Sp1 and Sp3 activate the testis-specific histone H1t promoter through the H1t/GC-box

The testis-specific linker histone H1t gene is transcribed exclusively in mid to late pachytene primary spermatocytes. Tissue specific expression of the gene is mediated in large part through elements located within the proximal promoter. Previous work in transgenic animals showed that a unique 40 bp promoter element designated H1t/TE is essential for spermatocyte-specific expression. The H1t/TE element contains a GC-box, which is a perfect consensus binding site for members of the Sp family of transcription factors. We have shown that Sp1 and Sp3 are present in testis cells from 9-day-old and adult rats and in pachytene primary spermatocytes and early spermatids and that they can bind to the H1t/GC-box. Mutagenesis of the GC-box reduced H1t promoter activity. Furthermore, a CpG dinucleotide within the GC-box was totally unmethylated in rat testis primary spermatocytes where the gene is transcribed but it was methylated in liver where the gene is silenced. These previous studies supported the importance of the GC-box and suggested that Sp transcription factors contribute to expression of the H1t gene. In this study, we show that co-transfection of Sp1 and Sp3 expression vectors leads to an upregulation of histone H1t promoter activity in several cell lines including testis GC-2spd cells. However, very low H1t promoter activity is seen in GC-2spd cells grown at 39 degrees C, which correlates with lower levels of Sp1 and Sp3 in these cells grown at this elevated temperature. Upregulation of the H1t promoter by Sp1 and Sp3 was also seen in cotransfected NIH3T3 and C127I cell lines. On the other hand, co-transfection of the Sp1 and Sp3 expression vectors does not lead to upregulation of activity of the cell-cycle dependent histone H1d promoter.