ALKBH4 depletion leads to a marked loss of male germ cells

To investigate the physiological role of ALKBH4 and Zatebradine circumvent embryonic lethality of Alkbh4 deletion, we depleted ALKBH4 in mice using a tamoxifen-inducible Alkbh4L/L CreEsr strain. Here, we report that ALKBH4 depletion leads to a marked loss of male germ cells during meiotic prophase and disorganization of the synaptonemal complex. Moreover, we describe the nuclear localization pattern of ALKBH4 in mitotic and premeiotic male germ cells in addition to Sertoli cells. To assess whether the reduced tubular diameter and loss of germinal epithelium were associated with either a reduced entry of cells in spermatogenesis or increased cell death, we performed a 2 hour-pulse of 5-bromo-29-deoxyuridine -incorporation and terminal deoxynucleotidyl transferase -mediated dUTP nick end labeling experiments. Synaptonemal complex protein 3 is one of the major components in the synaptonemal complex. This meiosisspecific protein is essential for the axial element assembly along the chromosomes and it is Rottlerin necessary for accurate synapsis formation. The nuclei of these cells were shrunken and the cells dislocated towards the lumen, indicative of cell death. Similar observations were made using spread pachytene spermatocytes derived from Alkbh4D/D mice, which occasionally lacked a mature synaptonemal complex. Furthermore, staining for the synaptonemal complex component SYCP1 also revealed the presence of pachytene spermatocyte. Selective loss of pachytene spermatocytes in Alkbh4D/D mice led us to examine the expression of ALKBH4 protein throughout spermatogenesis. We have previously characterized the localization of ALKBH4 in the cleavage furrow during cytokinesis. We were not able to assess a detailed cytoplasmic localization pattern for ALKBH4 in the testis sections studied in this report. However, in the nuclei of Sertoli cells, spermatogonia, and zygotene and pachytene spermatocytes of wild-type mice, ALKBH4 localized to distinct structures in euchromatin, often in close association with heterochromatin.In Sertoli cells, a single ALKBH4-rich structure was observed in each nucleus. In spermatogonia, ALKBH4 was distributed in fine granular patches under the nuclear envelope. In preleptotene and early leptotene spermatocytes, ALKBH4 formed 3 to 8 diffuse threads and patches per nucleus.