Dataset DOI: 10.5061/dryad.4xgxd25r0

Description of the data and file structure

This dataset contains the primary data supporting the manuscript “Selective loss of primary cilia and neurotrophic signaling in G51D α-synuclein mice highlights a common pathway in Parkinson’s disease.” The dataset includes raw microscopy images (.czi), immunoblot images (.tif), quantitative analyses in GraphPad Prism format (.pzf), tabulated data files (.csv), and analysis scripts, including FIJI/ImageJ macros, R scripts, and CellProfiler pipelines. This dataset also includes a key resource table (Table S1).

Files and variables

File list:

Fig.1.zip file:

Folder Fig_1A_3M_ChAT_cilia

Microscopy images (.czi) of striatal cholinergic (ChAT) neurons from 8 animals of 3 months of age. Four animals are wild type (WT) mice (Br01, Br02, Br03, Br04), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05, Br06, Br07, Br08). Note that all images of Brains (01 to 08) are split into two files.

Folder Fig_1C_12M_ChAT_cilia

Microscopy images (.czi) of striatal cholinergic (ChAT) neurons from 14 animals of 12 months of age. Seven animals are wild type (WT) mice (Br01_set1, Br02_set1, Br03_set1, Br01_set2, Br02_set2, Br03_set2, Br04_set2), and seven are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br04_set1, Br05_set1, Br06_set1, Br05_set2, Br06_set2, Br07_set2, Br08_set2). Note that images of Br01_set1 are split into 3 files, images of Br04_set1 are split into 2 files, and images of Br05_set1 are split into 2 files.

Folder Fig_1E_3M_PV_cilia

Microscopy images (.czi) of striatal parvalbumin (PV) neurons from 8 animals of 3 months of age. Four animals are wild type (WT) mice (Br01, Br02, Br03, Br04), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05, Br06, Br07, Br08). Note that all images of Brains (01 to 08) are split into two files.

Folder Fig_1G_12M_PV_cilia

Microscopy images (.czi) of striatal parvalbumin (PV) neurons from 14 animals of 12 months of age. Seven animals are wild type (WT) mice (Br01_set1, Br02_set1, Br03_set1, Br01_set2, Br02_set2, Br03_set2, Br04_set2), and seven are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br04_set1, Br05_set1, Br06_set1, Br05_set2, Br06_set2, Br07_set2, Br08_set2). Note that images of Br01_set1 are split into 2 files, images of Br04_set1 are split into 2 files, images of Br05_set1 are split into 2 files, and Br06_set1 are split into 2 files.

Folder Fig_1I_3M_Astrocyte_cilia

Microscopy images (.czi) of striatal astrocytes from 8 animals of 3 months of age. Four animals are wild type (WT) mice (Br01, Br02, Br03, Br04), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05, Br06, Br07, Br08).

Folder Fig_1K_12M_Astrocyte_cilia

Microscopy images (.czi) of striatal astrocytes from 14 animals of 12 months of age. Seven animals are wild type (WT) mice (Br01_set1, Br02_set1, Br03_set1, Br01_set2, Br02_set2, Br03_set2, Br04_set2), and seven are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br04_set1, Br05_set1, Br06_set1, Br05_set2, Br06_set2, Br07_set2, Br08_set2).

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. 1B, 1D, 1F, 1H, 1J, and 1L.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. 1B, 1D, 1F, 1H, 1J, and 1L. The number of cells possessing a cilium was manually quantified, and values excluded as outliers (e.g., partially imaged cells) were labeled as NA.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) were imported into GraphPad Prism to generate Fig. 1B, 1D, 1F, 1H, 1J, and 1L.

Fig.2.zip file:

Folder Fig_2A_2B_12M_set2_striatal_ChAT_Ptch1_cilia

Microscopy images (.czi) of striatal cholinergic (ChAT) neurons from 8 animals of 12 months of age. Four animals are wild type (WT) mice (Br01_set2, Br02_set2, Br03_set2, Br04_set2), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2). One additional image set (no-probe control) was acquired from a WT animal. Ptch1 denotes Patched 1.

Folder Fig_2E_2F_12M_set2_striatal_PV_Ptch1_cilia

Microscopy images (.czi) of striatal parvalbumin (PV) neurons from 8 animals of 12 months of age. Four animals are wild type (WT) mice (Br01_set2, Br02_set2, Br03_set2, Br04_set2), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2). One additional image set (no-probe control) was acquired from a WT animal. Ptch1 denotes Patched 1.

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. 2C, 2D, 2G and 2H.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. 2C, 2D, 2G and 2H. The number of cells possessing a cilium and the number of Patched 1 (Ptch1) RNA dot were manually quantified, and values excluded as outliers (e.g., partially imaged cells) were labeled as NA.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) imported into GraphPad Prism to generate Fig. 2C, 2D, 2G and 2H.

Fig.3.zip file:

Folder Fig_3A_12M_ChAT_Gdnf_cilia

Microscopy images (.czi) of striatal cholinergic (ChAT) neurons from 8 animals of 12 months of age. Four animals are wild type (WT) mice (Br01_set2, Br02_set2, Br03_set2, Br04_set2), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2). One additional image set (no-probe control) was acquired from a WT animal. Gdnf denotes glial derived neurotrophic factor.

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. 3B and 3C.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. 3B and 3C. The number of cells possessing a cilium and the number of glial derived neurotrophic factor (Gdnf) RNA dot were manually quantified, and values excluded as outliers (e.g., partially imaged cells) were labeled as NA.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) imported into GraphPad Prism to generate Fig. 3B and 3C.

Fig.4.zip file:

Folder Fig_4A_12M_PV_Nrtn_cilia

Microscopy images (.czi) of striatal parvalbumin (PV) neurons from 8 animals of 12 months of age. Four animals are wild type (WT) mice (Br01_set2, Br02_set2, Br03_set2, Br04_set2), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2). One additional image set (no-probe control) was acquired from a WT animal. Note that images of Br02_set2 are split into 2 files. Nrtn denotes neurturin.

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. 4B and 4C.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. 4B and 4C. The number of cells possessing a cilium and the number of neurturin (Nrtn) RNA dot were manually quantified, and values excluded as outliers (e.g., partially imaged cells) were labeled as NA.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) imported into GraphPad Prism to generate Fig. 4B and 4C.

Fig.5.zip file:

Folder Fig_5A_3M_PCx_PV_cilia

Microscopy images (.czi) of piriform cortex (PCx) parvalbumin (PV) neurons from 8 animals of 3 months of age. Four animals are wild type (WT) mice (Br01, Br02, Br03, Br04), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05, Br06, Br07, Br08). 

Folder Fig_5C_12M_ PCx_PV_cilia

Microscopy images (.czi) of piriform cortex (PCx) parvalbumin (PV) neurons from 14 animals of 12 months of age. Seven animals are wild type (WT) mice (Br01_set1, Br02_set1, Br03_set1, Br01_set2, Br02_set2, Br03_set2, Br04_set2), and seven are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br04_set1, Br05_set1, Br06_set1, Br05_set2, Br06_set2, Br07_set2, Br08_set2).

Folder Fig_5E_12M_PCx_PV_Nrtn_cilia

Microscopy images (.czi) of piriform cortex (PCx) parvalbumin (PV) neurons from 8 animals of 12 months of age. Four animals are wild type (Br01_set2, Br02_set2, Br03_set2, Br04_set2), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2). Nrtn denotes neurturin. 

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. 5B, 5D, 5F and 5G.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. 5B, 5D, 5F, and 5G. The number of cells possessing a cilium and the number of neurturin (Nrtn) RNA dot were manually quantified, and values excluded as outliers (e.g., partially imaged cells) were labeled as NA.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) imported into GraphPad Prism to generate Fig. 5B, 5D, 5F, and 5G.

Fig.6.zip file:

Folder Fig_6A_12M_WT_G51D_HBC_data

Microscopy images (.czi) of horizontal basal cells (HBSs) in the olfactory epithelium from 8 animals of 12 months of age. Four animals are wild type (WT) mice (wt1, wt2, wt3, wt4), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (g51d1, g51d2, g51d3, g51d4). Note that images of wt3 are split into 2 files.

Folder Fig_6D_12M_OSN_ac-tubulin_regular_staining

Microscopy images (.czi) of olfactory sensory neurons (OSNs) in the olfactory epithelium from 8 animals of 12 months of age. Four animals are wild type (WT) mice (wt1, wt2, wt3, wt4), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (g51d1, g51d2, g51d3, g51d4). Note that images of wt3 are split into 2 files.

Folder Fig_6G_6H_12M_expansion_microscopy_WT_G51D

Expansion microscopy images (.czi) of olfactory sensory neurons (OSNs) in the olfactory epithelium from 4 animals of 12 months of age. Two animals are wild type (WT) mice (wt1, wt2), and two are G51D-alpha-synuclein (G51D) homozygous knock-in mice (g51d1, g51d2). Note that each mouse contributes 25 image files.

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. 6B, 6C, 6E, 6F and 6I.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. 6B, 6C, 6E, 6F, and 6I. The number of cells possessing a cilium was manually quantified. The length used to quantify the number of HBCs was measured in millimeters (mm). The intensity of acetylated tubulin was manually quantified. The area used to normalize the number of HBCs was measured in square micrometers (µm²).
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse epithelium) were imported into GraphPad Prism to generate Fig. 6B, 6C, 6E, 6F, and 6I. For visualization in Fig. 6F, values originally measured in square micrometers (µm²) were converted to square millimeters (mm²).

Fig.7.zip file:

Folder Fig_7A_7C_pSNCA_ChAT_WT_and_G51D

Microscopy images (.czi) of striatal cholinergic (ChAT) neurons from 8 animals of 12 months of age. Four animals are wild type (WT) mice (Br01_set2, Br02_set2, Br03_set2, Br04_set2), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2). Note that images of Br02_set2 are split into 2 files. pSNCA denotes phospho–α-synuclein.

Folder Fig_7E_pSNCA_ChAT_MSN_in_G51D_only

Microscopy images (.czi) of striatal cholinergic (ChAT) and medium spiny neurons (MSNs) from 4 animals of 12 months of age. Four animals are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2).

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. 7B and 7F.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. 7B, 7D, and 7F. The number of cells possessing a cilium was manually quantified. Quantification of phospho–α-synuclein (pSNCA) in cholinergic (ChAT) and medium spiny neurons (MSNs) was performed using FIJI/ImageJ macros, R scripts, and CellProfiler pipelines, which are included in the Fig. 7 pipeline files.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) imported into GraphPad Prism to generate Fig. 7B and 7F.

Fig.7_pipeline.zip file:

Folder scripts_for_macro

FIJI/ImageJ scripts used to generate non-overlapping 3-slice maximum-intensity projections from microscopy z-stacks.

Folder cellprofiler

A CellProfiler pipeline used to quantify phospho–α-synuclein (pSNCA) in cholinergic (ChAT) and medium spiny neurons (MSNs).

Folder Rscripts

R scripts used to analyze the output tables generated by the CellProfiler pipeline.

Fig.S1.zip file:

.tif file for Fig_S1A_ChAT_control_set2_low_mag_(RGB)

.tif file for Fig_S1A_ChAT_G51D_set2_low_mag_(RGB)

Folder Fig_S1B_3M_MSN_cilia

Microscopy images (.czi) of striatal medium spiny neurons (MSNs) from 8 animals of 3 months of age. Four animals are wild type (WT) mice (Br01, Br02, Br03, Br04), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05, Br06, Br07, Br08).

Folder Fig_S1D_12M_MSN_cilia

Microscopy images (.czi) of striatal medium spiny neurons (MSNs) from 14 animals of 12 months of age. Seven animals are wild type (WT) mice (Br01_set1, Br02_set1, Br03_set1, Br01_set2, Br02_set2, Br03_set2, Br04_set2), and seven are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br04_set1, Br05_set1, Br06_set1, Br05_set2, Br06_set2, Br07_set2, Br08_set2).

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. S1C and S1E. Dopamine- and cAMP-Regulated Phosphoprotein (DARPP32) is a cell marker for medium spiny neuron (MSN).

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. S1C and S1E. The number of cells possessing a cilium was manually quantified, and values excluded as outliers (e.g., partially imaged cells) were labeled as NA.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) imported into GraphPad Prism to generate Fig. S1C and S1E.

Fig.S2.zip file:

Folder Fig_S2A_12M_Aldh1l1_Bdnf_cilia

Microscopy images (.czi) of striatal Aldh1l1 astrocyte from 8 animals of 12 months of age. Four animals are wild type (WT) mice (Br01_set2, Br02_set2, Br03_set2, Br04_set2), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (Br05_set2, Br06_set2, Br07_set2, Br08_set2). One additional image set (no-probe control) was acquired from a WT animal. Aldh1l1 denotes Aldehyde Dehydrogenase 1 Family Member L1, a marker for the astrocyte. Bdnf denotes Brain derived neurotrophic factor.

GraphPad Prism files

GraphPad Prism files (.pzf) containing the graphs and analyses used to generate Fig. S2B and S2C.

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. S2B and S2C. The number of cells possessing a cilium and the number of Brain derived neurotrophic factor (Bdnf) RNA dot were manually quantified, and values excluded as outliers (e.g., partially imaged cells) were labeled as NA.
• Processed tabular data for Prism (.csv):
  Averaged values (per mouse brain) imported into GraphPad Prism to generate Fig. S2B and S2C.

Fig.S3.zip file:

Folder Fig_S3A_2B_2C_2D_12M_expansion_microscopy_WT_G51D_images

Expansion microscopy images (.czi) of olfactory sensory neurons (OSNs) in the olfactory epithelium from 4 animals of 12 months of age. Two animals are wild type (WT) mice (wt1, wt2), and two are G51D-alpha-synuclein (G51D) homozygous knock-in mice (g51d1, g51d2). Note that each mouse contributes 25 image files.

Fig.S4.zip file:

.tif file for Fig_S4A_relative_ciliary_vulnerability_index

Folder Fig_S4B_WB_pRab10_pRab12_images

Western blotting (WB) images (.tif) of the whole brain lysates from 8 animals of 9 months of age. Four animals are wild type (WT) mice (WT1, WT2, WT3, WT4), and four are G51D-alpha-synuclein (G51D) homozygous knock-in mice (G51D1, G51D2, G51D3, G51D4).

Tabular data

• Raw tabular data (.csv):
  Individual (raw) values used for analysis of Fig. S4C. Quantitative immunoblotting analysis with the indicated antibodies (pRab10, total Rab10, pRab12, total Rab12) and data analyzed using the LI-COR Odyssey IRdye imaging and the gel scanning plugin in ImageJ.

Key_Resource_Table_Table_S1_ver2.csv:

This table (.csv) lists all key resources used in this study.

• Resource Type: Categorizes each entry (e.g., genetic reagent such as Mus musculus (mouse), antibody, chemical, peptide, recombinant protein, critical commercial assay, dataset, protocol, or software/code).

• Resource Name: Lists the specific name of each resource or reagent.

• Source: Indicates the vendor or company, as well as repositories or references where the resource was obtained, deposited, or cross-referenced.

• Identifier: Provides unique identifiers for each resource, including catalog numbers, RRIDs, DOIs, or relevant publications.

• New/Reuse: Specifies whether the resource was newly generated in this study or reused from prior work.

• Additional Information: Includes essential details for reproducibility, such as antibody titers used, availability of RRIDs, and any other relevant study-specific notes.

Code/software

• Microsoft Excel (or equivalent spreadsheet software) to open .csv files
• GraphPad Prism 10 or newer to open Prism files (.pzf)
• ZEISS ZEN 3.4 or newer to open .czi files (alternatively, FIJI/ImageJ can also be used)
• FIJI/ImageJ to run the macros and view .tif files.
• RStudio to open and run the R scripts
• CellProfiler 4.2.6 or newer to open the CellProfiler pipeline