Human cells have a finite number of divisions before the activation of cell death or growth arrest. Telomeres are clock-like in that they define this limit as they shorten with each successive cell division. To overcome mortality due to telomere shortening, cancer cells can reactivate telomerase, an enzyme that is expressed in embryonic and adult stem cells, to maintain telomere length and enable infinite cellular divisions. One mechanism of reactivating telomerase in cancer cells is acquisition of mutations in the TERT promoter. In fact, mutations in the TERT promoter (mainly the two hotspot mutations G228A/C228T and G250A/C250T) are the most common non-coding mutations found in human cancers1-5.
But how do mutations in a promoter, a region that is not coding for proteins, activate telomerase? In 2015, Bell et al. found that these hotspot mutations generate an ETS transcription factor binding motif in the TERT promoter which, together, with a native ETS motif recruit the ETS transcription factor GABP to bind and selectively activate the mutant TERT promoter. Functionally, these two ETS binding motifs are positioned at an ideal distance within the TERT promoter for the binding of the GABP tetramer (consisting of two GABPA and two GABPB1 subunits), thus initiating TERT expression, which results in reactivating telomerase activity6.
What’s novel about our paper
We found short duplications of wild type sequence in the TERT promoter in 7 cancer types that typically display a high frequency of TERT promoter mutations, but these were mutually exclusive events. All 22 duplications that we identified had a similar length and insertion site. But most importantly, all duplications created a novel ETS binding motif in the ideal position for GABP to bind as a tetramer. We show that the duplication, inserted adjacent to the native ETS motif within in the TERT promoter, increases TERT promoter activity and that GABP is binding to the TERT duplication to a similar level as to the TERT hotspot mutations (Figure 1). In a patient with glioblastoma, a tumor that is known to acquire TERT promoter mutations at the early stage of tumorigenesis, we could show that the TERT promoter duplication was clonal (found in every tissue sample analyzed) and TERT expression was at a similar level as compared to glioblastomas with hotspot TERT promoter mutations.
In summary, we show that TERT promoter duplications and TERT promoter hotspot mutations share a common mechanism to activate telomerase through the recruitment of GABP. These results emphasize GABP’s role as an essential activator of the mutant TERT promoter in cancer, suggesting a strong selective pressure for this mechanism of immortality. Furthermore, our data show that TERT promoter duplications are functionally equivalent and should be valued equally pathogenic as the hotspot mutations G228A/C228T and G250A/C250T. This holds true especially for cancers where TERT promoter mutations are crucial for diagnosis (e.g. diffuse IDH-wildtype gliomas)7.
Figure 1. TERT promoter duplications mimic hotspot mutations for GABP tetramer recruitment. (a) Schematic of the wildtype TERTp. The native ETS motif is shown relative to the TERT ATG translational start site. GABPB1L-GABPA heterodimer is not bound to the native ETS motif and an inactive TERTp. (b) Schematic of the TERTp with a hotspot mutation. The native ETS motif and de novo ETS motifs are shown relative to the TERT ATG (denoted by the arrow). Distance in base pair (bp) between the native ETS motif and de novo motifs (hotspot mutation) is shown along with the associated helical turns (HT). GABPB1L-GABPA tetramer binding to the native ETS motif and de novo ETS motif (G228A, hotspot mutation) to activate the TERTp. (c) Schematic of the TERTp duplication. Native ETS motif and de novo ETS motif (c.-100_-79, duplication) are shown relative to the TERT ATG. Distance (bp) between the native ETS motif and de novo ETS motif (duplication) is shown along with the associated helical turns (HT). GABPB1L-GABPA tetramer binding to the native ETS motif and de novo ETS motif to activate the TERTp. The figures were created with BioRender (a-c).
1 Huang, F. W. et al. Highly recurrent TERT promoter mutations in human melanoma. Science 339, 957-959, doi:10.1126/science.1229259 (2013).
2 Horn, S. et al. TERT promoter mutations in familial and sporadic melanoma. Science 339, 959-961, doi:10.1126/science.1230062 (2013).
3 Killela, P. J. et al. TERT promoter mutations occur frequently in gliomas and a subset of tumors derived from cells with low rates of self-renewal. Proc Natl Acad Sci U S A 110, 6021-6026, doi:10.1073/pnas.1303607110 (2013).
4 Arita, H. et al. TERT promoter mutations rather than methylation are the main mechanism for TERT upregulation in adult gliomas. Acta Neuropathol 126, 939-941, doi:10.1007/s00401-013-1203-9 (2013).
5 Arita, H. et al. Upregulating mutations in the TERT promoter commonly occur in adult malignant gliomas and are strongly associated with total 1p19q loss. Acta Neuropathol 126, 267-276, doi:10.1007/s00401-013-1141-6 (2013).
6 Bell, R. J. et al. Cancer. The transcription factor GABP selectively binds and activates the mutant TERT promoter in cancer. Science 348, 1036-1039, doi:10.1126/science.aab0015 (2015).
7 Louis, D. N. et al. The 2021 WHO Classification of Tumors of the Central Nervous System: a summary. Neuro Oncol 23, 1231-1251, doi:10.1093/neuonc/noab106 (2021).