Figure 1.
SC-null spermatocytes lack nuclear structures resembling the axial elements or the central region of the SC.
Electron microscopy analysis of nuclei from spermatocytes derived from of wild-type (A, E) and SC-null (B–D) mice. Wild-type pachytene meiocytes show synaptonemal complexes (SC) and normally condensed chromatin (A, E). In SC-null meiocytes, chromatin is less condensed and axial structures are absent (B). The arrow in (B) points to dense regions of centromeric heterochromatin located close to the nuclear envelope. Attachment plates of the nuclear envelope in wild-type and SC-null meiocytes are denoted by arrowheads (C–D). NE, nuclear envelope; XY, XY body.
Figure 2.
SC proteins, but not cohesin proteins, are lost from the chromosome axes in SC-null spermatocytes.
(A) SC-null spermatocytes were stained with antisera against the meiosis-specific cohesins REC8 (red) and SMC1β (red) and the cohesin protein STAG3 (green). Centromeres, labeled by CREST, are shown in white. (B) SC-null spermatocytes were labeled with antisera against the axial element protein SYCP2 (red), the central element protein TEX12 (red) or the central element protein SYCE3 (red). The chromosomal axes are identified by labeling of the cohesion protein STAG3 (green). Centromeres, labeled by CREST, are shown in white. Bars, 10 µm.
Figure 3.
An axial cohesin structure develops in the absence of the SC.
SC-null and wild-type oocytes from prenatal ovaries were labeled with antisera that recognize the cohesion complex (STAG3-red) and the centromeres (CREST-white), and analyzed by immunofluorescent microscopy. Mutant oocytes were classified according to their day of appearance during development (see Materials and Methods). Cohesin cores that showed extensive alignment were found in SC-null oocytes. The cores failed to synapse, as judged by the number of CREST foci seen at late pachytene. The integrity of the cohesion axes rapidly declined from the pachytene stage and onwards in SC-null oocytes. Bars, 10 µm.
Figure 4.
Cohesin proteins label the chromosomal axes in SC-null oocytes, but the SC proteins are lost from the chromosomal axes.
(A) SC-null oocytes were stained with antisera recognizing cohesins REC8 (red), RAD21/RAD21L (magenta) and STAG3 (green). (B) SC-null oocytes were labeled with antisera against the central element proteins SYCE1 and SYCE2 (red). Chromosomal axes were identified by labeling of the cohesion protein STAG3 (green). Centromeres, labeled by CREST, are shown in white. Bars, 10 µm.
Figure 5.
The SYCP1 protein does not form extended fiber-like structures in TEX12/SYCP3 double-null oocytes.
Mutant oocytes were labeled with antisera against SYCP1 (red) and STAG3 (blue). Centromeres were identified by CREST staining (blue). Bar, 10 µm.
Figure 6.
Repair of DNA DSBs are impaired in the absence of the SC.
SC-null and wild-type oocytes from prenatal ovaries were labeled with antisera that recognize the cohesion complex (STAG3-blue) and the centromeres (CREST-blue), and analyzed by immunofluorescent microscopy. (A) H2AX phosphorylation (red) persists in SC-null oocytes until the diplotene stage. (B) Mean intensity of the γH2AX signal in nuclei of wild-type and mutant oocytes. (C) RPA foci (green) co-localize with γH2AX (red) in diplotene stage SC-null oocytes. Bars, 10 µm.
Figure 7.
Temporal expression of RAD51 and DMC1 in wild-type and SC-null oocytes.
Oocytes at different stages of meiotic prophase were labeled with antisera to RAD51 or DMC1 (red). The chromosomal axes were visualized by STAG3 protein labeling (blue). The RAD51 and DMC1 recombination-related proteins disappear from the chromosomal axes in wild-type oocytes by late pachytene. In SC-null oocytes DMC1 show a pattern similar to that seen in wild-type oocytes, while RAD51 foci persist until the diplotene stage. Bar, 10 µm.
Figure 8.
Temporal expression of RPA and MSH4 in wild-type and SC-null oocytes.
Oocytes at different stages of meiotic prophase were labeled with antisera to RPA or MSH4 (red). The chromosomal axes were visualized by STAG3 protein labeling (blue). The RPA and MSH4 recombination-related proteins disappear from the chromosomal axes in wild-type oocytes by late pachytene. In SC-null oocytes MSH4 shows a pattern similar to that seen in wild-type oocytes, while RPA foci persist until the diplotene stage. Bar, 10 µm.
Figure 9.
The DNA recombination process is correctly initiated in SC-deficient oocytes, but the repair process is severely obstructed.
The temporal and spatial distribution of RAD51, DMC1, RPA and MSH4 was analyzed at different stages of meiosis in wild-type, SYCP1-null, TEX12-null, TEX12/SYCP3 double-null and SC-null ovaries. (A) The number of axis-associated RAD51, DMC1, RPA and MSH4 foci in wild-type and mutant oocytes was revealed using immunofluorescent microscopy (Figs. 7–8) and scored. The recombination-related proteins disappear from the chromosomal axes in wild-type oocytes by late pachytene. In mutant oocytes, DMC1 and MSH4 show a similar turnover, while RAD51 and RPA persist to the diplotene stage. (B) The level of interference between MSH4 foci is similar in wild-type, SYCP1-null and SC-null oocytes, as estimated by the value of shape parameter νof gamma-distribution. A value of 1 indicates the absence of interference. Bars, s.e.m.
Figure 10.
The recombination process in SC-null oocytes stops prior to the formation of recombination structures that contain MLH1 or MLH3.
The chromosomal axes were labeled by STAG3 (blue), and centromeres by CREST (blue). MLH1 (red)/MLH3 (green) axis-associated complexes are absent in SC-null oocytes at the pachytene stage. Bars, 10 µm.
Figure 11.
Inactivation of the SYCP3 gene in a SYCP1-null background transiently suppresses oocyte loss.
(A) Sections of ovaries taken from mutant and wild-type females were stained by GCNA (at 1dpp and 8dpp), or with hematoxylin and eosin (at 4 weeks and 8 weeks). Bars, 100 µm. (B) Oocyte numbers in wild-type and mutants animals and the ratio of mutant/wild-type oocytes were scored at 1 day (1 dpp), 8 days (8 dpp), 4 weeks and 8 weeks after birth. Bars, s.d.
Figure 12.
A small number of chiasmata forms in SC-null oocytes.
(A) Wild-type, SC-null and TEX12/SYCP3 double-null oocytes at the first meiotic metaphase stage were stained with DAPI. Arrows indicate bivalents. The occurrence of bivalents strongly suggests that homologous chromosomes are held together by chiasmata. Bars, 10 µm. (B) Percentage of SC-null (n = 55) and TEX12/SYCP3 double-null (n = 57) oocytes that contain 0–3 bivalents per cell.
Table 1.
Phenotypes identified for SYCP1-null, SYCP3-null and SC-null mutant mice.
Figure 13.
RAD51 (red), but not DMC1 (red), is found on the asynapsed axes in wild-type oocytes.
Chromosomal axes are labeled by STAG3 (blue). Bars, 10 µm.
Table 2.
Phenotypes described for SC-null mutants.
Table 3.
The number of cells used for the statistical analysis of the recombination process.