Expression of the microglial INPP5D isoforms as a function of Alzheimer's disease status and genetics.

Abstract

Genome wide association studies have identified single nucleotide polymorphisms (SNPs) within INPP5D (Phosphatidylinositol-3,4,5-trisphosphate 5-phosphatase 1), the gene encoding SHIP1, that are strongly associated with Alzheimer's Disease (AD) risk. INPP5D is expressed in the brain, mostly in microglia and can include up to 27 exons. In the brain, INPP5D encodes several isoforms. The full-length 27 exon isoform encodes an amino-terminal SH2 domain followed by the phosphatase domain. Truncated isoforms lacking the SH2 domain begin from internal transcription start sites. Whether these isoforms are relevant to the SNP's actions in AD is unclear. RNA from AD and non-AD anterior cingulate human brain samples was analyzed via quantitative polymerase chain reaction for the copy number of each isoform relative to INPP5D standard curves. The AD-associated SNPs were identified using TaqMan (ABI). Isoform expression results were analyzed as a function of microglial gene expression (ITGAM and AIF1), total INPP5D expression, AD status, and SNP status. We also evaluated INPP5D expression as a function of SNPs by using data from GTEX. Single cell RNAseq was performed on APP/PS1 mice to understand the relationship between INPP5D expression and microglial activation. Lastly, we performed SHIP1 immunocytochemistry on AD and non-AD human brain slices. Expression of each INPP5D isoform was strongly correlated with microglial gene expression and showed an increase with AD neuropathology. None of the isoforms showed a significant association with AD-associated SNPs. In contrast, GTEX analysis suggested a modest but significant association between rs35349669 and INPP5D expression; the minor T allele, which is associated with increased AD risk, was associated with decreased INPP5D expression in spleen and blood. INPP5D was expressed at equivalent levels in microglia, regardless of homeostatic versus disease-associated microglial status. Immunocytochemistry supported that SHIP1 expression was restricted to microglia in the human brain. INPP5D is expressed as multiple isoforms, and expression of these isoforms is increased with AD neuropathology. INPP5D expression does not change in disease-associated microglia. The mechanisms whereby AD genetics influence INPP5D expression is not clear, but the rs35349669 allele that increases AD risk may be associated with a decrease in INPP5D expression.