Skip to main content
. 2006 Nov 20;103(48):18314–18319. doi: 10.1073/pnas.0606528103

Fig. 4.

Fig. 4.

Delivery of purified PAR polymer into neurons induces AIF translocation and nuclear condensation. (A) Representative confocal images of AIF translocation and nuclear condensation after BioPorter mediated delivery of purified PAR polymer into wild-type (WT) cortical cultures with and without pretreatment with PARG. AIF (red) is seen in well defined mitochondria like structures in the neuronal cell body as well as long axonal processes and no AIF is seen in the nucleus (blue) in control or PAR + PARG-treated cultures, whereas a clear nuclear translocation and shrinkage of AIF (Pink) is seen in PAR treated cultures. (Scale bar, 20 μm.) (B) Quantitative analysis of AIF translocation and nuclear condensation in the neurons treated with PAR polymer with and without pretreatment with PARG or PD1. BioPorter mediated delivery of poly(A) serves as a control. ∗, P < 0.05, Student's t test. Data represent the mean ± SD; n = 3. (C) Western blots of subcellular fractions of WT cultures, showing nuclear AIF translocation after PAR delivery. Synthetic polymer, poly(A), or enzymatic digestion of PAR with PARG or PD1 did not induce any nuclear AIF translocation. Integrity of mitochondrial fraction (M) and nuclear fraction is shown with antibodies against MnSOD or histone, respectively. (D) Nuclear translocation of AIF accompanies decrease of AIF in mitochondrial fraction. PAR polymer was delivered into cortical neurons using BioPorter for 6 h and subjected to subcellular fractionation into nuclear and mitochondrial fractions. MnSOD and histone were probed as mitochondrial and nuclear markers, respectively. PAR polymer induces the nuclear translocation of AIF, concurrent with loss of AIF in the mitochondrial fraction. (E) Representative confocal images of AIF translocation and nuclear condensation after BioPorter mediated delivery of purified PAR polymer into PARP-1 knockout (KO) cultures with and without pretreatment with PARG. (Scale bar, 20 μm.) (F) Western blots of subcellular fractions of PARP-1 KO cultures, showing nuclear AIF translocation after PAR delivery. Synthetic polymer poly(A) or enzymatic digestion of PAR with PARG or PD1 did not induce any nuclear AIF translocation. These experiments have been replicated in separate experiments at least three times with similar results.