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Abstract

From the genome to the proteome, each molecular step is tightly controlled. Advances in RNA-sequencing technologies show that a system of RNA modifiers extensively and dynamically regulates the transcriptome. The posttranscriptional changes alter mRNA stability, localisation, and translational efficiency. However, the effects and dynamics of one of the least studied RNA modifiers, Apolipoprotein B mRNA editing enzyme, catalytic polypeptide 1 (APOBEC1), are unknown. APOBEC1 catalyses cytosine deamination to uracil in a process known as RNA editing. APOBEC1 has a vital role in lipid metabolism, modifies phagocytosis in mouse macrophages, and its activity contributes to the heterogenic nature of macrophages. This work aimed to study how editing modifies macrophage phenotype, transcript-level effects, and dynamics during macrophage activation. To determine whether APOBEC1 editing is temporally regulated after activation, I analysed RNA-sequencing data from polarised mouse bone-marrow-derived macrophages and the macrophage cell line RAW264.7. I show that global editing levels are altered over time with an early increase followed by a drastic decrease. I report a striking distinction in the pattern of transcript level editing changes after proinflammatory stimulation; transcripts involved in phagosome maturation (processes that acidify phagosomes) have decreased editing while transcripts essential for antigen presentation and processing (processes that increase or require higher pH levels) have increased editing. I find that APOBEC1 increases antigen presentation and processing machinery protein levels and decreases factors involved in acidification and antigen digestion. I demonstrate that the lack of editing decreases the translational efficiency of Cytochrome B-245 Beta Chain (CYBB). CYBB is part of an enzyme complex that produces reactive oxygen species in phagosomes consuming protons during the process, which is critical for preventing antigen over-digestion. The loss of APOBEC1 causes dysfunction in the regulation of phagosomal pH resulting in lower-than-normal pH, which in part is responsible for the previous observations of increased phagocytosis in knockout macrophages. Overall, my work highlights a novel role of APOBEC1 editing in regulating transcript translational efficiency and control of the primary functions of macrophages in phagocytosis and antigen presentation.