Figure 4.

Progression of flagellum abnormalities in germ cell–specific Lrrc8a^−/− mice. A, fluorescent labeling of Lrrc8a^lox/lox control and GC-Δ8A spermatozoa isolated from cauda epididymis. Acrosomal cap labeled with PNA (red), the mitochondrial sheath with MitoTracker® (green) and nuclei with DAPI (blue). B, green spermatozoa (expressing EGFP-tagged acrosin and DsRed2-tagged mitochondria) isolated from cauda epididymis of Lrrc8a^lox/lox control and GC-Δ8A green sperm mice. Sperm were fixed and stained with anti-GFP (green) and anti-DsRed (red) antibodies. DAPI reveals nuclei (blue). C, immunohistochemical analysis of testes from Lrrc8a^lox/lox control and GC-Δ8A green sperm mice. Magnified regions show elongated mitochondrial sheaths of sperm in both genotypes. D, immunohistochemical analysis of cauda epididymis from Lrrc8a^lox/lox control and GC-Δ8A green sperm mice. Magnified regions show elongated mitochondrial sheaths in Lrrc8a^lox/lox control mice and mitochondria forming a compact mass in GC-Δ8A mice. E, fluorescent labeling of spermatozoa isolated from testes and different epididymal compartments of Lrrc8a^lox/lox control and GC-Δ8A mice, showing progression of flagellar abnormalities. Mitochondrial sheaths indicated by black arrowheads. White arrowheads highlight excess cytoplasm. Acrosomal cap labeled with PNA (red) and nuclei with DAPI (blue). Scale bars, 5 μm (A, B, and E), 10 μm (D), and 20 μm (C). Mito, mitochondria.