Figure 6.Human V315A mutant MYH protein expressed in mouse cells cannot associate with damaged telomeres and interrupting the MYH interactions with its partners can increase cell’s sensitivity to H2O2 and/or elevate cellular 8-oxoG levels. (A) Sequence alignment of the IDC regions of eukaryotic MYH proteins. V315 and Q324 of hMYH are important for interaction with Hus1 [55, 66] (red stars). However, residue Q324 of hMYH is dispensable for interacting with SIRT6 [40]. (B) and (C) GFP-MYHWT, GFP-MYHV315A, and GFP-MYHQ324H along with KR-TRF1 were expressed in MEF cells to determine their association with damaged telomeres. The association of GFP-MYHV315A, but not GFP-MYHQ324H, with damaged telomeres is substantially attenuated. (D) HEK-293T cells transformed with pEGFP-N1 vector or vector with IDC sequences were treated with 700 mM H2O2 for 1 h and recovered in fresh medium for two days. Cell viability was measured as described in Materials and Methods. (E) HEK-293T cells transformed with pEGFP-N1 vector or vector with IDC sequences were treated with 700 mM H2O2 for 1 h and recovered in fresh medium for 1 h. 8-oxoG levels were measured. Each average value was obtained by subtracting the value of cells containing pEGFP-N1 vector without H2O2 treatment from H2O2-treated cells with listed plasmids. *, **, and *** represent P <0.1, P <0.05, and P < 0.01, respectively.
