title: HLTF and SHPRH are not essential for PCNA polyubiquitination, survival and somatic hypermutation: existence of an alternative E3 ligase.
abstract: DNA damage tolerance is regulated at least in part at the level of proliferating cell nuclear antigen (PCNA) ubiquitination. Monoubiquitination (PCNA-Ub) at lysine residue 164 (K164) stimulates error-prone translesion synthesis (TLS), Rad5-dependent polyubiquitination (PCNA-Ub(n)) stimulates error-free template switching (TS). To generate high affinity antibodies by somatic hypermutation (SHM), B cells profit from error-prone TLS polymerases. Consistent with the role of PCNA-Ub in stimulating TLS, hypermutated B cells of PCNA(K164R) mutant mice display a defect in generating selective point mutations. Two Rad5 orthologs, HLTF and SHPRH have been identified as alternative E3 ligases generating PCNA-Ub(n) in mammals. As PCNA-Ub and PCNA-Ub(n) both make use of K164, error-free PCNA-Ub(n)-dependent TS may suppress error-prone PCNA-Ub-dependent TLS. To determine a regulatory role of Shprh and Hltf in SHM, we generated Shprh/Hltf double mutant mice. Interestingly, while the formation of PCNA-Ub and PCNA-Ub(n) is prohibited in PCNA(K164R) MEFs, the formation of PCNA-Ub(n) is not abolished in Shprh/Hltf mutant MEFs. In line with these observations Shprh/Hltf double mutant B cells were not hypersensitive to DNA damage. Furthermore, SHM was normal in Shprh/Hltf mutant B cells. These data suggest the existence of an alternative E3 ligase in the generation of PCNA-Ub(n).
authors.affiliation: Division of Immunology, The Netherlands Cancer Institute, 1066 CX Amsterdam, The Netherlands.
authors.forename: Peter H L
authors.initials: PH
authors.lastname: Krijger
authors.forename: Kyoo-Young
authors.initials: KY
authors.lastname: Lee
authors.forename: Niek
authors.initials: N
authors.lastname: Wit
authors.forename: Paul C M
authors.initials: PC
authors.lastname: van den Berk
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Wu
authors.forename: Henk P
authors.initials: HP
authors.lastname: Roest
authors.forename: Alex
authors.initials: A
authors.lastname: Maas
authors.forename: Hao
authors.initials: H
authors.lastname: Ding
authors.forename: Jan H J
authors.initials: JH
authors.lastname: Hoeijmakers
authors.forename: Kyungjae
authors.initials: K
authors.lastname: Myung
authors.forename: Heinz
authors.initials: H
authors.lastname: Jacobs
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004265:DNA Helicases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D053595:Embryonic Stem Cells / Q000378:metabolism; D055785:Gene Knockdown Techniques; D023061:Gene Order; D018390:Gene Targeting; D017578:Immunoglobulin Class Switching / Q000187:drug effects; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics / Q000378:metabolism*; D027041:Somatic Hypermutation, Immunoglobulin / Q000235:genetics*; D014157:Transcription Factors / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination / Q000235:genetics / Q000502:physiology* / Q000528:radiation effects; D014466:Ultraviolet Rays / Q000009:adverse effects
chemical_list: D003432:Cross-Linking Reagents; D008070:Lipopolysaccharides; D018809:Proliferating Cell Nuclear Antigen; D014157:Transcription Factors; D044767:Ubiquitin-Protein Ligases; D004265:DNA Helicases