Physical and Functional Interactions of Human DNA Polymerase {eta} with PCNA

doi:10.1128/MCB.21.21.7199-7206.2001

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Molecular and Cellular Biology, November 2001, p. 7199-7206, Vol. 21, No. 21
0270-7306/01/$04.00+0 DOI: 10.1128/MCB.21.21.7199-7206.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Physical and Functional Interactions of Human DNA Polymerase $eta$ with PCNA

Lajos Haracska,¹ Robert E. Johnson,¹ Ildiko Unk,¹ Barbara Phillips,² Jerard Hurwitz,² Louise Prakash,¹ and Satya Prakash¹^,^*

Sealy Center for Molecular Science, University of Texas Medical Branch, Galveston, Texas 77555-1061,¹ and Department of Molecular Biology and Virology, Memorial Sloan-Kettering Cancer Center, New York, New York 10021-6007²

Received 5 July 2001/Returned for modification 20 July 2001/Accepted 27 July 2001

Human DNA polymerase $eta$ (hPol $eta$ ) functions in the error-free replication of UV-damaged DNA, and mutations in hPol $eta$ cause cancer-pronesyndrome, the variant form of xeroderma pigmentosum. However,in spite of its key role in promoting replication through a varietyof distorting DNA lesions, the manner by which hPol $eta$ is targetedto the replication machinery stalled at a lesion site remainsunknown. Here, we provide evidence for the physical interactionof hPol $eta$ with proliferating cell nuclear antigen (PCNA) and showthat mutations in the PCNA binding motif of hPol $eta$ inactivate thisinteraction. PCNA, together with replication factor C and replicationprotein A, stimulates the DNA synthetic activity of hPol $eta$ , andsteady-state kinetic studies indicate that this stimulation accruesfrom an increase in the efficiency of nucleotide insertion resultingfrom a reduction in the apparent K_m for the incomingnucleotide.

^* Corresponding author. Mailing address: Sealy Center for Molecular Science, University of Texas Medical Branch, 6.104 BlockerMedical Research Building, 11th and Mechanic Streets, Galveston,TX 77555-1061. Phone: (409) 747-8602. Fax: (409) 747-8608. E-mail:sprakash{at}scms.utmb.edu.

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Physical and Functional Interactions of Human DNA Polymerase with PCNA

Physical and Functional Interactions of Human DNA Polymerase $eta$ with PCNA