Priority Research Paper Volume 12, Issue 18 pp 17761—17785

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-MYH^WT, GFP-MYH^V315A, and GFP-MYH^Q324H along with KR-TRF1 were expressed in MEF cells to determine their association with damaged telomeres. The association of GFP-MYH^V315A, but not GFP-MYH^Q324H, 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.