Project 2: Impact of H1/H2 haplotypes on cellular disease-associated phenotypes driven by FTD-causing MAPT mutations

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PROJECT SUMMARY (PROJECT 2) Common and rare variants at the 17q21.31/MAPT locus contribute to Frontotemporal Dementia (FTD-tau) and the Frontotemporal Lobar Degeneration (FTLD) spectrum disorder, Progressive Supranuclear Palsy (PSP). Dominantly acting, rare missense and splice-site mutations in the MAPT gene have been reported to cause both FTD-tau and PSP. Interestingly, it appears as though the majority of reported pathogenic MAPT mutations causing familial PSP and FTD-tau occur on the H1 haplotype background, which is consistent with the increased genetic risk associated with this haplotype in sporadic disease, although P301L, N279K and IVS10+16 have been described on both haplotypes. However, it is unknown whether these haplotypes influence the phenotypic expression of MAPT mutations. In addition to rare variation contributions to disease in this region, the strongest genome-wide association signal for common variation in PSP is in the 17q21.31/MAPT locus. Identification of causal variants in this region has been hampered by the broad patterns of linkage disequilibrium created by the 970 Kb chromosomal inversion, which limits local recombination. Work by our group and by others, including our preliminary data using massively parallel reporter assays (MPRA), indicates that this 17q21.31/MAPT region harbors several different risk loci and hundreds of contributory common causal variants. Furthermore, because >96% of patients with PSP have the H1 haplotype, the effects of these SNPs driving this major association signal likely occur in the context of the H1 haplotype, and not H2. These observations lead to two major hypotheses that drive this project: 1) H1/H2 haplotypes not only influence risk for sporadic FTD-tau/PSP but also modulate expression of the pathogenic phenotypes associated with specific MAPT variants. Therefore, we hypothesize that the changes to chromatin structure and gene expression or splicing that occur in H2/H2 neural cells compared to H1/H1 will ameliorate the pathogenic effects of PSP/FTD MAPT mutations; and 2) That the regulatory effects of common PSP-associated variation in this region will be similarly blunted on an H2 haplotype. We will use a combination of single-cell genomic, and proteomic approaches to characterize the impact of PSP/FTD-associated MAPT mutations in human brain tissue and in iPSC-derived assembloids. We will use CRISPR genome editing to introduce PSP/FTD-associated MAPT mutations into cell lines from Project 1 with either H1/H1 or H2/H2 backgrounds, and will use single-cell multi-OMICs (RNAseq and ATACseq) combined with proteomics and ISOseq to comprehensively assess the impact of the mutations on each background. We will further characterize the impact of MAPT mutations on disease-associated phenotypes on each background using cellular and molecular assays, and will then use CRISPRa/i screens to validate the functional consequences of key haplotype-associated regulatory regions containing candidate common causal variants on disease-related Tau phenotypes on gene expression in both H1 and H2 haplotypes, in order to test the hypothesis that causal disease variants have more activity on the H1 background.