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. 2024 Jun 12;121(25):e2320995121. doi: 10.1073/pnas.2320995121

Fig. 1.

Fig. 1.

RNF212B colocalizes with RNF212, its loading to the chromosome axis is dependent on synapsis and is independent of Spo11-induced DSB. (A) Colabeling of RNF212B and SYCP3 from Zygonema to Diplonema. (B) Colabeling of spermatocytes spreads with SYCP3, central element protein SYCE1, and RNF212B. Arrow in (A) and (B) indicate the positive labeling of the PAR. (C) Plot quantitation of the number of RNF212B foci. LZ (Late Zygonema), EP (early Pachynema), MP (mid pachynema), and D (Diplonema). (D) Colabeling of RNF212B and SYCP3 in human and dog spermatocytes showing conserved pattern of localization onto the chromosome axes. (E) STED microscopy images of RNF212 and RNF212B colocalization in pachytene spermatocytes. (F) Double immunostaining of RNF212B and RNF212. Immunofluorescence signal levels were measured on synapsed chromosome axes. Upper plot represents normalized signal intensity profiles of RNF212B (magenta) with RNF212 (green). Lower plot shows regression analysis of the correlation between each pair corresponding to chromosome axes. Each of the dots represents the RNF212B fluorescence intensity in axes “x” and RNF212 fluorescence intensity in axe “y” for a given point along the AE. (G) STED images of pachynema chromosome immunostained for SYCP3 and RNF212B. (H) Double immunolabeling of RNF212B and SYCP3 in arrested spermatocytes of Rec8−/−, Rad21l−/−, Six6os1−/−, and Spo11−/−. Bar in panels (A), (B), (D), and (H) 10 μm. Bars in panels (G) and (F) represents 2.5 μm. All experiments have been carried out in at least three mice and 15 cells per mouse.