HIV-1 Tat Interacts with and Regulates the Localization and Processing of Amyloid Precursor Protein
Figure 4
HIV-1 Tat alters the processing of APP and increases the levels of β-CTF and Aβ42.
(A) Level of β-CFT was increased in Lenti-Tat, or Lenti-mTat infected cells while the amount of total APP was not changed. U-87 MG cells were transduced with mock, Lenti-Tat, or Lenti-mTat virus and incubated for 14 days. The cell number was counted, and equal numbers of cells from each sample were analyzed in 16.5% Tris-Tricine gels to detect CTF or 8% SDS-PAGE for the detection of APP and β-actin. Both α- and β-CTF were detected with an anti-APP C-terminal antibody (A8717, top panel), and β-CTF was detected with 6E10 (upper middle panel). The total amounts of APP (22C11, lower middle panel) and β-actin (bottom panel) were not changed by the expression of Tat or mutant Tat protein. (B) Aβ42 level was increased in Lenti-Tat and Lenti-mTat infected U-87 MG cells. Average of Aβ42 concentration for 9 days was calculated from individual Aβ42 concentration. The Aβ42 concentrations in Lenti-Tat- or Lenti-mTat-infected cells were increased by 5.58±0.83 (mean ± SE, P<0.005)-fold or 3.64±0.38 (mean ± SE, P<0.005)-fold, respectively, compared with mock-infected cells. (C) Treatment with the neprilysin inhibitor thiorphan further increased the level of Aβ42 in the presence of Tat. Mock or Lenti-Tat-infected U-87 MG cells were cultured for 3 days in the absence or presence of 10 µM thiorphan. Culture supernatant was harvested from each cell culture and analyzed by Aβ42 ELISA. Error bars represent the mean ± SD. *P<0.05, **P<0.001.