Synonymous variant rs2405442 in PILRA is associated with Alzheimer's disease and affects RNA expression by destroying a ramp sequence

Abstract

AbstractBackgroundPaired Immunoglobulin‐like Type 2 Receptor Alpha (PILRA) recognizes specific O‐glycosylated proteins on various cell types, including microglia. A common missense mutation in PILRA, rs1859788, that likely causes the confirmed Alzheimer's disease risk locus at 7q21 is in linkage disequilibrium with synonymous variant rs2405442:T>C. This synonymous variant affects a ramp of rare codons at the beginning of PILRA, which may decrease overall transcriptional efficiency. Since reduced inhibitory signaling in microglia has a protective effect on Alzheimer's disease, we propose that destroying a ramp sequence in PILRA will likewise protect against Alzheimer's disease by reducing the amount of PILRA inhibitory receptor.MethodsWe recently developed an algorithm, ExtRamp, to identify ramp sequences within specific gene sequences. Using ExtRamp, we calculated the relative codon adaptiveness of PILRA using all isoforms in GRCh38. We then calculated the relative codon adaptiveness of PILRA with synonymous variant rs2405442:T>C and found that a ramp sequence is present in the reference isoforms but not the mutant isoforms. We predicted that although the mutant has a more common codon, it would have less overall mRNA expression because it lacks a ramp sequence. We performed biological validation using four sets of three qPCR replicates on the longest isoform to determine the transcriptional efficiency of the mutant versus the reference. We normalized the qPCR output by the number of cells.ResultsFigure 1 shows the relative adaptiveness of the longest PILRA reference isoform and the mutant gene averaged over a nine codon window. The mutant gene has a higher codon adaptiveness at the beginning of the gene sequence, which destroys the predicted ramp sequence. Figure 2 shows that our biological validation using qPCR confirms our prediction that the mutant gene has lower mRNA expression (p‐value = 4.45 × 10−9).ConclusionsAlthough higher GC content and more common codons generally increase mRNA levels, the ramp sequence appears to play an integral role in transcriptional efficiency in PILRA. Our results show that synonymous variant rs2405442:T>C directly impacts transcriptional efficiency in PILRA and indicates that synonymous variants should be considered in future association analyses for their causal effects on transcriptional efficiency by modifying ramp sequences.