title: Reporter mice express green fluorescent protein at initiation of meiosis in spermatocytes.
abstract: Transgenic mice were generated using a heat shock protein 2 (Hspa2) gene promoter to express green fluorescent protein (GFP) at the beginning of meiotic prophase I in spermatocytes. Expression was confirmed in four lines by in situ fluorescence, immunohistochemistry, western blotting, and PCR assays. The expression and distribution of the GFP and HSPA2 proteins co-localized in spermatocytes and spermatids in three lines, but GFP expression was variegated in one line (F46), being present in some clones of meiotic and post-meiotic germ cells and not in others. Fluorescence activated cell sorting (FACS) was used to isolate purified populations of spermatocytes and spermatids. Although bisulfite sequencing revealed differences in the DNA methylation patterns in the promoter regions of the transgene of the variegated expressing GFP line, a uniformly expressing GFP reporter line, and the Hspa2 gene, these differences did not correlate with variegated expression. The Hspa2-GFP reporter mice provide a novel tool for studies of meiosis by allowing detection of GFP in situ and in isolated spermatogenic cells. They will allow sorting of meiotic and post-meiotic germ cells for characterization of molecular features and correlation of expression of GFP with stage-specific spermatogenic cell proteins and developmental events.
authors.affiliation: Gamete Biology Section, Laboratory of Reproductive and Developmental Toxicology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina.
authors.forename: Paula R
authors.initials: PR
authors.lastname: Brown
authors.forename: Fanny
authors.initials: F
authors.lastname: Odet
authors.forename: Carl D
authors.initials: CD
authors.lastname: Bortner
authors.forename: Edward M
authors.initials: EM
authors.lastname: Eddy
mesh_terms: D000818:Animals; D019175:DNA Methylation; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008540:Meiosis; D051379:Mice; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013087:Spermatids / Q000166:cytology / Q000378:metabolism; D013090:Spermatocytes / Q000166:cytology* / Q000378:metabolism*
chemical_list: D018840:HSP70 Heat-Shock Proteins; C490434:Hspa2 protein, mouse; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins
keywords: fluorescence activated cell sorting; heat shock protein; spermatogenesis; variegated gene expression