Rad21 depletion increases tert expression and γH2AX TIFs in brain tumors. (A) Representative fluorescent microscope image of Q-FISH and γH2AX immunostaining in RAS and rad21;RAS tumor cells. Scale bar: 5 µm. Insets show examples of co-localization of the Q-FISH and antibody signals, in foci pointed by the yellow arrows. (B) Representative fluorescent microscope image of Q-FISH and Rad51 immunostaining in RAS and rad21;RAS tumor cells. Scale bar: 5 µm. Insets show examples of co-localization of the Q-FISH and antibody signals, in foci pointed by the yellow arrows. (C) Quantification of the numbers of telomere foci which are positive for γH2AX immunostaining (nuclei = 27), also known as TIFs, in RAS and rad21;RAS brain tumors cells. Each dot represents a nucleus. ** p < 0.01. (D) Quantification of the numbers of telomere foci which are positive for rad51 immunostaining (RAS nuclei = 22; rad21;RAS nuclei =19) in in RAS and rad21;RAS brain tumors cells. ns = not significant. Each dot represents a nucleus. (E) Expression of zebrafish tert mRNA in brain tumors measured by RT-qPCR. Values were normalized to rps11 mRNA levels and are relative to tert expression in control brains (dashed line set at 1.0). * p < 0.05. Each dot represents a sample. (F) Relative telomerase activity measured by Q-TRAP in control, RAS, and rad21;RAS brains, using 1 μg of protein extracts. RNase treatment (+) was used as a negative control to confirm the specificity of the assay; each dot represents one sample.
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