EXPRESSION OF HUMAN PLACENTAL-TYPE 6-PHOSPHOFRUCTO-2-KINASE FRUCTOSE-2,6-BISPHOSPHATASE IN VARIOUS CELLS AND CELL-LINES/

The expression of the human placental-type phosphofructo-2-kinase/fruc tose-2,6-bisphosphatase (HP2K) in various human cells and cell lines w as investigated at the levels of transcription and translation. Analys es by both Northern blotting and a reverse transcription-polymerase ch ain reaction (RT-PCR) showed that BeWo, U-937, SupT1, H9, HeLa, HepG2, and human mononuclear cells, as well as human placental chorionic cel ls, expressed HP2K mRNA. All the nucleotide sequences of RT-PCR produc ts from these cell lines were identical to that of HP2K. The expressio n of HP2K protein was determined by Western blot analysis of fractions from POROS-HQ column chromatography of the cell extracts from U-937 c ells, which was used as an example of HP2K-mRNA positive cell lines. A s with the 6-phosphofructo-2-kinase activity of HP2K, the activity of 6-phosphofructo-2-kinase in extracts of U-937 cells was not inhibited by glycerol 3-phosphate, a known 6-phosphofructo-2-kinase inhibitor of liver- and testis-type isozymes. These results strongly suggested tha t various cell lines, in particular U-937 cells, express functional HP 2K enzyme. Furthermore, 6-phosphofructo-2-kinase in U-937 cells was fo und to be activated by treatments with isoproterenol and phorbol la-my ristate 13-acetate, indicating regulation of 6-phosphofructo-2-kinase activity in U-937 cells by protein kinases A and C. (C) 1998 Academic Press.