Figure 4

Expression of an alternative Gpi1 transcript in mouse spermatogenic cells. (A) Diagram of RT-PCR approach used to distinguish expression of Gpi1-related transcripts. Gray arrows denote the primer set used to differentiate transcripts containing alternatively spliced exons 5 and 6 (boxes with diagonal lines). Black arrows denote the Gpi1-rs1-specific primer set. (B) Transcripts from Gpi1 were detected in all mouse tissues and isolated testicular cells. Gpi1-rs1 was amplified from genomic DNA to identify the expected size of PCR products from Gpi1 transcripts not containing exons 5 and 6. A product of the same size was detected in isolated pachytene spermatocytes (PS) and round spermatids (RS), but not condensing spermatids (CS). This PCR fragment appears to be derived from Gpi1_v2, since Gpi1_rs1- specific primers did not amplify a product. (C) A smaller GPI1_V2 protein was not detected by western analysis using a polyclonal antibody raised against human GPI1. A larger protein product was seen in isolated testicular cell, but not in mouse or human (Hs) sperm. S/N fraction contains proteins solubilized from sperm tail following brief sonication and centrifugation. Tail fraction contains proteins left insoluble following sonication and centrifugation.