EMBO/soda-nel huggingface dataset:
Molecular flexibility of DNA as a key determinant of RAD51 recruitment (2019)
Figure 3

Figure 3. Mathematical models describing WT RAD51 polymerisation kinetics on ssDNA and dsDNA.

(A, B) A. Kinetic representation of the ordinary differential equation (ODE) model describing WT RAD51 polymerisation on ssDNA, consisting of five parameters: kp (polymerisation forward rate), ku (unstable reverse rate), kq (quasi-stable reverse rate), ks (stable reverse rate) and KD (protomer-protomer interaction affinity). KD predicts the concentrations of RAD51 polymers of variable length in solution, while kp, ku, kq and ks predict the speed of formation of RAD51 polymers on ssDNA. B. Kinetic representation of the ODE model describing WT RAD51 polymerisation on dsDNA, consisting of four parameters: kp, ku, ks, KD. KD predicts the concentrations of RAD51 polymers of variable length in solution, and kp, ku and ks predict the speed of formation of RAD51 polymers on dsDNA. In panels A and B, purple arrows depict a simplified cartoon representation of RAD51 polymerisation in solution. The model allows for any RAD51 n-mer (1 ≤ n < 16) to associate with any other RAD51 m-mer (1 ≤ m < 16) to form an (m+n)-mer (1 < m + n ≤ 16), and any RAD51 k-mer to dissociate into any combination of n-mers and m-mers (k = n + m). The ssDNA and dsDNA models were calibrated by simultaneously fitting the SPR curves of ssDNA dN-X (dN-8, dN-14, dN-17, dN-50) and dsDNA dN-Xp (dN-5p, dN-8p, dN-11p, dN-50p) using the mode ABC-SMC particles. Mean values ± 1 SD of 3 mode ABC-SMC particles derived from model fits of 3 dN-X and dN-Xp repeats (n = 3). ssDNA kp, ku, kq and ks were fit to the ssDNA SPR curves, and dsDNA kp, ku ks were fit to the dsDNA SPR curves. A single KD was fit to all curves.

(C, D) C. Predicted concentrations of WT RAD51 polymers in solution at equilibrium as a function of WT RAD51 monomer concentration (150 nM, 3 µM, 30 µM), prior to RAD51 injection onto DNA-coated SPR CM5 chips. D. Predicted % WT RAD51 within each polymer state in solution at equilibrium as a function of WT RAD51 monomer concentration (150 nM, 3 µM, 30 µM), prior to RAD51 injection onto DNA-coated SPR CM5 chips. In panels C and D, the KD was fixed to the mean value identified in Figs. 3 A and B (i.e. KD = 1.14 nM), and the concentration of WT RAD51 monomer concentration was varied accordingly. In panel D, the % WT RAD51 values were calculated by multiplying the polymer concentrations by their respective polymer length, and dividing each value by the total RAD51 monomer concentration (i.e. % WT RAD51 = [n-mer] * n / [WT RAD51monomer]). [WT RAD51monomer] was calculated via coomassie staining image quantification using a BSA standard curve. It is important to note that the assumed maximum polymer length in solution (16-mer) leads to an overestimation of the concentration of each n-mer in solution, given that RAD51 is likely to form polymers of length greater than 16 at 150 nM, 3 µM and 30 µM WT RAD51 monomer concentration. However, this overestimation is likely to not alter the conclusion that WT RAD51 forms long polymers in solution at 150 nM - 30 µM concentration.
Molecular flexibility of DNA as a key determinant of RAD51 recruitment (2019)
Figure 4

Figure 4. Kinetics of WT RAD51 binding to DNA of varying flexibility.

A. A depiction of the Bend-to-Capture (BTC) mechanism. RAD51 generates two sequential, non-covalent interactions (a RAD51 protomer-protomer interaction and a RAD51-DNA interaction, or vice versa) faster on flexible DNA (depicted as ssDNA) compared to rigid DNA (depicted as dsDNA). The free RAD51 monomer or polymer in solution (here depicted as a monomer) to be incorporated into the growing polymer is shown with an asterisk. ΔTS is the entropic penalty of restricting DNA bending fluctuations, which depends on DNA persistence length. ΔGRAD51-RAD51 and ΔGRAD51-DNA are binding energies, which do not depend on DNA persistence length, i.e., degrees of freedom within the macrostate of being bound or unbound.

B. Biotinylated DNA molecules of varying flexibility were separately immobilised onto SPR CM5 chips via biotin-streptavidin interaction, and WT RAD51 was injected at the indicated concentrations to measure association and dissociation kinetics. The flexibility of respective DNA, the expected entropic penalties upon RAD51 binding to corresponding DNA and protomer-protomer binding energy contribution of WT RAD51 are indicated in gray, green and blue boxes. H and L in each box denote high and low, respectively.

C. The mean number of WT RAD51 bound to respective DNA oligonucleotides, as measured by SPR (dotted lines) and ODE model fits (solid lines).

D, E. Bar plots of fitted kp values (D) and ks values (E) for each SPR curves. In panel D, all parameters except kp were fixed to the mean values as identified in Fig. 3 A and B, and kp was fitted using lsqcurvefit (MATLAB). In panel E, all parameters except ks and kp were fixed to the mean values as identified in Fig. 3 A and B, and ks and kp were fitted using lsqcurvefit (MATLAB). Only the ks values are reported here. Mean ± 1 SD of 3 ODE model fits (n=3). D: un-paired, one-tailed Mann-Whitney-Wilcoxon tests. E: un-paired, two-tailed Mann-Whitney-Wilcoxon tests. * p ≤ 0.05; ns = non-significant.
Divergent androgen regulation of unfolded protein response pathways drives prostate cancer. (2015)
Figure 1

Correlation of AR and UPR gene expression in prostate cancer cohorts

Possible correlation between AR- and UPR-associated gene expression was assessed in the global gene expression data available in the TCGA Prostate Adenocarcinoma cohort (n = 190) (http://www.cbioportal.org/public-portal/index.do). Tumors were stratified according to AR status into three groups, that is ARlow (n = 60), ARmedium (n = 70), and ARhigh (n = 60). The levels of UPR gene expression in the three groups were compared using Pearson's correlation analysis by the R software and presented as a heatmap. There were significant differences between the three groups (Supplementary Table S2).

The expression profiles of some prominent UPR genes from the data in (A), including ERN1 (IRE1), EDEM1, ATF6, and DNAJC3 (P58IPK), are presented. P-values of the different genes are given.
Divergent androgen regulation of unfolded protein response pathways drives prostate cancer. (2015)
Figure 2

Androgens divergently regulate the UPR arms

LNCaP cells were cultured and treated with R1881 for the indicated times.

A, B mRNA expression levels in LNCaP cells for the indicated genes were investigated using quantitative PCR (qPCR). Controls were treated with vehicle for 84 h and set to 100. Data represent the mean of three independent experiments in triplicate, and bars represent SE. P-values ranged between 1.66 × 10−5 and 0.025, and IRE1α expression in R1881 48 h was *P = 0.013 with respect to vehicle-treated cells using unpaired Student's t-test.

C–E IRE1α, XBP-1S, and XBP-1U mRNA in CWR22 xenografts grown in nude mice and collected at the indicated times after castration. The value at t = 0 was set to 1. Columns represent the mean of at least three independent tumors for each time point, and bars represent SE. P-values ranged between 5.7  ×  10−10 and 0.0002. IRE1α expression at 48 h post-castration was 3.17 × 10−6 with respect to t = 0 using unpaired Student's t-test.

F, G Protein expression in LNCaP cells upon treatment with R1881 for the indicated times by Western blot analysis. Data presented are representative of three independent experiments.

Source data are available online for this figure.
Divergent androgen regulation of unfolded protein response pathways drives prostate cancer. (2015)
Figure 3

AR knockdown differentially influences transcription of the different UPR members

After starvation, LNCaP cells were transfected with control (CTRL) siRNA or AR siRNA. Cells were then treated with R1881 for the indicated times. Controls were treated with vehicle for 48 h.

A Expression level of AR mRNA upon siRNA treatment for 48 h assessed by qPCR in LNCaP cells. Expression in cells transfected with CTRL siRNA was set to 1. Bars represent SE with *P = 0.001 indicating significant difference between AR siRNA- and control siRNA-transfected cells using paired Student's t-test.

B–E Same as in (A), but mRNA expression levels of the indicated UPR genes were determined by qPCR at indicated time points after R1881 stimulation. Expression in cells transfected with CTRL siRNA was set to 100. Bars represent SE. P-values are shown indicating significant difference between AR siRNA- and control siRNA-transfected cells using unpaired Student's t-test.

F Expression of the indicated proteins under conditions indicated on the top label was determined by Western blot analysis. Representative blots for three independent experiments are shown.

Source data are available online for this figure.
Divergent androgen regulation of unfolded protein response pathways drives prostate cancer. (2015)
Figure 4

AR directly binds in the vicinity of different UPR genes

A, B LNCaP (A) or VCaP cells (B) were cultured and treated with vehicle (C) for 48 h or R1881 for 24 and 48 h. The cells were then fixed, and ChIP assay was performed as described in Materials and Methods using AR antibody. The data shown are representative of one experiment in duplicate. Error bars represent SE. *P < 0.01 for LNCaP and *P < 0.04 for VCaP indicate significant difference between C (control) and R1881 using unpaired Student's t-test.
Divergent androgen regulation of unfolded protein response pathways drives prostate cancer. (2015)
Figure 5

IRE1α and XBP-1 are proliferative factors in PCa cells both in vitro and in vivo

A Knockdown of IRE1α or XBP-1 leads to a decrease in cell survival. LNCaP cells were transfected with siRNA targeting either IRE1α or XBP-1 (5 nM) and starved in 2% CT-FCS medium for 3 days before cell viability was measured using the CCK-8 assay. The graph is representative of one experiment in triplicate and was repeated three times with similar results. Error bars represent SD with *P = 6.6 × 10−5 and 4.5 × 10−5 for comparison between Ctrl and siRNA against IRE1α and XBP-1, respectively, using paired Student's t-test.

B XBP-1 rescues the growth defect of siIRE1α-transfected LNCaP cells. LNCaP cells were transfected with 5 nM of indicated siRNA using Lipofectamine RNAiMax reagent. One day after siRNA transfection, the cells were transfected with either vector control (Empty) or Flag-XBP-1S (XBP-1S). Three days after transfection, cells were harvested for Western analysis or cultured for three more days before being applied to cell proliferation assay using the CCK-8 reagent. The data are representative of two experiments in triplicate. Error bars represent SE. *P = 0.02, **P = 8.54 × 10−7 using paired Student's t-test.

C, D IRE1α and XBP-1 knockdown inhibits clonogenic capacity of LNCaP cells. Control LN-Scr (Scr), LN-shIRE1 (shIRE1), or LN-shXBP1 (shXBP-1) cells were cultured for 3 weeks. The colonies formed were stained with crystal violet and photographed. The extent of IRE1α and XBP-1 knockdown was determined by Western blot analysis. The area covered by colonies was quantified using the Gene Tools software (SynGene). The data are representative of three experiments in triplicate. Error bars represent SEM. *P = 4.38 × 10−7 and **P = 3.39 × 10−20 using paired Student's t-test.

E Growth analysis of xenografted LNCaP tumors in nude mice. LNCaP cells expressing shRNA against IRE1α (LN-shIRE1), XBP1 (LN-shXBP-1), or control shRNA (LN-Scr) were subcutaneously implanted into both flanks of male nude mice (6 mice per group). Tumor size was measured at the indicated time points. Representative pictures of the tumors at harvest are shown. Error bars indicate SEM. *P = 0.03 for shIRE1 at week 7, P = 0.02 for shXBP-1 at week 7, **P = 0.01 for both shIRE1 and shXBP-1 at week 8 using unpaired Student's t-test.

F PCNA immunostaining in tumors from animals bearing LN-shIRE1, LN-shXBP-1, or LN-Scr tumors. Scale bars: 100 μm.
Divergent androgen regulation of unfolded protein response pathways drives prostate cancer. (2015)
Figure 6

A small molecule IRE1α inhibitor interferes with prostate cancer cell growth in vivo

LNCaP xenografts were grown in nude mice until palpable. Mice were then intraperitoneally injected with 0.5 mg/kg toyocamycin or saline (Vehicle) (tumor numbers: n = 15, or n = 10, respectively) twice weekly.

Tumor sizes were measured weekly with calipers. Error bars indicate SEM. *P = 0.04; **P = 0.0004; ***P = 0.002 using unpaired Student's t-test.

Same procedure was repeated on VCaP xenografts, with similar findings as in LNCaP xenografts (tumor numbers: n = 9, n = 11). Error bars indicate SEM. *P = 0.02 and **P = 0.0006 using unpaired Student's t-test.

XBP-1S mRNA levels in LNCaP xenografts from animals treated with toyocamycin or vehicle. Tumors were harvested, RNA-extracted, and qPCR performed. Error bars indicate SEM. *P = 0.04 for LNCaP and **P = 0.02 for VCaP using unpaired Student's t-test.

PCNA staining in tumors from animals treated with either toyocamycin or saline. Scale bar: 100 μm.
Divergent androgen regulation of unfolded protein response pathways drives prostate cancer. (2015)
Figure 7

XBP-1S expression in clinical prostate cancer specimens

XBP-1S expression was determined by IHC in two different cohorts of human prostatectomy samples.

Representative pictures of benign and tumor samples.

Tissue microarrays (Wang et al, 2010) containing 25 benign and 283 tumor samples were stained with a XBP-1S specific antiserum and scored by a pathologist. The P-value indicates the difference between XBP-1S staining (strong and moderate) in normal vs cancer cells using Mann–Whitney test.

XBP-1S expression was determined by IHC of a neoadjuvant hormone therapy (NHT) tissue microarray containing samples from hormone naïve (untreated) (n = 25), patients that received NHT for 1–6 months (n = 33), and patients that received NHT for 7–12 months (n = 50), as indicated. Representative images are shown. Scale bars: 100 μm.

Quantitative presentation of the data from (C). *P = 0.006 in unpaired Student's t-test.

A model for AR regulation of UPR in PCa cells: Liganded AR activates the IRE1α pathway and coordinately inhibits the PERK arm of the UPR. In addition, AR inhibits the proapoptotic JNK pathway that may be activated by IRE1α or other pathways. The end result of these AR effects is PCa cell proliferation and survival. An arrow with a solid line indicates direct promotion. An arrow with dashed line indicates indirect/unexplored interactions.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 1

Figure 1. TMEM106B deficiency in mice results lysosome trafficking defects near AIS and ALS/FTLD-related pathology.

(A-B) Spinal cord sections from 5-month-old WT and Tmem106b-/- mice were stained with anti- neurofascin, anti- LAMP1 and anti-Cath D antibodies. Accumulation of LAMP1 and CathD positive vacuoles at the distal end of AIS was observed in Tmem106b-/- motor neurons. Representative images from three independent mice were shown. Scale bar = 10 µm. n=3. Data presented as mean ± SEM. Unpaired Student's t test. ****, p<0.0001.

(C-F) Western blot analysis of p62, ubiquitin (Ub), TDP-43 and p-TDP-43 in RIPA- and urea-soluble fractions from brain (C and E) and spinal cord (D and F) of 16-month-old WT and Tmem106b-/- mice. n=5. Data presented as mean ± SEM. Unpaired Student's t test. *, p<0.05, ***, p<0.001, ****, p<0.0001.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 2

Figure 2. Behavioral deficits of Tmem106b-/-Grn-/- mice.

(A) Tmem106b-/-Grn-/- mice do not show any changes in body weight. Body weight of 5-month-old mice of indicated genotypes were measured and analyzed. n=6-10.

(B, C) Tmem106b-/-Grn-/- mice show reduced activities in the open field test. 4.5-month-old mice were used in the study. Total movement was quantified. n=8-10. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: ****, p<0.0001.

(D, E) Tmem106b-/-Grn-/- mice show abnormal hindlimb clasping behavior. 5-month-old mice were used in the study. Severity score was quantified. n= 8-10. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: ***, p<0.001, ****, p<0.0001.

Data information: For all the behavioral studies, mixed male and female mice were used.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 3

Figure 3. Neuronal loss and increased gliosis in Tmem106b-/-Grn-/- spinal cord.

(A,B) Spinal cord sections (C1-C4) from 5-month-old mice of indicated genotypes were stained with indicated antibodies (A) and the numbers of NeuN, or CHAT positive cells, as well as IBA1 and GFAP intensities were quantified (B). 3-5 sections/mouse were used for quantification. n=3. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: *, p<0.05, **, p<0.01, ***, p<0.001, ****, p<0.0001. Scale bar: 20 µm for CHAT staining, 200 µm for other images.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 4

Figure 4. Altered protein levels of NeuN, PSD95, SYN and GFAP in Tmem106b-/-Grn-/- mice.

(A-D) Western blot analysis of NeuN, PSD95, SYN, GFAP, TMEM106B, PGRN and GAPDH in spinal cord (C5-C8) (A, B) and brain (C, D) lysates from 5-month-old WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- mice.

Data information: n=3. Data presented as mean ± SEM.. One-way ANOVA tests with Bonferroni's multiple comparisons: *, p<0.05, **, p<0.01***, p<0.001, ****, p<0.0001.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 5

Figure 5. Retinal neurodegeneration and gliosis in Tmem106b-/-Grn-/- mice.

(A, B) Retinal sections from 5-month-old mice were stained with indicated antibodies. Thickness of each layer, number of Brn3a positive cells (retinal ganglion cells) per section and length of outer segment as indicated by opsin staining are quantified. (C, D) Increase in apoptosis in Grn-/- and Tmem106b-/-Grn-/- retina as indicated by TUNEL assay. TUNEL positive signals per section were quantified.

(E, F) Increase in Müller glial activation in Tmem106b-/-Grn-/- retina as indicated by GFAP staining. GFAP intensity per section was quantified and normalized to WT control.

(G, H) Increase in microglia activation in Grn-/- and Tmem106b-/-Grn-/- retina as indicated by IBA1 staining. The number of IBA1 positive cells per section was quantified.

Data information: ONL: outer nuclear layer; INL: Inner nuclear layer; OPL: outer plexiform layer; IPL: inner plexiform layer; GCL: ganglion cell layer. n=3-6. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: *, p<0.05, **, p<0.01, ***, p<0.001, ****, p<0.0001. Scale bar = 50 µm.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 6

Figure 6. Transcriptome analysis reveals increased expression of inflammation and lysosome genes in Tmem106b-/-Grn-/- mice.

(A, B) Heatmap illustrating gene expression changes in WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- cortex (A) and spinal cord (B) samples isolated from 2.7-month-old mice. n=3. Genes with FDR p≤0.05, LogFC≥0.5 or ≤0.5 between WT and Tmem106b-/-Grn-/- groups are plotted based on hierarchical clustering analysis.

(C) Heatmap illustrating gene expression changes in the inflammation and lysosome pathways in WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- spinal cord samples.

Data information: Inflammation genes are highlighted in magenta and lysosomal genes are highlighted in blue.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 7

Figure 7. Increases in microglia activation in Tmem106b-/-Grn-/- mice.

(A-D) Western blot analysis of Galectin-3 and GPNMB in the spinal cord (A, B) and brain (C, D) lysates from 5-month-old WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- mice. n=3. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: ***, p<0.001, ****, p<0.0001.

(E) Immunostaining of Galectin-3, CathD and CD68 in spinal cord sections from 5-month-old WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- mice. Scale bar = 10 µm.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 8

Figure 8. Increases in the levels of lipofuscin and lysosome proteins in Tmem106b-/-Grn-/- mice.

(A-B) Lipofuscin accumulation in the retina: the number of auto fluorescent puncta per section was quantified. ONL: outer nuclear layer; INL: Inner nuclear layer; OPL: outer plexiform layer; IPL: inner plexiform layer; GCL: ganglion cell layer. n=3-6. Scale bar = 50 µm.

(C-F) TMEM106B and PGRN deletion results in increased autofluorescence in the spinal cord (C, D) and thalamus (E, F) of 5-month-old mice. n=3. Scale bar = 10 µm.

(G, H) Western blot analysis of lysosomal proteins in the spinal cord lysates from 5-month-old WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- mice. Asterisk indicates non-specific bands. n=3.

Data information: For all the analyses, data are presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: *, p<0.05; **, p<0.01; ***, p<0.001; ****, p<0.0001; ns, no significance.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 9

Figure 9. Lysosomal vacuolization in the axon initial segments of motoneurons in spinal cord sections of Tmem106b-/- and Tmem106b-/-Grn-/- mice.

(A) Spinal cord sections from 5-month-old WT, Tmem106b-/-, Grn-/- and Tmem106b-/-Grn-/- mice were stained with anti-neurofascin, anti-LAMP1 and anti-CathD antibodies. Representative images from three independent mice were shown. Scale bar = 10 µm.

(B) Quantification of LAMP1 positive vacuoles (> 2 µm) for experiments in (A). 4-6 sections/spinal cord from 3 mice were analyzed for each genotype. Data presented as mean ± SEM. Unpaired Student's t test: ****, p<0.0001.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 10

Figure 10. Lysosome abnormalities in Tmem106b-/-Grn-/- astrocytes and microglia.

(A, B) Immunostaining of GFAP (A) or IBA1 (B) with LAMP1 and Cath D in the spinal cord sections from 5-month-old WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- mice. Scale bar = 5 µm.
Loss of TMEM106B and PGRN leads to severe lysosomal abnormalities and neurodegeneration in mice (2020)
Figure 11

Figure 11. Autophagy-lysosome dysfunction in the Tmem106b-/-Grn-/- spinal cord.

(A) Accumulation of autophagosomes and lysosomes in the spinal cord sections from a 5-month-old Tmem106b-/-Grn-/- mouse as shown by transmission electron microscope analysis. Scale bar = 1µm. Arrows: autophagosome; arrow heads: lysosomes; (a): axon.

(B) Accumulation of myelin debris and amorphous inclusions in the microglia in the spinal cord sections from a 5-month-old Tmem106b-/-Grn-/- mouse as shown by transmission electron microscope analysis. Scale bar = 1µm. Arrows: myelin debris; arrow heads: dark amorphous inclusions; (m): microglia.

(C) Western blot analysis of LC3-I and LC3-II in the spinal cord lysates from 5-month-old WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- mice. n=3. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: ****, p<0.0001.

(D, E) Immunostaining of TFE3 and IBA1 in the spinal cord sections from 5-month-old WT, Tmem106b-/-, Grn-/-, and Tmem106b-/-Grn-/- mice. Nuclear signals of TFE3 in IBA1 positive microglia were quantified. n=3. Data presented as mean ± SEM. One-way ANOVA tests with Bonferroni's multiple comparisons: ***, p<0.001, ****, p<0.0001. Scale bar = 10 µm.