Expression of p300-truncated fragments results in the modulation of apoptosis in rat mesangial cells

Background. Mesangial cell proliferation, apoptosis, and matrix deposition have pivotal roles in the pathogenesis of renal diseases such as diabetic n ephropathy and glomerulonephritis. The behavior of mesangial cells depends on the integration of intracellular signals elicited by hormones and cytoki nes. We hypothesized that p300 is primarily involved in the integration of signal transduction pathways in rat mesangial cells (RMCs) and that interfe rence with p300 function will alter apoptotic signals. Methods. We established an RMC cell line expressing the Tet-activator (tTA) . RMC-tTA cells were transiently transfected with vectors coding for either the N-terminal third or the C-terminal third of p300. Expression was induc ed by the addition of doxycycline [Dox; 1 mu g/mL; 5% fetal bovine serum (F BS)]. The percentage of apoptosis was determined using the TUNEL technique. Specific protein-protein interactions were determined by Western blot anal ysis of immunoprecipitated complexes. Cells were treated with 5% FBS or wit h H2O2 (500 mu mol/L, 1 h) with and without Dox. Results. The expression of p300-C resulted in increased susceptibility to l ow serum-induced (20.0 +/- 4.6 vs. 3.0 +/- 1.7%) and to H2O2-induced apopto sis (75.3 +/- 13.3 vs. 50.8 +/- 6.5%) compared with controls. Immunoprecipi tation of p300-C showed an interaction with the transcription factor c-Fos, which was enhanced by H2O2 treatment. Expression of the p300-N resulted in a rescue (34.8 +/- 6.4 vs. 50.8 +/- 6.5%) from H2O2-induced apoptosis comp ared with controls. P300-N was shown to form a complex with the transcripti on factor nuclear factor-kappa B (NF-kappa B). Conclusions. The data indicate that endogenous p300 is involved in apoptosi s in mesangial cells. We propose that interference or enhancement of endoge nous p300 function, by expression of exogenous fragments, can alter interac tions with c-Fos or NF-kappa B and modulate signals during cellular stress.