Structural basis for telomerase catalytic subunit TERT binding to RNA template and telomeric DNA

Mitchell, Meghan; Gillis, Andrew; Futahashi, Mizuko; Fujiwara, Haruhiko; Skordalakes, Emmanuel

doi:10.1038/nsmb.1777

Cited by 185 publications

(295 citation statements)

References 58 publications

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“…Intriguingly, the catalytic domain of telomerase reverse transcriptase (TERT) has structural and functional characteristics in common with picornaviral RNA-dependent RNA polymerases. The catalytic domain of TERT assumes a right-hand conformation, with thumb, palm, and fingers domains encircling the telomerase RNA template and cDNA products (53)(54)(55). Furthermore, TERT uses a template-dependent reiterative transcription mechanism to synthesize repetitive DNA sequences of variable length at the 3= end of eukaryotic chromosomes (56).…”

Section: Discussionmentioning

confidence: 99%

Structural Features of a Picornavirus Polymerase Involved in the Polyadenylation of Viral RNA

Kempf

Kelly²,

Springer

et al. 2013

J Virol

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Picornaviruses have 3= polyadenylated RNA genomes, but the mechanisms by which these genomes are polyadenylated during viral replication remain obscure. Based on prior studies, we proposed a model wherein the poliovirus RNA-dependent RNA polymerase (3D pol ) uses a reiterative transcription mechanism while replicating the poly(A) and poly(U) portions of viral RNA templates. To further test this model, we examined whether mutations in 3D pol influenced the polyadenylation of virion RNA. We identified nine alanine substitution mutations in 3D pol that resulted in shorter or longer 3= poly(A) tails in virion RNA. These mutations could disrupt structural features of 3D pol required for the recruitment of a cellular poly(A) polymerase; however, the structural orientation of these residues suggests a direct role of 3D pol in the polyadenylation of RNA genomes. Reaction mixtures containing purified 3D pol and a template RNA with a defined poly(U) sequence provided data consistent with a template-dependent reiterative transcription mechanism for polyadenylation. The phylogenetically conserved structural features of 3D pol involved in the polyadenylation of virion RNA include a thumb domain alpha helix that is positioned in the minor groove of the double-stranded RNA product and lysine and arginine residues that interact with the phosphates of both the RNA template and product strands.P icornaviruses, like many other positive-strand RNA viruses, have RNA genomes with variable-length 3= poly(A) tails (1). The poly(A) tails of picornaviruses are important for viability (2), with the length of the poly(A) tails influencing the magnitudes of both viral mRNA translation and RNA replication (3, 4). RNA sequences and structures within the 3= nontranslated region are reported to influence both the length of poly(A) tails in picornaviral RNA (5) and the efficiency of virus replication (6, 7). Viral RNA-dependent RNA polymerases (3D pol s) are predicted to catalyze the polyadenylation of picornaviral RNA in a template-dependent manner during viral RNA replication (8); however, the mechanisms by which RNA genomes are polyadenylated during viral RNA replication remain obscure. In particular, there is little understanding of the mechanisms regulating the length of poly(A) tails synthesized during RNA replication. The RNA-dependent RNA polymerases of negative-strand RNA viruses reiteratively transcribe a small poly(U) sequence within intergenic regions of the viral RNA genome to produce long poly(A) tails on viral mRNAs (9). In a similar manner, the poliovirus RNA-dependent RNA polymerase can reiteratively transcribe poly(A) and poly(U) sequences during viral RNA replication, producing poly(A) and poly(U) sequences longer than those in the respective template RNAs (8).Viral RNA-dependent RNA polymerases are well studied at the structural level (10-12), yet there have been no reports describing the structural features of viral polymerases involved in the polyadenylation of viral RNA. The RNA-dependent RNA polymerase of picornaviruses a...

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Section: Discussionmentioning

confidence: 99%

Structural Features of a Picornavirus Polymerase Involved in the Polyadenylation of Viral RNA

Kempf

Kelly²,

Springer

et al. 2013

J Virol

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“…Impressive progress toward achieving this goal was made by the determination of the atomic resolution structure of TERT from Tribolium castaneum (19). At the colloquium and in an accompanying perspective, Juli Feigon summarized advances on the structure of human telomerase RNA (TER) and the RNP (20,21).…”

Section: Telomerases and Retrotransposons Function Within The Contextmentioning

confidence: 99%

Telomerase and retrotransposons: Reverse transcriptases that shaped genomes

Belfort

Curcio

Lue

2011

Proc. Natl. Acad. Sci. U.S.A.

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“…The ribonucleic acid component of telomerase, TR, containing the template, associates with the complementary 3 single-stranded overhang of telomeric DNA. This is followed by processive repeat addition synthesis of DNA in which a telomeric repeat DNA sequence is synthesised, followed by dissociation of the singlestranded product from the template, and translocation of the template (RNA) to enable synthesis of the next telomeric repeat [20][21][22][23][24]. The mechanism by which translocation occurs has not been fully elucidated, and the role(s), if any, of secondary DNA structures (e.g.…”

Section: Introductionmentioning

confidence: 99%

ESI-MS Investigation of an Equilibrium between a Bimolecular Quadruplex DNA and a Duplex DNA/RNA Hybrid

Birrento

Bryan

Samosorn

et al. 2015

J. Am. Soc. Mass Spectrom.

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, J. L. (2015). ESI-MS investigation of an equilibrium between a bimolecular quadruplex DNA and a duplex DNA/RNA hybrid. Journal of the American Society for Mass Spectrometry, 26 (7), 1165-1173.ESI-MS investigation of an equilibrium between a bimolecular quadruplex DNA and a duplex DNA/RNA hybrid AbstractElectrospray ionization mass spectrometry (ESI-MS) conditions were optimized for simultaneous observation of a bimolecular qDNA and a Watson-Crick base-paired duplex DNA/RNA hybrid. The DNA sequence used was telomeric DNA, and the RNA contained the template for telomerase-mediated telomeric DNA synthesis. Addition of RNA to the quadruplex DNA (qDNA) resulted in formation of the duplex DNA/ RNA hybrid. Melting profiles obtained using circular dichroism spectroscopy confirmed that the DNA/RNA hybrid exhibited greater thermal stability than the bimolecular qDNA in solution. Binding of a 13-substituted berberine (1) derivative to the bimolecular qDNA stabilized its structure as evidenced by an increase in its stability in the mass spectrometer, and an increase in its circular dichroism (CD) melting temperature of 10°C. The DNA/RNA hybrid did not bind the ligand extensively and its thermal stability was unchanged in the presence of (1). The qDNA-ligand complex resisted unfolding in the presence of excess RNA, limiting the formation of the DNA/RNA hybrid. Previously, it has been proposed that DNA secondary structures, such as qDNA, may be involved in the telomerase mechanism. DNA/RNA hybrid structures occur at the active site of telomerase. The results presented in the current work show that if telomeric DNA was folded into a qDNA structure, it is possible for a DNA/RNA hybrid to form as is required during template alignment. The discrimination of ligand (1) AbstractElectrospray ionization mass spectrometry (ESI-MS) conditions were optimized for simultaneous observation of a bimolecular qDNA and a Watson-Crick base-paired duplex DNA/RNA hybrid. The DNA sequence used was telomeric DNA, and the RNA contained the template for telomerase-mediated telomeric DNA synthesis. Addition of RNA to the qDNA resulted in formation of the duplex DNA/RNA hybrid. Melting profiles obtained using circular dichroism spectroscopy confirmed that the DNA/RNA hybrid exhibited greater thermal stability than the bimolecular qDNA in solution. Binding of a 13-substituted berberine (1) derivative to the bimolecular qDNA stabilized its structure as evidenced by an increase in its stability in the mass spectrometer, and an increase in its (CD) melting temperature of 10 C. The DNA/RNA hybrid did not bind the ligand extensively and its thermal stability was unchanged in the presence of (1). The qDNA-ligand complex resisted unfolding in the presence of excess RNA, limiting the formation of the DNA/RNA hybrid.Previously, it has been proposed that DNA secondary structures, such as qDNA, may be involved in the telomerase mechanism. DNA/RNA hybrid structures occur at the active site of telomerase. The results presented in the current work show that if tel...

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Structural basis for telomerase catalytic subunit TERT binding to RNA template and telomeric DNA

Cited by 185 publications

References 58 publications

Structural Features of a Picornavirus Polymerase Involved in the Polyadenylation of Viral RNA

Structural Features of a Picornavirus Polymerase Involved in the Polyadenylation of Viral RNA

Telomerase and retrotransposons: Reverse transcriptases that shaped genomes

ESI-MS Investigation of an Equilibrium between a Bimolecular Quadruplex DNA and a Duplex DNA/RNA Hybrid

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