PMID: 9171090Jun 15, 1997Paper

Expression of herpes virus thymidine kinase in Neurospora crassa

Nucleic Acids Research
Matthew S SachsB S Margolin

Abstract

The expression of thymidine kinase in fungi, which normally lack this enzyme, will greatly aid the study of DNA metabolism and provide useful drug-sensitive phenotypes. The herpes simplex virus type-1 thymidine kinase gene ( tk ) was expressed in Neurospora crassa. tk was expressed as a fusion to N.crassa arg-2 regulatory sequences and as a hygromycin phosphotransferase-thymidine kinase fusion gene under the control of cytomegalovirus and SV40 sequences. Only strains containing tk showed thymidine kinase enzyme activity. In strains containing the arg-2 - tk gene, both the level of enzyme activity and the level of mRNA were reduced by growth in arginine medium, consistent with control through arg-2 regulatory sequences. Expression of thymidine kinase in N.crassa facilitated radioactive labeling of replicating DNA following addition of [3H]thymidine or [14C]thymidine to the growth medium. Thymidine labeling of DNA enabled demonstration that hydroxyurea can be used to block replication and synchronize the N.crassa mitotic cycle. Strains expressing thymidine kinase were also more sensitive than strains lacking thymidine kinase to anticancer and antiviral nucleoside drugs that are activated by thymidine kinase, including 5-fluoro-2'...Continue Reading

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Citations

May 11, 2005·Biochemical and Biophysical Research Communications·Danton H O'DayMichael A Myre
May 17, 2005·Fungal Genetics and Biology : FG & B·Chang Hyun KhangSeogchan Kang
Feb 27, 2007·Fungal Genetics and Biology : FG & B·Nirmala Bardiya, Patrick K T Shiu
Aug 17, 2006·The Journal of Biological Chemistry·Barry J BowmanEmma Jean Bowman

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