Disease-associated mutations in topoisomerase IIβ result in defective NK cells

Lori Broderick,Gwendolyn M Clay,Robert H Blum,Yang Liu,Rachael Mcvicar,Fabio Papes,Laela M Booshehri,Ian G Cowell,Caroline A Austin,Christopher D Putnam,Dan S Kaufman

doi:10.1016/j.jaci.2021.12.792

Abstract

Hoffman syndrome is a syndromic, inborn error of immunity due to autosomal-dominant mutations in TOP2B, an essential gene required to alleviate topological stress during DNA replication and gene transcription. Although mutations identified in patients lead to a block in B-cell development and the absence of circulating B cells, an effect on natural killer (NK) cells was not previously examined. We sought to determine whether disease-associated mutations in TOP2B impact NK-cell development and function. Using a knockin murine model and patient-derived induced pluripotent stem cells (iPSCs), we investigated NK-cell development in mouse bone marrow and spleen, and performed immunophenotyping by flow cytometry, gene expression, and functional assessment of cytotoxic activity in murine NK cells, and human IPSC-derived NK cells. Mature NK cells were reduced in the periphery of TOP2B knockin mice consistent with patient reports, with reduced cytotoxicity toward target cell lines. IPSCs were successfully derived from patients with Hoffman syndrome, but under optimal conditions showed reduced cytotoxicity compared with iPSC-derived NK cells from healthy controls. Hoffman syndrome-associated mutations in TOP2B impact NK-cell development and function in murine and human models.

Highlights

Topoisomerases are essential enzymes required for relaxation of topological stress during DNA replication and gene transcription, via the generation of transient double-strand breaks in DNA
The probands from each family with Hoffman (BILU) syndrome presented with symptoms consistent with humoral deficiency including recurrent infections by polysaccharide-encapsulated bacteria, severe hypogammaglobulinemia, and absent cluster of differentiation (CD)[191] B cells, with normal T-cell responses to mitogens, more than two-third of the patients had low to lower limit of normal numbers of natural killer (NK) cells (Table I).[1,2,3,4]
Analysis of murine primary B cells from published Top2b ChIP-seq data and etoposide-induced DNA breaks by END-seq[9] showed the presence of Top2b in the promoter and enhancer regions of the key B-cell transcription factor Pax5.1 Remarkably, a similar analysis shows that Top2b in mice binds to the Nfil[3] promoter, a key regulator of NK-cell differentiation (Fig 1, A).[10]

Summary

Introduction

Topoisomerases are essential enzymes required for relaxation of topological stress during DNA replication and gene transcription, via the generation of transient double-strand breaks in DNA. We previously reported 3 unrelated families with autosomaldominant inherited syndromic B-cell immunodeficiency subsequently found to have heterozygous-dominant negative mutations in the TOPRIM domain of TOP2B,1 1 of the 2 type II topoisomerases in humans. This was later confirmed in 2 additional families.[2] the probands from each family with Hoffman (BILU) syndrome presented with symptoms consistent with humoral deficiency including recurrent infections by polysaccharide-encapsulated bacteria, severe hypogammaglobulinemia, and absent cluster of differentiation (CD)[191] B cells, with normal T-cell responses to mitogens, more than two-third of the patients had low to lower limit of normal numbers of natural killer (NK) cells (Table I).[1,2,3,4] Because B, T, and NK cells originate from a common lymphocyte precursor, we questioned whether NK-cell development was impaired when TOP2B is mutated. Conclusions: Hoffman syndrome–associated mutations in TOP2B impact NK-cell development and function in murine

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Results