Hypoxia Associated Proteolytic Processing of OS-9 by the Metalloproteinase Meprin β

Figure 7

Immunoblots for OS-9 and HIF-1α in meprin β transfected HEK293 cells exposed to CoCl₂. HEK293 cells were cultured in MEM with 10% FBS to 90% confluence, serum-starved overnight, and exposed to 125 μM CoCl₂ for 0–3 h. Proteins were extracted and fractionated into cytosolic- and nuclear-enriched fractions. Western blot analysis and optic densitometry were used to evaluate the levels of OS-9 and HIF-1α in the nuclear-enriched protein fractions. Tubulin was used as a loading control. CoCl₂ exposure resulted in a time-dependent decrease in OS-9 levels in the meprin β transfected cells but not in the mock transfected controls. Stabilization of HIF-1α was evidenced by increased nuclear protein levels in all the cells exposed to CoCl₂. The relative OD was determined by the intensity of the protein relative to the intensity of the loading control as measured by spectrophotometry. The fold-change represents a comparison of the relative OD at each time point to the relative OD at 0 h. ; .