Chemistry and Physics Faculty Articles

Document Type

Article

Publication Date

7-24-2021

Publication Title

Nucleic Acids Research

Keywords

Genome integrity, repair and replication

ISSN

1362-4962

First Page

gkab634

Abstract

PrimPol is the second primase in human cells, the first with the ability to start DNA chains with dNTPs. PrimPol contributes to DNA damage tolerance by restarting DNA synthesis beyond stalling lesions, acting as a TLS primase. Multiple alignment of eukaryotic PrimPols allowed us to identify a highly conserved motif, WxxY near the invariant motif A, which contains two active site metal ligands in all members of the archeo-eukaryotic primase (AEP) superfamily. In vivo and in vitro analysis of single variants of the WFYY motif of human PrimPol demonstrated that the invariant Trp87 and Tyr90 residues are essential for both primase and polymerase activities, mainly due to their crucial role in binding incoming nucleotides. Accordingly, the human variant F88L, altering the WFYY motif, displayed reduced binding of incoming nucleotides, affecting its primase/polymerase activities especially during TLS reactions on UV-damaged DNA. Conversely, the Y89D mutation initially associated with High Myopia did not affect the ability to rescue stalled replication forks in human cells. Collectively, our data suggest that the WFYY motif has a fundamental role in stabilizing the incoming 3′-nucleotide, an essential requisite for both its primase and TLS abilities during replication fork restart.

Comments

This article is dedicated to the memory of Samuel H. Wilson who passed away on 23 April 2021.

© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.

Additional Comments

BFU2015-65880-P (MINECO) and PGC2018-093576-B.C21 (MCI/AEI/FEDER, UE) to L.B., BFU2016-80402-R (MINECO/FEDER, UE) and PID2019-106707RB-100 (AEI/10.13039/501100011033) to J.M., Kempe JCK 1831 to G.S, Knut och Alice Wallenbergs Foundation KAW 2019.0307, VR-2018-02781, to S.W., and by institutional grants from Fundación Ramón Areces and Banco de Santander to the Centro de Biología Molecular Severo Ochoa; P.A.C. and M.D. were recipients of FPI-predoctoral fellowships from Spanish Ministry of Economy and Competitiveness. Funding for open access charge: Consejo Superior de Investigaciones Científicas.

Creative Commons License

Creative Commons Attribution 4.0 International License
This work is licensed under a Creative Commons Attribution 4.0 International License.

gkab634_supplemental_file.docx (1348 kB)
Supplementary data

ORCID ID

0000-0002-7836-7262

DOI

10.1093/nar/gkab634

Peer Reviewed

Included in

Chemistry Commons

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