Mechanism of dominant-negative telomerase function

Human telomerase uses its integral core components, hTR and hTE RT, to maintain telomeres in many cell types. Expression of a dominant-negative mutant of the catalytic subunit of telomerase, DN-hTE RT, has been shown to cause telomere shortening and ultimately cell death in a number of tumor-derived cell lines. However, the mechanism of dominant-negative hTE RT function and its fate inside the cell are still unknown. In order to understand the effect of the dominant-negative on wild-type hTE RT, each was fused with GFP and expressed in telomerase-positive cells. GFP-DN-hTE RT expression resulted in cytoplasmic exportation and degradation via ubiquitination. Co-expression of wild-type GFP-hTE RT with an untagged DN-hTE RT resulted in decreased wild-type hTE RT levels, export to the cytoplasm, and increased ubiquitination, suggesting that DN-hTE RT complexes with wild-type hTE RT to induce cytoplasmic localization. Based on the cytoplasmic degradation, we propose two new mechanisms of dominant-negative hTE RT, employing the theory of interactive dimerization. First, the heterodimer of DN-hTE RT with wild-type hTE RT is exported to the cytoplasm for ubiquitin-mediated protein degradation, and second, the heterodimer may be degraded at a faster rate than the wild-type hTE RT homodimer. Understanding mechanisms of telomerase degradation will guide future drug design to target sites on telomerase important for catalytic activity and protein stability.