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J Biol Chem. 2017 Aug 18;292(33):13853-13866. doi: 10.1074/jbc.M117.796425. Epub 2017 Jun 22.

Saccharomyces cerevisiae Red1 protein exhibits nonhomologous DNA end-joining activity and potentiates Hop1-promoted pairing of double-stranded DNA.

Author information

1
From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
2
From the Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India kmbc@biochem.iisc.ernet.in.

Abstract

Elucidation of the function of synaptonemal complex (SC) in Saccharomyces cerevisiae has mainly focused on in vivo analysis of recombination-defective meiotic mutants. Consequently, significant gaps remain in the mechanistic understanding of the activities of various SC proteins and the functional relationships among them. S. cerevisiae Hop1 and Red1 are essential structural components of the SC axial/lateral elements. Previous studies have demonstrated that Hop1 is a structure-selective DNA-binding protein exhibiting high affinity for the Holliday junction and promoting DNA bridging, condensation, and pairing between double-stranded DNA molecules. However, the exact mode of action of Red1 remains unclear, although it is known to interact with Hop1 and to suppress the spore viability defects of hop1 mutant alleles. Here, we report the purification and functional characterization of the full-length Red1 protein. Our results revealed that Red1 forms a stable complex with Hop1 in vitro and provided quantitative insights into their physical interactions. Mechanistically, Red1 preferentially associated with the Holliday junction and 3-way junction rather than with single- or double-stranded DNA with overhangs. Although Hop1 and Red1 exhibited similar binding affinities toward several DNA substrates, the two proteins displayed some significant differences. Notably, Red1, by itself, lacked DNA-pairing ability; however, it potentiated Hop1-promoted intermolecular pairing between double-stranded DNA molecules. Moreover, Red1 exhibited nonhomologous DNA end-joining activity, thus revealing an unexpected role for Red1 in recombination-based DNA repair. Collectively, this study presents the first direct insights into Red1's mode of action and into the mechanism underlying its role in chromosome synapsis and recombination.

KEYWORDS:

DNA recombination; DNA-protein interaction; chromosomes; molecular genetics; nucleic acid structure

PMID:
28642366
PMCID:
PMC5566537
DOI:
10.1074/jbc.M117.796425
[Indexed for MEDLINE]
Free PMC Article

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