Abstract
DNA cytosine deaminase APOBEC3B (A3B) is an endogenous source of mutations in many human cancers, including multiple myeloma. A3B proteins form catalytically inactive high molecular mass (HMM) complexes in nuclei, however, the regulatory mechanisms of A3B deaminase activity in HMM complexes are still unclear. Here, we performed mass spectrometry analysis of A3B-interacting proteins from nuclear extracts of myeloma cell lines and identified 30 putative interacting proteins. These proteins are involved in RNA metabolism, including RNA binding, mRNA splicing, translation, and regulation of gene expression. Except for SAFB, these proteins interact with A3B in an RNA-dependent manner. Most of these interacting proteins are detected in A3B HMM complexes by density gradient sedimentation assays. We focused on two interacting proteins, ILF2 and SAFB. We found that overexpressed ILF2 enhanced the deaminase activity of A3B by 30%, while SAFB did not. Additionally, siRNA-mediated knockdown of ILF2 suppressed A3B deaminase activity by 30% in HEK293T cell lysates. Based on these findings, we conclude that ILF2 can interact with A3B and enhance its deaminase activity in HMM complexes.
Highlights
DNA cytosine deaminase APOBEC3B (A3B) is an endogenous source of mutations in many human cancers, including multiple myeloma
We previously reported decreased deaminase activity and fewer Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) signature mutations upon shRNA-mediated A3B knockdown in myeloma cells, suggesting that, among APOBECs, A3B plays a major role in cytidine deamination-related mutagenesis in myeloma c ells[7]
RNA-mediated formation of high molecular mass (HMM) ribonucleoprotein complexes plays a crucial role in APOBEC deaminase activity
Summary
DNA cytosine deaminase APOBEC3B (A3B) is an endogenous source of mutations in many human cancers, including multiple myeloma. We performed mass spectrometry analysis of A3B-interacting proteins from nuclear extracts of myeloma cell lines and identified 30 putative interacting proteins These proteins are involved in RNA metabolism, including RNA binding, mRNA splicing, translation, and regulation of gene expression. SiRNA-mediated knockdown of ILF2 suppressed A3B deaminase activity by 30% in HEK293T cell lysates. Based on these findings, we conclude that ILF2 can interact with A3B and enhance its deaminase activity in HMM complexes. Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) family proteins have DNA cytidine deaminase activity that restricts retroviruses and retrotransposons by inducing hypermutations and degradation of replication intermediates. The estrogen receptor (ER) recruits A3B at ER binding regions and introduces C-to-U deamination which facilitates ER target gene
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