Determining the role of TCAB1 in shaping telomerase function

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Abstract Impaired maintenance of telomeres by telomerase causes cellular senescence and underlies many aging-related diseases in humans. Furthermore, telomere shortening is one of the most reproducible hallmarks of aging in human tissues, suggesting that telomere dysfunction contributes broadly to aging phenotypes in people. The human telomerase enzyme is comprised of a catalytic core – the telomerase reverse transcriptase (TERT) and the telomerase RNA component (TERC) – but also depends upon other holoenzyme proteins that bind the H/ACA element within TERC. During evolution, vertebrate telomerase hijacked this H/ACA sequence from other ancient non-coding RNAs, bringing the dyskerin core protein complex, which recognizes the H/ACA element, and TCAB1, which binds the CAB box element, into the telomerase holoenzyme. Telomerase function is critically dependent upon a series of steps, including assembly of the enzyme, trafficking within the nucleus, recruitment to telomeres and finally catalytic extension of telomeres. Each of these steps is poorly understood in humans and each step can be disrupted by germline mutations in patients with telomere diseases. The dyskerin complex is required for assembly and stability of telomerase. We identified TCAB1 as a protein that interacts with dyskerin and showed that TCAB1 binds the CAB box element common to TERC and to all small Cajal body RNAs (scaRNAs). We previously showed that TCAB1 is required for localization of telomerase in Cajal bodies, required for telomere maintenance and is important in recruitment to telomeres. We recently made the surprising finding that TCAB1 is also critical for telomerase catalytic activity and that TCAB1 is required for proper TERC conformation. Telomerase and other RNPs depend on an RNA molecule that needs to fold in a precise conformation, and this correct conformation represents one of countless potential structures. In this proposal, we pursue the hypothesis that TCAB1 is essential in shaping the telomerase RNA, and that this activity of TCAB1 is conserved in the splicing RNA pathway. We propose that the principal advantages conferred upon telomerase by adopting an H/ACA RNA fate is facilitating assembly and proper folding of TERC, and that numerous downstream steps depend upon this proper folding. We will pursue the following aims: (1) To understand how telomerase catalytic function depends upon TCAB1 (2) To determine how TCAB1 influences telomerase trafficking and recruitment (3) To determine how TCAB1 loss affects scaRNA function and RNA splicing.