EXPLORING THE ROLE OF ABI3 AND THE CONSEQUENCES OF THE ALZHEIMER’S DISEASE RISK VARIANT S209F ON MICROGLIAL FUNCTION

Genome-wide association studies have identified Alzheimer’s disease risk variants in several microglial genes, including ABI3 and members of the TREM2 signalling pathway. ABI3 is an adaptor protein, and part of the WAVE Regulatory Complex (WRC), a key regulator of the actin cytoskeleton. This work aims to understand the role of ABI3 in microglia and to characterise the functional consequences of the AD-associated risk variant, S209F. Isogenic human induced pluripotent stem cell (hiPSC) lines carrying ABI3 knock-out (KO) or S209F knock-in (KI) mutations were generated using CRISPR-Cas9 genome editing and differentiated alongside the parental line into tissue-resident macrophages. The effects of these mutations were assessed by Western blotting, qRT-PCR and high-content imaging focused on phagocytosis, migration and morphology. The S209F variant led to a reduction in ABI3 phosphorylation, a process that was found to be regulated by TREM2 signalling, as demonstrated by genetic and pharmacological manipulation of the pathway. Changes in ABI3 levels were associated with altered levels of the WRC components CYFIP1 and NCKAP1L, suggestive of dysregulated WRC assembly. Despite these changes, cell morphology and total actin levels remained unchanged in both ABI3 KO and S209F KI lines under baseline conditions. However, following stimulation, ABI3 KO macrophages exhibited impaired phagocytosis of apoptotic SH-SY5Y cells, while S209F KI macrophages displayed enhanced migration toward C5a. Collectively, these findings provide insights into the role of ABI3 and the S209F risk variant in modulating microglial functions. Understanding how this variant alters microglial behaviour may help inform future therapeutic strategies for Alzheimer’s disease.