The RNF8/RNF168 ubiquitin ligase cascade facilitates class switch recombination.
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.
Ramachandran
Shaliny
S
Department of Immunology, University of Toronto, Toronto, Ontario, Canada.
Chahwan
Richard
R
Nepal
Rajeev M
RM
Frieder
Darina
D
Panier
Stephanie
S
Roa
Sergio
S
Zaheen
Ahmad
A
Durocher
Daniel
D
Scharff
Matthew D
MD
Martin
Alberto
A
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
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