The hTERT core promoter forms three parallel G-quadruplexes

Abstract

The structure of the 68 nt sequence with G-quadruplex forming potential within the hTERT promoter is disputed. One model features a structure with three stacked parallel G-quadruplex units, while another features an unusual duplex hairpin structure adjoined to two stacked parallel and antiparallel quadruplexes. We report here the results of an integrated structural biology study designed to distinguish between these possibilities. As part of our study, we designed a sequence with an optimized hairpin structure and show that its biophysical and biochemical properties are inconsistent with the structure formed by the hTERT wild-type sequence. By using circular dichroism, thermal denaturation, nuclear magnetic resonance spectroscopy, analytical ultracentrifugation, small-angle X-ray scattering, molecular dynamics simulations and a DNase I cleavage assay we found that the wild type hTERT core promoter folds into a stacked, three-parallel G-quadruplex structure. The hairpin structure is inconsistent with all of our experimental data obtained with the wild-type sequence. All-atom models for both structures were constructed using molecular dynamics simulations. These models accurately predicted the experimental hydrodynamic properties measured for each structure. We found with certainty that the wild-type hTERT promoter sequence does not form a hairpin structure in solution, but rather folds into a compact stacked three-G-quadruplex conformation.

hTERT Promotes CRC Proliferation and Migration by Recruiting YBX1 to Increase NRF2 Expression

“…hTERT is the catalytic subunit of telomerase, which is located at the ends of chromosomes and protects them from degradation. It has been reported that hTERT and telomerase activity play a vital role in cell immortalization [32], transformation [33], proliferation [34] and metastasis [35]. A previous study indicated that hTERT could affect cancer development via activity not involving telomere elongation [36].…”

Télomères et télomérase : des cibles toujours pertinentes en oncologie ?

“…La régulation transcriptionnelle d'hTERT est complexe, avec une soixantaine d'activateurs ou de répresseurs recensés. La formation de G-quadruplex en tandem par un fragment de 68 nucléotides riche en G de ce promoteur a récemment été étudiée par une approche de biologie structurale intégrée [Monsen 2020]. Mais, que l'on s'intéresse ou non à la formation de conformations inhabituelles d'acides nucléiques dans ce gène, il est frappant de constater que des mutations récurrentes dans ce promoteur sont présentes dans certaines tumeurs solides en particulier dans des gliomes, mélanomes et hépatocarcinomes (pour revue : [Hannen 2018]).…”

An Updated Focus on Quadruplex Structures as Potential Therapeutic Targets in Cancer

“…The hTERT core promoter, approximately from −180 to +1 of its TSS, contains twelve tracts with three or more guanines, which enable the formation of a 68 nucleotide (nt) long G4 and a structure with three stacked parallel units. The formation of this unusual tandem structure disables all three critical binding sites for SP1 transcription factor, thus dramatically downregulating hTERT expression and exerting telomere shortening [ 68 ].…”