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Telomerase is a ribonucleoprotein enzyme with an essential RNA component. Embedded within the telomerase RNA is a template sequence for telomere synthesis. We have characterized the structure of the 5' regions of the human and mouse telomerase-RNA genes, and have found a striking difference in the location of the template sequence: Whereas the 5'-end of the human telomerase RNA lies 45 nt from the telomerase-RNA template sequence, the 5'-end of the mouse telomerase RNA lies just 2 nt from the telomerase-RNA template sequence. Analysis of genomic sequences flanking the 5'-end of the human and mouse telomerase RNA-coding sequences reveals similar promoter-element arrangements typical of mRNA-type promoters: a TATA box-like element and an upstream region containing a consensus CCAAT box. This putative promoter structure contrasts with that of the ciliate telomerase-RNA genes whose structure resembles RNA polymerase III U6 small nuclear RNA (snRNA) promoters. These and other comparisons suggest that, during evolution, both the RNA-polymerase specificity of telomerase RNA-gene promoters and, more recently, the position of the template sequence in the telomerase RNA changed.

Recently, a novel PCR-based technique, differential display (DD), has facilitated the study of differentially expressed genes at the mRNA level. We report here an improved version of DD, which we call Enhanced Differential Display (EDD). We have modified the technique to enhance reproducibility and to facilitate sequencing and cloning. Using EDD, we have generated and verified a catalog of genes that are differentially expressed between young and senescent human diploid fibroblasts (HDF). From 168 genetags that were identified initially, 84 could be sequenced directly from PCR amplified bands. These sequences represent 27 known genes and 37 novel genes. By Northern blot analysis we have confirmed the differential expression of a total of 23 genes (12 known, 11 novel), while 19 (seven known, 12 novel) did not show differential expression. Several of the known genes were previously observed by others to be differentially expressed between young and senescent fibroblasts, thereby validating the technique.