PARADOXICAL EFFECTS OF PHOSPHATE TO DIRECTLY REGULATE THE LEVEL OF SKELETAL ALKALINE-PHOSPHATASE ACTIVITY IN HUMAN OSTEOSARCOMA (SAOS-2) CELLS AND INVERSELY REGULATE THE LEVEL OF SKELETAL ALKALINE-PHOSPHATASE MESSENGER-RNA

Recent studies indicate that the amount of alkaline phosphatase (ALP) activity in human osteoblast-line cells is proportional to the concent ration of phosphate in the culture medium. The current studies were in tended to extend those observations and to determine whether the effec ts of phosphate (and phosphate esters and analogs) to alter the cellul ar level of ALP activity, in human osteosarcoma SaOS-2 cells, reflecte d regulation at the level of transcription. Consistent with previous f indings, we found direct, time- and dose-dependent correlations betwee n the concentration of phosphate and the amount of ALP activity/mg cel l protein (P < 0.05). Surprisingly, we also found a negative correlati on between the phosphate concentration in the medium and the level of skeletal ALP mRNA (e.g., r = -0.98, P < 0.01 at 24 hours). As the high est cellular levels of skeletal ALP activity were associated with the lowest levels of ALP mRNA, these data indicated that the phosphate-dep endent increase in ALP activity was not mediated by an increase in tra nscription and, conversely, that the effect of phosphate withdrawal to decrease ALP activity was not mediated by a decrease in transcription . Two additional studies have further suggested that these paradoxical effects of phosphate-increasing ALP activity while decreasing ALP mRN A-may be unique to inorganic phosphate: (1) beta-glycerol-phosphate (a t 10 mmol/liter) mimicked the action of phosphate to increase the cell ular level of ALP activity (P < 0.001), but did not mimick the action of phosphate to decrease the level of ALP mRNA; and (2) neither phenyl phosphonate or molybdate (both of which, like phosphate, increased ALP activity, P < 0.05) or berate (which decreased ALP activity, P < 0.05 ) altered the level of ALP mRNA. In summary, these studies show that t he effect of phosphate to regulate the level of ALP activity in human osteoblast-line cells is not determined by effects on ALP mRNA synthes is, suggesting that the regulation may depend on phosphate-dependent c hanges in the translation of ALP mRNA and/or a modification of ALP act ivity.