Journal: Neuron

Article Title: Acute MeCP2 loss in adult mice reveals transcriptional and chromatin changes that precede neurological dysfunction and inform pathogenesis.

doi: 10.1016/j.neuron.2024.11.006

Figure Lengend Snippet: Figure 1. Acute depletion of MeCP2 protein in adult knockout mice (A) Schematic of conditional deletion of Mecp2 in 16-week old, adult Mecp2flox/y;UBC-CreERT2 mice using tamoxifen (TMX). Male mice were treated with 33 doses of either TMX (100 mg/kg) or peanut oil vehicle. TMX-treated animals are referred to as adult knockout (aKO) and shown in red, while vehicle-treated animals are referred to as control and shown in black.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit monoclonal anti-MeCP2 (clone D4F3) Cell Signaling Technology #3456; RRID: AB_2143894 Rabbit monoclonal anti-Tri-Methyl-Histone H3 (Lys27) [H3K27me3] (clone C36B11) Cell Signaling Technology #9733; RRID: AB_2616029 Rabbit monoclonal anti-Acetyl-Histone H3 (Lys27) [H3K27ac] (clone D5E4) Cell Signaling Technology #8173; RRID: AB_10949503 Rabbit polyclonal anti-Tri-Methyl-Histone H3 (Lys9) [H3K9me3] Abcam #ab8898; RRID: AB_306848 Rabbit monoclonal anti-Acetyl-Histone H3 (Lys9) [H3K9ac] (clone C5B11) Cell Signaling Technology #9649; RRID: AB_823528 Mouse monoclonal anti-Di and Tri-Methyl-Histone H3 (Lys20) [H4K20me2/3] (clone 6F8-D9) Abcam #ab78517; RRID: AB_1951279 Rabbit polyclonal IgG Millipore #12-370; RRID: AB_145841 Mouse monoclonal anti-Vinculin (clone hVIN-1) MilliporeSigma #V9131; RRID: AB_477629 Mouse monoclonal anti-GAPDH (clone 6C5) Advanced ImmunoChemical Inc. #2-RGM2; RRID: AB_2721282 Chicken polyclonal anti-alpha tubulin Abcam #ab89984; RRID: AB_10672056 Chemicals, peptides, and recombinant proteins Tamoxifen Sigma-Aldrich #T5648 Peanut oil Sigma-Aldrich #P2144 OptiPrep density gradient Sigma-Aldrich #D15556 Spermidine Sigma-Aldrich #S0266 Ultrapure BSA Invitrogen #AM2618 cOmplete protease inhibitor Roche #11873580001 Digitonin Calbiochem #11024-24-1 pAG-MNase Epicypher #15-1016 Critical commercial assays Qiagen miRNeasy Mini Qiagen #217004 Qiagen RNase free DNase Qiagen #79254 NEB Next II DNA Ultra Kit New England Biolabs #E7645S Unique Combinatorial Dual index kit New England Biolabs #E6440S, #E6442S, #E6444S Deposited data Mecp2 Adult Knockout RNA-seq (raw and processed) This study GEO: GSE246119 Mecp2 Adult Knockout CUT&RUN (raw and processed) This study GEO: GSE246664 Mecp2-G118E RNA-seq (raw and processed) This study GEO: GSE277077 Mecp2-null Cortex RNA-seq Zhou et al.36 GEO: GSE179229 Mecp2-null Cortex RNA-seq Clemens et al.10 GEO: GSE123372 Mecp2-null Cortex RNA-seq Boxer et al.9 GEO: GSE128178 Mecp2-R306C Cortex RNA-seq Boxer et al.9 GEO: GSE128178 Mecp2-null Hypothalamus RNA-seq Chen et al.20 GEO: GSE66870 Mecp2-null Cerebellum RNA-seq Raman et al.37 GEO: GSE105045 Reduced representation bisulfite sequencing of hippocampus Reizel et al.39 GEO: GSE85251 Hippocampal RNA-seq used in deconvolution Habib et al.64 GEO: GSE143758 Hippocampal RNA-seq used in deconvolution Methi et al.65 GEO: GSE234278 (Continued on next page) Neuron 113, 1–16.e1–e8, February 5, 2025 e1

Techniques: Knock-Out, Control

Journal: Neuron

Article Title: Acute MeCP2 loss in adult mice reveals transcriptional and chromatin changes that precede neurological dysfunction and inform pathogenesis.

doi: 10.1016/j.neuron.2024.11.006

Figure Lengend Snippet: Figure 2. Progressive global transcriptional dysregulation in the hippocampus after acute depletion of MeCP2 (A) Schematic of conditionally deleting Mecp2 in 16-week-old, adult Mecp2flox/y;UBC-CreERT2 mice using tamoxifen (TMX). Male mice were treated with 33 doses of either TMX (100 mg/kg) or peanut oil vehicle. Total RNA from the hippocam- pus was collected at the time points indicated by a check mark after TMX treatment in adult knockout (aKO), control, and treatment control mice. Bulk mRNA sequencing was performed, and differen- tially expressed genes (DEGs) between (1) treat- ment controls and control (Table S2) and (2) aKO and controls were tabulated (Table S4). DEGs de- tected in treatment controls were removed as non- specific gene expression changes. (B) Volcano plots plotting log2 fold change (aKO/ control) versus padj highlighting DEGs at each time point. The size of the data points (see right upper center of week 8 graph) reflects the number of total time points a given gene was classified as a DEG (padj < 0.05 and | log2 fold change | > 0.15).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit monoclonal anti-MeCP2 (clone D4F3) Cell Signaling Technology #3456; RRID: AB_2143894 Rabbit monoclonal anti-Tri-Methyl-Histone H3 (Lys27) [H3K27me3] (clone C36B11) Cell Signaling Technology #9733; RRID: AB_2616029 Rabbit monoclonal anti-Acetyl-Histone H3 (Lys27) [H3K27ac] (clone D5E4) Cell Signaling Technology #8173; RRID: AB_10949503 Rabbit polyclonal anti-Tri-Methyl-Histone H3 (Lys9) [H3K9me3] Abcam #ab8898; RRID: AB_306848 Rabbit monoclonal anti-Acetyl-Histone H3 (Lys9) [H3K9ac] (clone C5B11) Cell Signaling Technology #9649; RRID: AB_823528 Mouse monoclonal anti-Di and Tri-Methyl-Histone H3 (Lys20) [H4K20me2/3] (clone 6F8-D9) Abcam #ab78517; RRID: AB_1951279 Rabbit polyclonal IgG Millipore #12-370; RRID: AB_145841 Mouse monoclonal anti-Vinculin (clone hVIN-1) MilliporeSigma #V9131; RRID: AB_477629 Mouse monoclonal anti-GAPDH (clone 6C5) Advanced ImmunoChemical Inc. #2-RGM2; RRID: AB_2721282 Chicken polyclonal anti-alpha tubulin Abcam #ab89984; RRID: AB_10672056 Chemicals, peptides, and recombinant proteins Tamoxifen Sigma-Aldrich #T5648 Peanut oil Sigma-Aldrich #P2144 OptiPrep density gradient Sigma-Aldrich #D15556 Spermidine Sigma-Aldrich #S0266 Ultrapure BSA Invitrogen #AM2618 cOmplete protease inhibitor Roche #11873580001 Digitonin Calbiochem #11024-24-1 pAG-MNase Epicypher #15-1016 Critical commercial assays Qiagen miRNeasy Mini Qiagen #217004 Qiagen RNase free DNase Qiagen #79254 NEB Next II DNA Ultra Kit New England Biolabs #E7645S Unique Combinatorial Dual index kit New England Biolabs #E6440S, #E6442S, #E6444S Deposited data Mecp2 Adult Knockout RNA-seq (raw and processed) This study GEO: GSE246119 Mecp2 Adult Knockout CUT&RUN (raw and processed) This study GEO: GSE246664 Mecp2-G118E RNA-seq (raw and processed) This study GEO: GSE277077 Mecp2-null Cortex RNA-seq Zhou et al.36 GEO: GSE179229 Mecp2-null Cortex RNA-seq Clemens et al.10 GEO: GSE123372 Mecp2-null Cortex RNA-seq Boxer et al.9 GEO: GSE128178 Mecp2-R306C Cortex RNA-seq Boxer et al.9 GEO: GSE128178 Mecp2-null Hypothalamus RNA-seq Chen et al.20 GEO: GSE66870 Mecp2-null Cerebellum RNA-seq Raman et al.37 GEO: GSE105045 Reduced representation bisulfite sequencing of hippocampus Reizel et al.39 GEO: GSE85251 Hippocampal RNA-seq used in deconvolution Habib et al.64 GEO: GSE143758 Hippocampal RNA-seq used in deconvolution Methi et al.65 GEO: GSE234278 (Continued on next page) Neuron 113, 1–16.e1–e8, February 5, 2025 e1

Techniques: Knock-Out, Control, Sequencing, Gene Expression