EMBO/soda-nel huggingface dataset:
Streptolysin O and its co-toxin NAD-glycohydrolase protect group A Streptococcus from Xenophagic killing. (2013)
Figure 5

SLO and NADase inhibit lysosomal fusion to GAS-containing autophagosomes in oropharyngeal keratinocytes.

A. Confocal microscopy of keratinocytes infected with GAS strain 188, 188SLO-, or 188NADase- demonstrating the association of the lysosomal marker LAMP-1 (red) with GAS (blue) contained within EGFP-LC3 (green)-positive compartments at 1 h, 3 h, and 6 h post-infection. Scale bar = 10 µm. B. Quantification of the percent of GAS within EGFP-LC3-positive compartments that are co-localized with LAMP-1 at 1 h, 3 h, and 6 h. Data represent mean values from three independent experiments in which at least 100 intracellular GAS were quantified for each time point in each experiment.
A force balance model for a cell size-dependent meiotic nuclear oscillation in fission yeast (2022)
Figure 1

Figure 1. Robust oscillation with a distinct frequency in wild-type S. pombe

(A) A x-y plot of a representative track of the SPB during meiotic S phase to prophase in a wild-type zygotic cell.

(B) Tracks of the SPB along the longitudinal (x, blue line) and lateral (y, red line) axes. The 64-minute period that gives the fastest movement of the SPB is indicated by the green bar and analyzed in (C).

(C) The power spectral density of the longitudinal movement during the period indicated by the green bar in (B). A frequency that gives the highest peak of the power spectral density is indicated in the graph.

(D) Spectrogram of the longitudinal movement in (B), which shows temporal transition of the power spectral density.

(E, F) Kymograph and individual images of time-lapse observation of Mal3-GFP (green) and Dli1-3mCherry (magenta) along the longitudinal axis. Yellow arrowheads indicate temporary accumulation of growing plus tips during the SPB is turning. Blue asterisks indicate dynein accumulating at the distal cell end cortex in the direction of the SPB movement. Individual images indicated by (a-d) are shown in (F).

(G) An image of microtubules (GFP-α-tubulin, green), the SPB (Sad1-CFP, blue) and dynein (Dli1-3mCherry, red). The image corresponds to the dotted line shown in (H).

(H) Tracks of the SPB (thick black line) and total lengths of individual microtubules (colored thin lines) along the longitudinal axis. The inner horizontal lines indicate the cell ends. Microtubules that are curled and elongate along the cortex are drawn as if they excess the cell length.

(I) Kymographs of the SPB (Sfi1-mCherry, yellow) and histone H2B (Htb1-CFP, blue) in dhc1∆ mutant. At 20 min (white solid lines), dimethyl sulfoxide (DMSO, as a negative control) or MBC (final concentration of 20 µg/mL) were added. Dashed lines indicate cell shape.

Data information: Scale bars, 5 µm.
A force balance model for a cell size-dependent meiotic nuclear oscillation in fission yeast (2022)
Figure 5

Figure 5. Loss of a plus-end depolymerizing kinesin perturbs the oscillation

(A) Time-lapse observation of microtubules (GFP-Atb2, green), dynein (Dli1-3mCherry, red) and the SPB (Sad1-CFP, blue) in wild-type and klp6∆ cells. Arrowheads indicate the SPB. The arrow shows an extensively elongated microtubule along the cell cortex in a klp6∆ cell.

(B and C) Time-lapse observation of Klp6 (Klp6-GFP, green), dynein (Dli1-3mCherry, red) and microtubules (CFP-Atb2, blue). A magnified view of the square area is shown in (C). Arrowheads indicate accumulation of Klp6 on microtubules extending along the cell cortex.

(D) Tracks of the SPB along the longitudinal (x, blue line) and lateral (y, red line) axes. The 64-minute period that gives the fastest movement of the SPB is indicated by the green bar and analyzed in (E).

(E) The power spectral density of the longitudinal movement during the period indicated by the green bar in (D). A frequency that gives the highest peak of the power spectral density is indicated in the graph.

(F) Spectrogram of the longitudinal movement in (D), which shows temporal transition of the power spectral density.

Data information: Scale bars, 5 µm.

Expanded View Figure Legends
Viral Architecture of SARS-CoV-2 with Post-Fusion Spike Revealed by Cryo-EM (2020)
Figure 1.

Isolation and identification of the SARS-CoV-2 virus.(A) Vero cell was inoculated with BALF sample, and the CPEs were observed at 4-dpi. (B) RNAs were extracted from the cell culture supernatant, and detected using a commercial kit targeting the ORF 1ab (red) and N (blue) genes of SARS-CoV-2. Testing the convalescent plasma IgG antibody (C) and SARA-CoV-2-RBD specific human monoclonal antibodies (D) using the purified SARS-CoV-2 particles. The control plasmas 1 and 2 were obtained from a patient recovered from influenza A virus infection and a healthy volunteer, respectively. The control monoclonal antibody is a human monoclonal antibody specific to influenza A virus generated by our laboratory. The PBS control was serum-free. (E) Viruses were detected by IFA using the patient's plasma, and plasma from a healthy control was used as negative control. Scale bar, 100 μm. (F) Phylogenetic characteristics of BetaCoV/Shenzhen/SZTH-003/2020, and this isolate was indicated with a triangle.
Viral Architecture of SARS-CoV-2 with Post-Fusion Spike Revealed by Cryo-EM (2020)
Figure 2.

Negative stain EM results of SARS-CoV-2.(A). Image of negative stained SARS-CoV-2. Nail-like spikes can be clearly seen. (B). Enlarged view of virion boxed in (A). (C) Zoom-in view of a spike boxed in (B). The shape is depicted by red dot line. Length, the diameter of stem and spike's head are 23nm, 4nm and 7nm, respectively. (D). Three-dimensional surface of post-fusion state S2 protein (EMDB code: 9597) [15]. (E). Projection of post-fusion state S2 protein [15].
Viral Architecture of SARS-CoV-2 with Post-Fusion Spike Revealed by Cryo-EM (2020)
Figure 3.

Cryo-EM results of SARS-CoV-2.(A) and (B). Cryo-EM images of SARS-CoV-2. (C). Zoom-in view of the virion showed in (A). Envelope and nucleocapsid are indicated by green and blue respectively, remarkable spikes are indicated by red triangles. (D). Zoom-in views of the two virions showed in (B), remarkable spikes are indicated by red triangles. (E). Zoom-in view of the spike indicated by yellow triangle in (C). The shape is depicted by yellow dot lines. (F). Zoom-in view of the spike indicated by yellow triangle in (D). The shape is depicted by yellow dot lines.
Cited4 is a sex-biased mediator of the antidiabetic glitazone response in adipocyte progenitors (2018)
Figure 1

Figure 1. Cited4 is a target of rosiglitazone in murine and human adipocyte progenitors promoting beige differentiation.

A mRNA expression in FACS-isolated Lin(Ter119/CD31/CD45)-Sca1+ progenitor cells from female mouse subcutaneous fat, differentiated in the presence of 1 µM cPGI2 or vehicle for the indicated time, as determined by expression profiling (n=3, E-MTAB-3693). ****P=3*10-6 (Day 2), ****P=4*10-7 (Day 4), ****P=1*10-6 (Day 6) in 2x2 ANOVA with Bonferroni posttests (cPGI2 vs. vehicle).

B mRNA expression in MACS-isolated Lin-Sca1+ progenitor cells from female mouse subcutaneous fat, differentiated in the presence of 100 nM Rosi or vehicle for the indicated time, as determined by qRT-PCR (n=4). ****P=1*10-10 (Day 1 and 2), **P=0.001, in 2x2 ANOVA with Bonferroni posttests (Rosi vs. vehicle).

C mRNA expression in female Lin-Sca1+ cells, differentiated in the presence of 100 nM Rosi or vehicle for 8 days, as determined by qRT-PCR (n=3). t-test Cited4-/- vs. Cited4+/+ (Rosi) *P=0.013 (Ucp1), **P=0.004 (Cpt1b), *P=0.026 (Dio2).

D-F mRNA expression in primary SVF cells from human subcutaneous fat, differentiated in the presence of 100 nM Rosi (D) or vehicle (D-F), as determined by qRT-PCR at the indicated time points (n=5 patients). ♀/♂ represent individual data. (D) ****P=3*10-5 (Day 2), ****P=3*10-6 (Day 6), ****P=4*10-9 (Day 10), ****P=3*10-7 (Day 14), in 2x2 ANOVA with Bonferroni posttests (Rosi vs. vehicle). (E, F) Pearson correlation coefficient (r) and P-value are shown.

G mRNA expression in primary SVF cells from human female subcutaneous fat transfected with the indicated siRNA prior to differentiation in the presence of 100 nM Rosi for 9 days, as determined by qRT-PCR (n=3). ***P=0.0002 (CITED4), **P=0.002 (UCP1), *P=0.02 (UCP1), *P=0.035/0.026 (PPARG), ***P=0.0006 (SLC2A4), **P=0.002 (ADIPOQ) in one-way ANOVA with Tukey posttests (vs. siCtrl).

Data information: Data are presented as mean ±SEM except for (D) ♀/♂, (E and F) individual data.
Cited4 is a sex-biased mediator of the antidiabetic glitazone response in adipocyte progenitors (2018)
Figure 2

Figure 2. Cited4 deficiency in progenitors affects Ucp1 protein expression and β3-adrenoreceptor mediated uncoupled respiration.

A-C Quantitative fluorescence microscopy of LipidTOX- and DAPI-stained female Cited4F/F Lin-Sca1+ progenitor cells transfected with Cre or control mRNA prior to differentiation in the presence of 100 nM Rosi for 8 days (n=5). **P=0.002 in t-test (Cre vs. Ctrl). Scale bar is 10 µm.

D, E Ucp1 expression in female Cited4F/F Lin-Sca1+ progenitor cells transfected with Cre or control mRNA prior to differentiation in the presence of 100 nM Rosi for 8 days, as determined by Western blot with VCP as loading control (n=3). ***P=0.0008 in t-test (Cre vs. Ctrl).

F Cellular respiration in female Lin-Sca1+ progenitor cells differentiated in the presence of 100 nM Rosi for 8 days. The extracellular oxygen consumption rate (OCR) was determined upon injection of the indicated substances and normalized to DNA content. CL: CL-316243. Representative experiment (n=9). *P=0.041 (47 min), **P=0.009 (55 min), **P=0.003 (62 min) in 2x2 ANOVA with Holm-Sidak posttests (Cited4-/- vs. Cited4+/+).

G Normalized CL-stimulated uncoupled respiration of cells in (F). Values represent the means of the 3 time points after CL injection after subtraction of the means of the oligomycin time points (n=9). *P=0.027 in t-test (Cited4-/- vs. Cited4+/+).

Data information: Data are presented as mean ±SEM.
Cited4 is a sex-biased mediator of the antidiabetic glitazone response in adipocyte progenitors (2018)
Figure 4

Figure 4. Cited4 deficiency specifically affects rosiglitazone-mediated thermogenic expression in subcutaneous fat.

A, B mRNA expression in scWAT of mice fed a diet with 0.0075% Rosi or control diet for 2.5 weeks, determined by qRT-PCR. t-test Cited4-/- vs. Cited4+/+ (Rosi), (A) n=5/5/6/6, **P=0.003 (Ucp1), **P=0.008 (Cidea), **P=0.007 (Cyc1), *P=0.048 (Ndufb3), (B) n=5/4/5/5, *P=0.022 (Ucp1), *P=0.037 (Cidea).

C, D Ucp1 expression in scWAT of mice treated as in A, as determined by Western blot with VCP as loading control. t-test Cited4-/- vs. Cited4+/+ (Rosi), females: n=5/5/6/6, *P=0.011, males: n=5/4/5/5.

E mRNA expression in scWAT of female mice treated with CL-316,243 (CL) (1 mg/kg/day via Alzet minipumps) or vehicle for 10 days (n=5/7/13, t-test Cited4-/- vs. Cited4+/+ (CL)).

F mRNA expression in scWAT of female mice exposed to 5°C or 23°C for 2 weeks (n=5/10/6, t-test Cited4-/- vs. Cited4+/+ (5°C)).

Data information: Data are presented as mean ±SEM.
Cited4 is a sex-biased mediator of the antidiabetic glitazone response in adipocyte progenitors (2018)
Figure 5

Figure 5. Cited4 deficiency affects energy expenditure under rosiglitazone treatment in female mice.

A Oxygen consumption rate of female mice fed a diet with 0.0075% Rosi for 2.5 weeks, determined by indirect calorimetry and normalized to body weight (n=9/10). Grey bars represent the dark phase.

B Oxygen consumption rate of female mice shown in (A) adjusted for body weight by ANCOVA. 3-day averages of VO2 were calculated for each mouse (n=9/10). *P=0.025 in ANCOVA with Bonferroni posttest (Cited4-/- vs. Cited4+/+).

C Oxygen consumption rate of female mice fed a diet with 0.0075% Rosi for 3.5 weeks and injected with 1 mg/kg CL-316,243 (CL) at the indicated time point, as determined by indirect calorimetry (n=9/10). *P=0.013 (-1 h), *P=0.045 (3 h), *P=0.015 (5 h), *P=0.016 (6 h), *P=0.019 (8 h), *P=0.046 (10 h) in repeated measures 2x2 ANOVA with Holm-Sidak posttests (Cited4-/- vs. Cited4+/+).

D Maximal CL-induced oxygen consumption rate of female mice shown in (C) adjusted for body weight by ANCOVA. Averages of VO2 throughout 3-8 hours post CL injection were calculated for each mouse (n=9/10). *P=0.049 in ANCOVA with Bonferroni posttest (Cited4-/- vs. Cited4+/+).

E Oxygen consumption rate of male mice treated as in (A), determined as in (C) (n=9/10). ANCOVA with Bonferroni posttest (Cited4-/- vs. Cited4+/+).

F Oxygen consumption rate of male mice treated as in (C), determined as in (C) (n=9/10). *P=0.044 (9 h), *P=0.013 (12 h) in repeated measures 2x2 ANOVA with Holm-Sidak posttests (Cited4-/- vs. Cited4+/+).

G Maximal CL-induced oxygen consumption rate of male mice shown in (F) and calculated as in (D) (n=9/10). ANCOVA with Bonferroni posttest (Cited4-/- vs. Cited4+/+).

Data information: Data presented as mean ±SEM.
Cited4 is a sex-biased mediator of the antidiabetic glitazone response in adipocyte progenitors (2018)
Figure 6

Figure 6. Sex-specific involvement of Cited4 in thermogenic expression upon therapeutic rosiglitazone treatment.

A, B mRNA expression in scWAT of mice fed a high-fat diet (HFD) for 11 weeks followed by 5 weeks of HFD with 0.0075% Rosi (qRT-PCR). t-test Cited4-/- vs. Cited4+/+, (A) n=8/7, *P=0.029 (Ucp1), **P=0.009 (Elovl3), **P=0.006 (Cpt1b), **P=0.004 (Cyc1), *P=0.016 (Ndufb3), **P=0.003 (Cd36), **P=0.009 (Gyk), (B) n=7/8, *P=0.026 (Pck1).

C, D Ucp1 protein expression in scWAT of mice treated as in (A), determined by Western blot with VCP as loading control (n=8/7 for females and n=7/8 for males). Arrow indicates the band specific to Ucp1 (see Appendix Fig 4F).

Data information: Data presented as mean ±SEM (A and B), individual mice (D, males) or means of replicate blots with independent lysates from the same mice (D, females).
Cited4 is a sex-biased mediator of the antidiabetic glitazone response in adipocyte progenitors (2018)
Figure 7

Figure 7. Sex-specific involvement of Cited4 in insulin sensitization upon therapeutic rosiglitazone treatment.

A Blood glucose during glucose tolerance test (GTT) on female mice after 11 weeks of HFD and 3 weeks of HFD+Rosi (n=8). ***P=0.001, in repeated measures 2x2 ANOVA with Holm-Sidak posttests (Cited4-/- vs. Cited4+/+).

B Fasting serum insulin in female mice after 10 weeks of HFD ("Before Rosi") or 11 weeks of HFD and 2.5 weeks of HFD+Rosi ("After Rosi") (n=8). ***P=0.0009 in repeated measures 2x2 ANOVA with Holm-Sidak posttests (After Rosi vs. Before Rosi); P=0.725 (Cited4-/- vs. Cited4+/+ Before Rosi).

C Blood glucose during GTT on male mice as in (A) (n=5/8). Repeated measures 2x2 ANOVA with Holm-Sidak posttests (Cited4-/- vs. Cited4+/+).

D Fasting serum insulin in male mice as in (C) (n=7/7/8/8). ***P=0.001 (Cited4+/+), ***P=0.0008 (Cited4-/-) in repeated measures 2x2 ANOVA with Holm-Sidak posttests (After Rosi vs. Before Rosi).

E, F Blood glucose during insulin tolerance test (ITT) with 1 U insulin per kg body mass on female mice after 12 weeks of HFD (n=8) (E) or 16 weeks of HFD and 4 weeks of HFD+Rosi (n=7) (F), expressed as % of the 0-time point value. **P=0.003 in repeated measures 2x2 ANOVA with Holm-Sidak posttests (Cited4-/- vs. Cited4+/+).

G Area under the curve (AUC) of blood glucose during the ITT (0-120 min) in (E,F). 2x2 ANOVA with Holm-Sidak posttests (After Rosi vs. Before Rosi).

H, I Blood glucose during insulin tolerance test (ITT) on male mice as in (E,F), n=8 (H), n=7/8 (I). Repeated measures 2x2 ANOVA with Holm-Sidak posttests (Cited4-/- vs. Cited4+/+).

J Area under the curve (AUC) of blood glucose during the ITT in (H,J). **P=0.001, ***P=0.00006 in 2x2 ANOVA with Holm-Sidak posttests (After Rosi vs. Before Rosi).

K, L Fasting serum non-esterified fatty acids (NEFA) in female (K) or male (L) mice after 10 weeks of HFD ("Before Rosi") or 11 weeks of HFD and 2.5 weeks of HFD+Rosi ("After Rosi"), n=7/8/8/8 (K), n=6/6/6/8 (L). **P=0.001 in 2x2 ANOVA with Holm-Sidak posttests (After Rosi vs. Before Rosi).

Data information: Data presented as mean ±SEM.

Expanded View figure legends
A conserved cysteine-based redox mechanism sustains TFEB/HLH-30 activity under persistent stress (2020)
Figure 2

Figure 2. TFEB and TFE3 oligomerization is mediated by a single cysteine residue.

A, Multi sequence alignment of TFEB orthologs showing conserved homology between different species of the region containing TFEB Cysteine 212.

B, Sequence homology analysis between human TFEB and TFE3 depicting the region containing the single cysteine residue in the entire proteins. Multi sequence alignment of TFE3 orthologs showing conserved homology between different species of the region containing TFE3 Cysteine 322.

C, Immunoblot analysis of protein lysates from ARPE-19 cells expressing TFEB-FLAG-WT or mutants treated with 250 μM NaAsO2 or EBSS for 1h and run under non-reducing condition.

D, Immunoblot analysis of protein lysates from ARPE-19 cells expressing TFE3-GFP-WT or mutants run under non-reducing condition.

E, Immunoblot analysis of co-immunoprecipitated TFEB-FLAG and 14-3-3 from ARPE-19 cells over-expressing TFEB-WT or TFEB-C212S mutant.

F, Immunoblot analysis of co-immunoprecipitated TFEB-FLAG, 14-3-3 and endogenous TFE3 from ARPE-19 cells over-expressing TFEB-WT or TFEB-C212A mutant. Immunoblots are representative of at least three independent experiments.
A conserved cysteine-based redox mechanism sustains TFEB/HLH-30 activity under persistent stress (2020)
Figure 4

Figure 4. TFEB activation in response to stress does not require oligomerization.

A, Immunofluorescence confocal microscopy of ARPE-19 cells overexpressing TFEB-FLAG-WT or TFEB-FLAG-C212A showing the subcellular distribution of recombinant TFEB in response to treatments with EBSS for 4 h or 250 μM NaAsO2 for 1 h. Scale bars, 10 μm.

B, Quantification of the nuclear localization of recombinant TFEB in ARPE-19 cells treated with 250 μM NaAsO2 for 1 h or EBSS for 4h and incubated in complete medium (refeed) for the indicated times. Data are presented as mean ± SD using two-way ANOVA, (ns) not significant as compared to the same treatment in TFEB-WT overexpressing cells, with >200 cells counted per trial from three independent experiments.

C, Immunoblot analysis of protein lysates from MEF cells treated with EBSS, 250 nM Torin-1 or 100 μM NaAsO2 for 1 h, subjected to subcellular fractionation and run under non-reducing condition.

D, Relative quantitative RT-PCR analysis of the mRNA expression of autophagy- (UVRAG), mitochondria- (PGC-1α) and lysosome (MCOLN1, ATPV1C1 and HEXA)-related genes in ARPE-19 cells overexpressing TFEB-FLAG-WT or TFEB-FLAG-C212A. Data are presented as mean ± SD using two-way ANOVA, (ns) not significant as compared to the same gene in TFEB-WT overexpressing cells, from three independent experiments.
A conserved cysteine-based redox mechanism sustains TFEB/HLH-30 activity under persistent stress (2020)
Figure 5

Figure 5. Increased stability of TFEB and TFE3 oligomers in response to prolonged stress conditions.

A, Immunoblot analysis of protein lysates from ARPE-19 and HeLa cells treated for the indicated times with 100 μM and 50 μM NaAsO2 respectively and run under non-reducing conditions.

B, Quantification of immunoblot data shown in (A). Data are presented as mean ± SD using one-way ANOVA (*)p<0.05, (**)p < 0.01, (***)p<0.001, and (****)p<0.0001 from three independent experiments.

C, Quantification of immunoblot data shown in (A). Data are presented as mean ± SD using one-way ANOVA (*)p<0.05, (**)p< 0.01, and (***)p<0.001 from three independent experiments.

D, Immunoblot analysis of protein lysates from ARPE-19 cells overexpressing TFEB-FLAG-WT or TFEB-FLAG-C212A treated with 100 μM NaAsO2 for the indicated times and run under non-reducing conditions.

E, Quantification of immunoblot data from TFEB-WT shown in (D). Data are presented as mean ± SD using one-way ANOVA (*)p<0.05, (**)p< 0.01, and (***)p<0.001 from three independent experiments. Some error bars are the same size or smaller than the data point symbols.

F, Immunofluorescence confocal microscopy of ARPE-19 cells showing the nuclear localization of TFE3 in response to treatment with 100 μM NaAsO2 for 8 h. Scale bars, 10 μm.

G, Relative quantitative RT-PCR analysis of the mRNA expression of lysosome-related genes (MCOLN1 and ATPV61C1) in ARPE-19 and HeLa cells treated with 100 μM and 50 μM NaAsO2 respectively for the indicated times. Data are presented as mean ± SD using one-way ANOVA (*)p<0.05, (**)p< 0.01, (***)p<0.001 from two independent experiments.
A conserved cysteine-based redox mechanism sustains TFEB/HLH-30 activity under persistent stress (2020)
Figure 6

Figure 6. Oligomers make TFEB and TFE3 less accessible to inactivation by mTORC1.

A, Immunoblot analysis of protein lysates from HeLa cells treated with EBSS for 1 h and then incubated with complete medium (refeed) for the indicated times. Samples were run under non-reducing conditions.

B, Quantification of immunoblot data shown in (A). Data are presented as mean ± SD using one-way ANOVA (*)p<0.05, and (***)p<0.001 from three independent experiments.

C, Quantification of immunoblot data shown in (A). Data are presented as mean ± SD using one-way ANOVA (*)p<0.05, and (***)p<0.001 from three independent experiments.

D, Relative quantitative RT-PCR analysis of the mRNA expression of autophagy- (UVRAG), lysosome- (ATP6V1C1), oxidative stress response- (TXNRD1 and HMOX1), cell cycle control- (CDKN1a) and cytokine production (IL6) -related genes in TFEB/TFE3-double knockout MEFs cells stably expressing TFEB-FLAG-WT or TFEB-FLAG-C212A and treated with 5 μM arsenic trioxide (ATO) or 12.5 μM NaAsO2 for 12 h. Data are presented as mean ± SD using two-way ANOVA (*)p<0.05, (**)p<0.01, (***)p<0.001, and (****)p<0.0001 from three independent experiments.

E, Schematic representation of TFEB/TFE3 redox regulation in response to chronic stress conditions.
A conserved cysteine-based redox mechanism sustains TFEB/HLH-30 activity under persistent stress (2020)
Figure 7

Figure 7. Oligomer formation and subcellular localization of C. elegans HLH-30.

A, Immunoblot analysis of protein lysates from ARPE-19 cells over-expressing FLAG-tagged C. elegans HLH-30 wild-type or mutated HLH-30(C284A) mutant. Samples were run under non-reducing conditions.

B, Immunoblot analysis of protein lysates from C. elegans knock-in strains expressing HLH-30::3xFLAG::eGFP and HLH-30(C284A)::3xFLAG::eGFP subjected to 4 h of starvation. Samples were run under non-reducing conditions.

C, Representative examples of HLH-30::3xFLAG::eGFP subcellular localization: Cytoplasmic, diffuse GFP distribution along the whole body of the worm; Weak nuclear, some cells in the posterior part of the worm display nuclear GFP labeling but no nuclear labeling is found in the head cells; Strong nuclear, distinct nuclear GFP labeling in cells along the worm body, including neurons in the head area. Arrowheads denote nuclear localization of HLH-30::3xFLAG::eGFP. Scale bar 200 µm and 50 µm.

D, Quantification of the subcellular distribution of HLH-30::3xFLAG::eGFP and HLH-30(C284A)::3xFLAG::eGFP upon treatment with different stimuli. Data are from three independent experiments with at least 50 animals per experiment.