title: The RNF8/RNF168 ubiquitin ligase cascade facilitates class switch recombination.
abstract: An effective immune response requires B cells to produce several classes of antibodies through the process of class switch recombination (CSR). Activation-induced cytidine deaminase initiates CSR by deaminating deoxycytidines at switch regions within the Ig locus. This activity leads to double-stranded DNA break formation at the donor and recipient switch regions that are subsequently synapsed and ligated in a 53BP1-dependent process that remains poorly understood. The DNA damage response E3 ubiquitin ligases RNF8 and RNF168 were recently shown to facilitate recruitment of 53BP1 to sites of DNA damage. Here we show that the ubiquitination pathway mediated by RNF8 and RNF168 plays an integral part in CSR. Using the CH12F3-2 mouse B cell line that undergoes CSR to IgA at high rates, we demonstrate that knockdown of RNF8, RNF168, and 53BP1 leads to a significant decrease in CSR. We also show that 53BP1-deficient CH12F3-2 cells are protected from apoptosis mediated by the MDM2 inhibitor Nutlin-3. In contrast, deficiency in either E3 ubiquitin ligase does not protect cells from Nutlin-3-mediated apoptosis, indicating that RNF8 and RNF168 do not regulate all functions of 53BP1.
authors.affiliation: Department of Immunology, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Shaliny
authors.initials: S
authors.lastname: Ramachandran
authors.forename: Richard
authors.initials: R
authors.lastname: Chahwan
authors.forename: Rajeev M
authors.initials: RM
authors.lastname: Nepal
authors.forename: Darina
authors.initials: D
authors.lastname: Frieder
authors.forename: Stephanie
authors.initials: S
authors.lastname: Panier
authors.forename: Sergio
authors.initials: S
authors.lastname: Roa
authors.forename: Ahmad
authors.initials: A
authors.lastname: Zaheen
authors.forename: Daniel
authors.initials: D
authors.lastname: Durocher
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Scharff
authors.forename: Alberto
authors.initials: A
authors.lastname: Martin
mesh_terms: D000818:Animals; D002460:Cell Line; D003564:Cytidine Deaminase / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics*; D006801:Humans; D007070:Immunoglobulin A / Q000235:genetics; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D011995:Recombination, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000071857:Tumor Suppressor p53-Binding Protein 1; D044767:Ubiquitin-Protein Ligases / Q000235:genetics* / Q000378:metabolism; D054875:Ubiquitination / Q000235:genetics
chemical_list: D004268:DNA-Binding Proteins; D007070:Immunoglobulin A; D047908:Intracellular Signaling Peptides and Proteins; C486521:RNF8 protein, human; C498301:TP53BP1 protein, human; D000071857:Tumor Suppressor p53-Binding Protein 1; C535210:RNF168 protein, human; D044767:Ubiquitin-Protein Ligases; D003564:Cytidine Deaminase