Aberrant APOBEC3B Expression in Breast Cancer Is Linked to Proliferation and Cell Cycle Phase

Abstract

APOBEC3B (A3B) is aberrantly overexpressed in a subset of breast cancers, where it associates with advanced disease, poor prognosis, and treatment resistance, yet the causes of A3B dysregulation in breast cancer remain unclear. Here, A3B mRNA and protein expression levels were quantified in different cell lines and breast tumors and related to cell cycle markers using RT-qPCR and multiplex immunofluorescence imaging. The inducibility of A3B expression during the cell cycle was additionally addressed after cell cycle synchronization with multiple methods. First, we found that A3B protein levels within cell lines and tumors are heterogeneous and associate strongly with the proliferation marker Cyclin B1 characteristic of the G2/M phase of the cell cycle. Second, in multiple breast cancer cell lines with high A3B, expression levels were observed to oscillate throughout the cell cycle and again associate with Cyclin B1. Third, induction of A3B expression is potently repressed throughout G0/early G1, likely by RB/E2F pathway effector proteins. Fourth, in cells with low A3B, induction of A3B through the PKC/ncNF-κB pathway occurs predominantly in actively proliferating cells and is largely absent in cells arrested in G0. Altogether, these results support a model in which dysregulated A3B overexpression in breast cancer is the cumulative result of proliferation-associated relief from repression with concomitant pathway activation during the G2/M phase of the cell cycle.

Human APOBEC3B promotes tumor development in vivo including signature mutations and metastases

“…First, A3B expression is constitutively driven from a CAG promoter, which contrasts with the regulation of endogenous A3B in humans with peak protein levels reported in cell lines and tumors at the G2/M phase of the cell cycle. 56 , 57 The constitutive nature of the CAG promoter also makes it tricky to dissociate tumor-cell-autonomous from -non-autonomous roles for A3B in tumor formation. Second, CAG-A3B animals express A3B protein levels similar to those observed at the high end of cells within human tumors ( Figures 3 D–3F).…”

“…This separates expression of this gene from regulatory mechanisms that are normally operative in human cells including transcriptional repression and cell-cycle regulation (although some of the same regulatory mechanisms are often dysregulated in human tumors). 56 , 57 This study does did not experimentally address alternative mechanisms such as roles for A3B in other (non-tumor) cell types, roles for A3B in altering the epigenome (e.g., methyl-C landscape), 60 or roles for A3B in transcriptional regulation 53 or R-loop homeostasis. 37…”

Similar deamination activities but different phenotypic outcomes induced by APOBEC3 enzymes in breast epithelial cells

“…For the parental, non-transduced MCF10A and MCF7 cell lines, endogenous A3A mRNA was not detected, but there was basal endogenous mRNA expression detected for A3B and A3H Hap I after treatment with dox ( Figures 1D,E , MCF10A or MCF7 on x -axis). Endogenous low-level A3B expression has previously been detected in these cell lines ( Roelofs et al, 2023 ). After dox induction in transduced cells (labeled A3A, A3B, or A3H Hap I on the x -axis), the level of A3A mRNA for both MCF10A and MCF7 cells was low, showing only 0.75- and 1.42-fold increases, respectively, although the steady-state protein expression was similar to A3B ( Figures 1B–E ).…”

Whole-genome mapping of APOBEC mutagenesis in metastatic urothelial carcinoma identifies driver hotspot mutations and a novel mutational signature

“…The lack of correlation might be linked to the heterogeneous expression of APOBEC enzymes that oscillate throughout the cell cycle. 12 , 13 Furthermore, we detected that both APOBEC3A and APOBEC3B contributed to APOBEC-associated mutations (fold enrichment above 1.0). Nevertheless, APOBEC3A appeared to be the main contributor, as suggested in primary cancers.…”