Activation of telomerase is linked to tumorigenesis and has been observed in a variety of human tumors. Previous reports demonstrated that p53 represses human telomerase reverse transcriptase (hTERT), a key component for telomerase activity. The p73 protein displays a tumor suppressor activity similar to p53. In the present study, we examined the effect of transactivation competent p73 isoforms on hTERT expression in p53-negative human H1299 cells. Overexpression of C-terminal p73 isoforms (α, β, γ, δ) resulted in a clear down-regulation of hTERT promoter activity. The strongest inhibitory effect, comparable with p53, was observed for p73β. Moreover, suppression of hTERT expression was also mediated by endogenous p73 after activation of E2F1 in H1299ER-E2F1 cells. Mutations in the Sp1 transcription factor-binding sites of the proximal core promoter region significantly abolished p73-induced repression, suggesting that the effect is mediated by Sp1. Finally, we demonstrate that p73 directly interacts with Sp1, suggesting that formation of a p73-Sp1 complex is the underlying mechanism for p73-triggered inhibition of hTERT expression. Our findings provide additional evidence that p73 mimics p53 in many aspects in cells lacking functional p53, thereby contributing to tumor surveillance.