Polyubiquitination of proliferating cell nuclear antigen by HLTF and SHPRH prevents genomic instability from stalled replication forks.
Chronic stalling of DNA replication forks caused by DNA damage can lead to genomic instability. Cells have evolved lesion bypass pathways such as postreplication repair (PRR) to resolve these arrested forks. In yeast, one branch of PRR involves proliferating cell nuclear antigen (PCNA) polyubiquitination mediated by the Rad5-Ubc13-Mms2 complex that allows bypass of DNA lesion by a template-switching mechanism. Previously, we identified human SHPRH as a functional homologue of yeast Rad5 and revealed the existence of RAD5-like pathway in human cells. Here we report the identification of HLTF as a second RAD5 homologue in human cells. HLTF, like SHPRH, shares a unique domain architecture with Rad5 and promotes lysine 63-linked polyubiquitination of PCNA. Similar to yeast Rad5, HLTF is able to interact with UBC13 and PCNA, as well as SHPRH; and the reduction of either SHPRH or HLTF expression enhances spontaneous mutagenesis. Moreover, Hltf-deficient mouse embryonic fibroblasts show elevated chromosome breaks and fusions after methyl methane sulfonate treatment. Our results suggest that HLTF and SHPRH are functional homologues of yeast Rad5 that cooperatively mediate PCNA polyubiquitination and maintain genomic stability.
Motegi
Akira
A
Genome Instability Section, Genetics and Molecular Biology Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Liaw
Hung-Jiun
HJ
Lee
Kyoo-Young
KY
Roest
Henk P
HP
Maas
Alex
A
Wu
Xiaoli
X
Moinova
Helen
H
Markowitz
Sanford D
SD
Ding
Hao
H
Hoeijmakers
Jan H J
JH
Myung
Kyungjae
K
D000251:Adenosine Triphosphatases; D004249:DNA Damage; D004265:DNA Helicases / Q000378:metabolism*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000378:metabolism*; D042822:Genomic Instability; D006801:Humans; D025821:Polyubiquitin / Q000378:metabolism*; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D040681:Structural Homology, Protein; D014157:Transcription Factors / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination
D004268:DNA-Binding Proteins; C100023:HLTF protein, human; D018809:Proliferating Cell Nuclear Antigen; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D025821:Polyubiquitin; C471607:UBE2N protein, human; D044763:Ubiquitin-Conjugating Enzymes; C516226:SHPRH protein, human; D044767:Ubiquitin-Protein Ligases; D000251:Adenosine Triphosphatases; C076225:RAD5 protein, S cerevisiae; D004265:DNA Helicases