Figure 7: Dynamic interaction of DNA polymerase β and HSP90 with XRCC1 regulates BER sub-pathway choice.
From: HSP90 regulates DNA repair via the interaction between XRCC1 and DNA polymerase β

(a) Cell viability in response to MNNG, H2O2 or IR. Left panel: LN428/MPG or LN428/MPG/XRCC1-KD cells expressing Flag-Polβ(WT), Flag-Polβ(TM) or EGFP after MNNG treatment, as measured by the MTS assay 48 h after exposure. Results indicate the mean±s.d. of three independent experiments. Middle panel: LN428 or LN428/XRCC1-KD cells expressing Flag-Polβ(WT), Flag-Polβ(TM) or EGFP after H2O2 treatment, as measured by the CyQuant assay 10 days after exposure. Results indicate the mean±s.d. of three independent experiments. Right panel: Clonogenic survival assay of LN428/Polβ-KD, LN428/Puro and LN428/XRCC1-KD cells after exposure to IR. Data points represent means and s.d. of at least three experiments, each performed in triplicate. The arrows indicate the extent of the requirement for Polβ (red) or XRCC1 (blue). Plots show the relative surviving fraction as compared with untreated cells. (b) Polβ/XRCC1 and XRCC1/HSP90 heterodimer ratios in proliferating (P) and confluent (C) MEFs (92TAg) were probed for heterodimer formation by IP of XRCC1 (XRCC1-Ab) and probing for XRCC1, HSP90 and Polβ by immunoblot. A representative immunoblot is shown. *¼ input IP product for XRCC1; L, longer exposure time; S, short exposure time. Bar graphs are plotted with mean±s.d. of two independent experiments. (c) The relative ratio of Polβ/XRCC1 and HSP90/XRCC1 was quantified following IP of XRCC1 and analysis for Polβ or HSP90 (Supplementary Fig. 7C,E) from proliferating and confluent cells (92TAg MEFs) treated with different DNA-damaging agents. Bar graphs are plotted with mean±s.d. of two independent experiments. (d) Proposed model for the dynamic regulation of the stability and degradation of Polβ and XRCC1. Varied cellular conditions, such as HSP90 phosphorylation, alterations in the expression or function related to the cell cycle or DNA damage response and cell-type specificity promote the formation of one of the two heterodimers. Conditions that increase Polβ/XRCC1 levels would favour Polβ-dependent BER, whereas conditions that increase XRCC1/HSP90 levels would result in a preference for XRCC1-dependent or Polβ-independent BER. The degradation of XRCC1 is regulated by HSP90 and CHIP.