Rebecca M. Hauser, Henry L. Limbo, J. Nicholas Brazell, Belle A. Moyers, Shelby N. Lauzon et al.
Tau neurofibrillary tangles are a hallmark of several neurodegenerative diseases called tauopathies, including frontotemporal dementia and Alzheimer’s Disease. Ongoing clinical trials for tauopathies seek to reduce Tau in the brain through immunotherapy, antisense oligonucleotides, and siRNA. MAPT codes for Tau, therefore understanding how the MAPT gene is regulated and the effect of genetic variation at its regulatory elements is likely to have high relevance for tauopathies. We screened a ~3 Mb region including the MAPT locus using 2 different massively parallel reporter assay (MPRA) strategies in KOLF2.1J h-NGN2 neurons and HEK293FT cells, identifying previously unannotated cis-regulatory elements (CREs). Using CRISPR interference (CRISPRi) in mixed neuron cultures, we identified a new CRE for MAPT, as well as 2 CREs for another nearby gene of interest, KANSL1. Known genetic variation from the Alzheimer’s Disease sequencing project was tested in a separate MPRA at the top CREs near the MAPT gene, identifying variants with altered regulatory effects including those at previously identified CREs for MAPT. Using a saturation mutagenesis screen of a 2,000 bp region encompassing the MAPT promoter, we assessed regulatory effects of each possible single nucleotide variant in this region. We identified several neuron-specific regulatory variant effects at this region, including a high confidence binding site for the transcription factors EGR2, ZBTB14, and TCLF5 at a region of high MPRA activity and genetic conservation.
Rebecca M. Hauser, Henry L. Limbo, J. Nicholas Brazell, Belle A. Moyers, Shelby N. Lauzon, Erin A. Barinaga, S. Quinn Johnston, Brianne B. Rogers, Jared W. Taylor, J. Nicholas Cochran. Promoter mutagenesis and a massively parallel reporter screen of the MAPT locus identifies cis-regulatory elements and genetic variation effects. 2026-03-09. https://doi.org/10.64898/2026.03.06.710116
A synthesized oligo lentiMPRA tiled candidate regions that had low BAC-MPRA coverage and additional regions of interest, measuring activity in KOLF2.1J h-NGN2 excitatory neurons. The processed table reports MPRAnalyze activity statistics for 270 bp oligos and scrambled negative controls.
The MAPT promoter deep-mutational-scanning lentiMPRA tested all designed single-nucleotide substitutions and centered 5-bp deletions in HEK293FT cells. The processed table reports bcalm activity effects relative to matched reference sequences.
The ADSP SNV subset of the combined variation lentiMPRA library was assayed in HEK293FT cells. The processed table reports published bcalm alternate-versus-reference activity effects for the tested single-nucleotide variants.
The ADSP InDel subset of the combined variation lentiMPRA library was assayed in HEK293FT cells. The processed table reports published bcalm alternate-versus-reference activity effects for the tested insertions and deletions.
A lentiviral MPRA used randomly sheared BAC fragments spanning approximately 3 Mb around MAPT to measure cis-regulatory activity in KOLF2.1J h-NGN2 excitatory neurons. The processed table reports activity statistics for 100 bp genomic bins.
The BAC lentiMPRA library spanning approximately 3 Mb around MAPT was assayed in HEK293FT cells. The processed table reports MPRAnalyze activity statistics for 100 bp genomic bins.
A variant-focused lentiMPRA tested Alzheimer’s Disease Sequencing Project insertions and deletions overlapping MAPT-proximal candidate regulatory regions in KOLF2.1J h-NGN2 excitatory neurons. The processed table reports alternate-versus-reference activity effects from the published bcalm analysis.
A variant-focused lentiMPRA tested Alzheimer’s Disease Sequencing Project single-nucleotide variants overlapping MAPT-proximal candidate regulatory regions in KOLF2.1J h-NGN2 excitatory neurons. The processed table reports alternate-versus-reference activity effects from the published bcalm analysis.
The synthesized oligo lentiMPRA library tiled MAPT-region candidate elements and additional regions of interest in HEK293FT cells. The processed table reports MPRAnalyze activity statistics for 270 bp oligos and scrambled negative controls.
A deep mutational scanning lentiMPRA tested every possible single-nucleotide substitution and a centered 5-bp deletion across a 2,000 bp MAPT promoter region in KOLF2.1J h-NGN2 excitatory neurons. The processed table reports bcalm activity effects relative to matched reference sequences.
Original supplemental and deposited inputs retained for this study. Download files individually or together as a ZIP; nested folders are preserved. Source reuse terms apply, and sequencing reads may be omitted.
Download all 14 files (ZIP)GSE324616_dna_annot_HEKs.tsv.gzGSE324616_dna_annot_Neurons.tsv.gzGSE324616_family.soft.gzGSE324616_rna_annot_HEKs.tsv.gzGSE324616_rna_annot_Neurons.tsv.gzGSE325256_family.soft.gzGSE325585_dna_annot_Heks.tsv.gzGSE325585_dna_annot_Neurons.tsv.gzGSE325585_family.soft.gzGSE325585_rna_annot_Heks.tsv.gzGSE325585_rna_annot_Neurons.tsv.gzmedia-1.xlsxNIHPP2026.03.06.710116v1-supplement-2.pdfsource_notes.txt