Figure 6

Sly and Asty/Asty(rec) are expressed in spermatids. (a) A multi-tissue northern blot of polyA⁺ RNA probed with Sly clone MTnH-K10 and Asty exon 4 probe. Sly transcripts are restricted to the testis, but Asty/Asty(rec) transcripts are also seen in the heart. (b) A northern blot of total testis RNA from testes of mice aged 12.5-30.5 days postpartum (dpp) hybridized with the same Sly and Asty probes. No transcripts are detectable before 20.5 dpp, suggesting that the transcripts are restricted to spermatid stages. (c) RNA in situ analysis confirming that Sly transcription in the testis is predominantly if not exclusively in spermatids. (d) Diagram summarizing the expression of Sly in spermatids throughout the spermatogenic cycle. The spermatogenic stages starting from the basal layer and reading left to right are: spermatogonia: A1 through to B; meiotic prophase spermatocytes: preleptotene (PL), leptotene (LE), zygotene (Z), pachytene (P), diplotene (D); meiotic divisions (mm); postmeiotic spermatid stages: 1-13. (e) RNA in situ analysis with the Asty exon 4 probe that should detect Asty and Asty(rec) transcripts. The most convincing signal is in round spermatid nuclei, but this is seen with the antisense and sense (control) probes. However, the previously reported cDNA clone BU936708 (Additional data file 3) contains Asty exon 4 sequence in the antisense oritentation and which would hybridize with the sense probe. (f) Exon 1-4 Asty reverse transcriptase polymerase chain reaction from 9.5 dpp to adult testes. Sequencing of the cloned amplification products confirmed the presence of full-length Asty transcripts at 14.5 dpp, and a shorter transcript lacking exon 3 from 22.5 dpp.