Identification of DNA cleavage- and recombination-specific hnRNP cofactors for activation-induced cytidine deaminase

Proc Natl Acad Sci U S A. 2015 May 5;112(18):5791-6. doi: 10.1073/pnas.1506167112. Epub 2015 Apr 20.

Abstract

Activation-induced cytidine deaminase (AID) is essential for antibody class switch recombination (CSR) and somatic hypermutation (SHM). AID originally was postulated to function as an RNA-editing enzyme, based on its strong homology with apolipoprotein B mRNA-editing enzyme, catalytic polypeptide 1 (APOBEC1), the enzyme that edits apolipoprotein B-100 mRNA in the presence of the APOBEC cofactor APOBEC1 complementation factor/APOBEC complementation factor (A1CF/ACF). Because A1CF is structurally similar to heterogeneous nuclear ribonucleoproteins (hnRNPs), we investigated the involvement of several well-known hnRNPs in AID function by using siRNA knockdown and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9-mediated disruption. We found that hnRNP K deficiency inhibited DNA cleavage and thereby induced both CSR and SHM, whereas hnRNP L deficiency inhibited only CSR and somewhat enhanced SHM. Interestingly, both hnRNPs exhibited RNA-dependent interactions with AID, and mutant forms of these proteins containing deletions in the RNA-recognition motif failed to rescue CSR. Thus, our study suggests that hnRNP K and hnRNP L may serve as A1CF-like cofactors in AID-mediated CSR and SHM.

Keywords: B cell; IgH; activation-induced cytidine deaminase; class switch recombination; somatic hypermutation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • APOBEC-1 Deaminase
  • Cell Line, Tumor
  • Cell Separation
  • Cytidine Deaminase / chemistry*
  • Cytidine Deaminase / metabolism*
  • DNA / chemistry*
  • Flow Cytometry
  • Genetic Complementation Test
  • HEK293 Cells
  • Heterogeneous-Nuclear Ribonucleoprotein K / chemistry
  • Heterogeneous-Nuclear Ribonucleoprotein L / chemistry
  • Heterogeneous-Nuclear Ribonucleoproteins / chemistry*
  • Humans
  • Immunoglobulin A / chemistry
  • Immunoglobulin Class Switching
  • Immunoglobulin Heavy Chains / chemistry
  • Mutation
  • Protein Binding
  • Protein Structure, Tertiary
  • Proto-Oncogene Proteins c-bcl-2 / genetics
  • RNA, Small Interfering / metabolism
  • RNA-Binding Proteins / metabolism*
  • Recombination, Genetic
  • Somatic Hypermutation, Immunoglobulin

Substances

  • A1CF protein, human
  • BCL2 protein, human
  • Heterogeneous-Nuclear Ribonucleoprotein K
  • Heterogeneous-Nuclear Ribonucleoprotein L
  • Heterogeneous-Nuclear Ribonucleoproteins
  • Immunoglobulin A
  • Immunoglobulin Heavy Chains
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Small Interfering
  • RNA-Binding Proteins
  • DNA
  • AICDA (activation-induced cytidine deaminase)
  • APOBEC-1 Deaminase
  • APOBEC1 protein, human
  • Cytidine Deaminase