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Cell Biology International (2002) 26, 615–625 (Printed in Great Britain)
EXPRESSION AND POST-TRANSLATIONAL MODIFICATION OF HUMAN 4-HYDROXY-PHENYLPYRUVATE DIOXYGENASE
Lene Aarenstrupa, Anne‑Marie Falcha, Kirsten K Jakobsena, Søren Nevea, Linda Ø Henriksenb, Niels Tommerupc, Henrik Leffersd and Karsten Kristiansena,f1
aDepartment of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, DK-5230, Odense M, Denmark
bInstitute of Human Genetics, Aarhus University, Wilhelm Meyers Allé, DK-8000, Aarhus, Denmark
cDepartment of Medical Genetic, Building 24, Panum Institute, Blegdamsvej 3b, University of Copenhagen, DK-2200, Copenhagen N, Denmark
dDepartment of Growth and Reproduction, Section GR-5064, Rigshospitalet, Blegdamsvej 9, DK-2100, Copenhagen Ø Denmark


Abstract

4-hydroxyphenylpyruvate dioxygenase (HPD) (EC 1.13.11.27) is a key enzyme involved in tyrosine catabolism. Congenital HPD deficiency is a rare, relatively benign condition known as hereditary type III tyrosinemia. The severe type I tyrosinemia, caused by a deficiency of fumarylacetoacetate hydrolase which functions downstream of HPD in the tyrosine degradation pathway, is often associated with decreased expression of HPD, and interestingly, inhibition of HPD activity seems to ameliorate the clinical symptoms of type I tyrosinemia. The HPD gene was previously mapped to the chromosomal region 12q24→qter. In the present study high-resolution chromosome mapping localized the HPD gene to 12q24.31. DNase I footprinting, revealed that four regions of the HPD promoter were protected by rat liver nuclear proteins. Computer-assisted analyses suggested that these elements might bind Sp1/AP2, HNF4, HNF3/CREB, and C/EBP, respectively. In transient transfection experiments, the proximal 271bp of the promoter conferred basal transcriptional activation in human Chang cells. Sequences in intron 1 were able to enhance the activity of this basal promoter. Finally, vaccinia virus-based expression provided evidence that HPD is subject to phosphorylation, and furthermore, allowed mapping of the HPD protein in the human keratinocyte 2D database.


Key words: chromosomal localization, footprinting, phosphorylation, 2D gel-electrophoresis, tyrosinemia.

f1To whom correspondence should be addressed: Dr Karsten Kristiansen, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, DK-5320 Odense M, Denmark. Tel.: (+45) 6550 2408; Fax: (+45) 6550 2467; E-mail: kak@bmb.sdu.dk


Received 8 November 2001; accepted 2 April 2002

doi:10.1006/cbir.2002.0896


ISSN Print: 1065-6995
ISSN Electronic: 1095-8355
Published by Portland Press Limited on behalf of the International Federation for Cell Biology (IFCB)