FIG. 7.

Protein binding at the FP sequences in germ cell extracts. (A) Germ cells from male (♂) and female (♀) 15.5-dpc fetuses were obtained by flow sorting of EGFP-expressing germ cells. The populations shown higher in the graphs are comprised of germ cells, while the lower populations are comprised of gonadal somatic cells. SSC, side scatter. (B) Assessment by semiquantitative RT-PCR of relative Rxrα and Erβ transcript levels in germ cells. RNA prepared from equal numbers of purified germ cells (GC) or somatic cells (som) of 15.5-dpc female or male gonads was subjected to RT. Aliquots of the PCR were subjected to increasing number of cycles in the linear amplification range. A representative experiment is shown. (C) Male germ cell-specific complex at the FP4 sequence. (D) Protein-DNA complexes of identical levels of mobility are present in nuclear extracts from PEF and in extracts from 15.5-dpc male germ cells. Female germ cells from the same stage of development do not contain this DNA binding activity. The complex is also present in 18.5-dpc male germ cells and in spermatogonia (SG) but not in pachytene spermatocytes (PS). Arrowheads point to the specific gel shifts. Antibody supershift assays indicate the presence of RXRα and ERβ in the complex in male germ cells (indicated by asterisks).