Transcriptional Promiscuity of the Human α-Globin Gene

Abstract

The human alpha-globin gene displays the unusual property of transcriptional promiscuity: that is, it functions in the absence of an enhancer when transfected into nonerythroid cell lines. It is also unusual in that its promoter region lies in a hypomethylated HpaII tiny fragment (HTF) island containing multiple copies of the consensus sequence for the SP1-binding site. We have investigated whether there is a relationship between these two observations. First, we investigated the mouse alpha-globin gene since it does not lie in an HTF island. We have demonstrated that it was not transcriptionally promiscuous. Second, we studied the transcriptional activity of the human alpha-globin gene in the absence of the GC-rich region containing putative SP1-binding sites and found a small (two- to threefold) but consistent positive effect of this region on transcriptional activity in both nonerythroid and erythroid cell lines. However, this effect did not account for the promiscuous nature of the human alpha-globin gene. We found that in a nonreplicating system, the human alpha-globin gene, like that of the mouse, required a simian virus 40 enhancer in order to be transcriptionally active in nonerythroid and erythroid cell lines. Since we only observed enhancer independence of the human alpha-globin gene in a high-copy-number replicating system, we suggest that competition for trans-acting factors could explain these results. Finally, our experiments with the erythroid cell line Putko suggest that there are no tissue-specific enhancers within 1 kilobase 5' of the human alpha-globin cap site or within the gene itself.

Transcription of the human U2 snRNA genes continues beyond the 3′ box in vivo

“…For each 90 cm dish of HeLa cells, a 5 μg aliquot of each U2 construct was co-transfected with 500 ng of VAI using Lipofectamine (Gibco-BRL) as recommended by the manufacturers, and cytoplasmic RNA was collected as described by Whitelaw et al (1989). RNase protection was carried out as described by Eggemont and Proudfoot (1993).…”