DSB-2 marked nuclei require RAD-50 for formation of RAD-51 foci after irradiation.
Immunofluorescence images of rad-50 (A) and htp-1; rad-50 (B) mutant gonads from the distal pre-meiotic region to end of pachytene, stained with DAPI and antibodies that recognize DSB-2 and RAD-51. Worms were fixed and stained 1 hour after exposure to 5 kRad of gamma-irradiation. A reciprocal relationship is observed between DSB-2 and RAD-51 immunolocalization: in nuclei where DSB-2 signal is detected on chromatin, formation of irradiation-induced RAD-51 foci is inhibited, and in nuclei where IR-induced RAD-51 foci are present, DSB-2 is absent. The zone of DSB-2 staining/RAD-51 inhibition is indicated by brackets. (Occasional bright RAD-51 foci in the “inhibited” zone are thought to represent pre-existing DNA damage acquired during mitotic cell cycles in mutants lacking RAD-50, as they are both irradiation- and SPO11-independent .) Arrowheads point to examples of nuclei that retain DSB-2 staining/RAD-51 inhibition in a region of the germ line where their neighbors do not. Scale bar, 15 µm. While the zone of DSB-2 staining/RAD-51 inhibition in the irradiated rad-50 single mutant extends from meiotic prophase entry to late pachytene, the zone of DSB-2 staining/RAD-51 inhibition is limited to a smaller domain from meiotic entry to the early pachytene region in the irradiated htp-1; rad-50 double mutant.