Human genomic site-specific recombination catalyzed by coliphage HK022 integrase.
It has been previously demonstrated that the wild type integrase (Int) protein of coliphage HK022 can catalyze site-specific recombination in human cells between attachment (att) sites that were placed on extrachromosomal plasmids. In the present report it is shown that Int can catalyze the site-specific recombination reactions in a human cell culture on the chromosomal level. These include integrative (attP x attB) as well as excisive (attL x attR) reactions each in two configurations. In the cis configuration both sites are on the same chromosome, in the trans configuration one site is on a chromosome and the other on an episome. The reactions in cis were observed without any selection force, using the green fluorescent protein (GFP) as a reporter. The reactions in trans could be detected only when a selection force was applied, using the hygromycin-resistant (Hyg(R)) phenotype as a selective marker. All reactions were catalyzed without the need to supply any of the accessory proteins that are required by Int in its Escherichia coli host. The versatility of the att sites may be an advantage in the utilization of Int to integrate plasmid DNA into the genome, followed by a partial exclusion of the integrated plasmid.
Harel-Levi
Gali
G
Department of Biochemistry, Tel-Aviv University, Tel Aviv 69978, Israel.
Goltsman
Janna
J
Tuby
Chen Nahum Josef Haim
CN
Yagil
Ezra
E
Kolot
Mikhail
M
D025523:Bacteriophage HK022 / Q000201:enzymology*; D015139:Blotting, Southern; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D015894:Genome, Human / Q000235:genetics*; D006801:Humans; D019426:Integrases / Q000378:metabolism*; D008957:Models, Genetic; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic / Q000235:genetics*
D004247:DNA; D019426:Integrases