drainas
Denis
D
D017255:Acitretin / Q000494:pharmacology; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D006801:Humans; D015603:Keratinocytes / Q000201:enzymology*; D007641:Keratolytic Agents / Q000494:pharmacology*; D008853:Microscopy; D043262:Ribonuclease P / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014212:Tretinoin / Q000494:pharmacology
D007641:Keratolytic Agents; D014212:Tretinoin; D043262:Ribonuclease P; D017255:Acitretin