Figure 5
Huwe1 is not essential for completion of the later stages of meiosis. (a) Huwe1 is localized to the sex body and telomeres in pachytene spermatocytes. Representative immunofluorescence image from a chromosome spread prepared from a WT testis at 28 dpp stained for Huwe1 (green) and SYCP3 (red). White arrow points to a Huwe1 + telomere end, yellow arrowhead points to the Huwe1 + sex body. Scale bar = 20 μm. (b) The Spo11-Cre KO animals have normal spermatogenesis. Hematoxylin and eosin stained testicular tissue sections from adult WT and Spo11-Cre KO. The yellow arrowheads indicate spermatids in the lumen of the seminiferous tubules. Scale bar = 90 μm (top panel). (c) The Spo11-Cre KO mice show the retention of γH2AX on the autosomes after homologous recombination. Representative immunofluorescence images of chromosome spreads prepared from WT and Spo11-Cre KO testes at 28 dpp stained for γH2AX (green) and SYCP3 (red) (left panel). White arrow indicates γH2AX foci on an autosome of a pachytene spermatocyte, an abnormality significantly more common in the Spo11-Cre KO testes. Quantification of different cell types from the images and of retention of γH2AX (right panel) WT, n = 4 (402 spreads), Spo11-Cre KO, n = 6 (613 spreads). (d) Spo11-Cre KO of Huwe1 did not lead to increased apoptosis. Shown are typical images from TUNEL assay performed on adult testes sections of WT and Spo11-Cre KO testes (left panel). Scale bar = 90 μm. Quantification from images (right panel) WT, n = 4 (335 tubules), KO, n = 4 (466 tubules). (e) RNA polymerase II remains excluded from the XY body of the Spo11-Cre KO spermatocytes. Representative immunofluorescence images of chromosome spreads prepared from WT and Spo11-Cre KO testes at 28 dpp stained for γH2AX (red) and RNA polymerase II (green) (n = 5). The white arrows indicate the absence of RNA polymerase II stain on the XY body. Scale bar = 20 μm. (f) Normal meiotic sex chromosome inactivation (MSCI) in the Spo11-Cre KO mice. Q-PCR measurements of mRNA levels of selected X-linked and Y-linked genes (n = 5). Data are represented as mean ± SEM. Student’s t-test. *p < 0.05.
