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Expression of Human Chromosome 2 Ornithine Decarboxylase Gene in Ornithine Decarboxylase-deficient Chinese Hamster Ovary Cells¹

Department of Human Oncology, University of Wisconsin Clinical Cancer Center, Madison, Wisconsin 53792

Ornithine decarboxylase (ODC) belongs to a multigene family and some of these may very well be nonfunctional (pseudogenes). We isolated an ODC gene from a human chromosome 2-specific library and transfected the gene into ODC-deficient Chinese hamster ovary cells to directly demonstrate that this ODC gene is functional and ODC is essential for cell proliferation. After screening 2.5 x 10⁵ plaques using a human ODC complementary DNA probe, a typical clone with a 5.4-kilobase insert was isolated and then cloned into the HindIII site of the pGem-1 vector. One (phODC 2B1) of these clones containing a 5.4-kilobase ODC gene insert was identified. Restriction enzyme analysis and partial sequencing data revealed that phODC 2B1 contained the full length protein-coding sequences but lacked first exon and 3'-polyadenylation sequences. Primer extension analysis indicated that human ODC mRNA has homologous sequences with the ODC gene from human chromosome 2. To determine that the chromosome 2 ODC gene is functional, ODC-deficient Chinese hamster ovary cells were transfected with the ODC expression vector (phSV2B1-neo) and several G418-resistant transfectants were isolated which expressed 70- to 400-fold more ODC activity than parental or wild-type Chinese hamster ovary cells. Furthermore, these stable transfectants exhibited a higher growth rate than wild-type cells. These results indicate that the ODC gene from human chromosome 2 encodes functional ODC protein, and ODC (and its product putrescine) is required for cell growth.

¹ This work was supported in part by the United States Public Health Service Research Grants CA-35368 and CA-42585 from the National Cancer Institute. J. T. H. was a recipient of a merit scholarship for Ph.D. studies from the Ministry of Education, Republic of China. A preliminary report of this work was presented at the 80th Annual Meeting of the American Association for Cancer Research Inc. (28).

² To whom requests for reprints should be addressed.

Received 4/11/89. Revised 12/20/89.