The Regulation of hTERT by Alternative Splicing

Telomeres are non-coding DNA hexameric repeats (TTAGGG in mammals) located at the ends of linear chromosomes that, along with their associated proteins, protect against the loss of genomic material during cell division and prevent the recognition of chromosome ends as double-strand breaks. Human telomeres shorten with continued cell proliferation but are maintained by human telomerase reverse transcriptase (hTERT), an enzyme that synthesizes telomeric repeats using an RNA template. The regulation of telomerase has been studied at many levels--from epigenetic and transcriptional regulation to the alternative splicing of hTERT pre-mRNA into catalytically inactive splice variants. Our hypothesis is that if the regulation of telomerase reverse transcriptase splicing is necessary for telomere length homeostasis, altering telomerase splicing to decrease the production of full-length hTERT and will result in decreased telomerase activity and subsequently telomere shortening. We focused our efforts on identifying splicing factors are involved in hTERT splicing and characterized the role of two splicing factors, NOVA1 and PTBP1, in regulation of hTERT splicing in non-small cell lung cancer cells. We show that these splicing factors are important for full-length hTERT, telomerase activity and telomere length maintenance in vitro. Xenograft studies suggest that NOVA1 is also important for tumor growth in vivo. We found that these splicing factors are able to directly interact with hTERT in a region our group previously identified to be important for hTERT splicing. Altogether, our work suggests that splicing factors are important for hTERT regulation and telomerase activity in cancer. Since telomerase activity is undetectable in most somatic tissues but is increased in the vast majority of human cancers, dependence on telomerase represents a key vulnerability in cancer tissues which could be therapeutically targetable.