Journal: Communications Biology

Article Title: PACS1 syndrome mutation disrupts dynein-mediated cargo transport via HDAC6 and BICD2

doi: 10.1038/s42003-026-09924-0

Figure Lengend Snippet: a FLAG-HDAC6 co-expressed with BICD2–eGFP and HA-tagged PACS1 or PACS1 R203W was immunoprecipitated from HCT116 cells (M2 agarose); co-precipitating BICD2–eGFP and HA-tagged PACS1 variants detected by Western blot. Data are mean ± SEM, ANOVA with Dunnett’s post hoc, n = 3 independently normalized experiments. b Control (160) and patient (159) fibroblasts nucleofected with BICD2 or nonspecific (ns) siRNAs. After 48 h, cells were collected for Western blot ( bottom left ) or treated overnight with G1 inhibitor ribociclib (3 μM), G2/M inhibitor Ro-3306 (9 μM), or DMSO. Top: Cells were fixed, stained for Giantin (Golgi, red), α-tubulin (green), and DAPI (blue), then imaged by confocal microscopy. Scale bar, 20 μm. Bottom right : Quantification of Golgi dispersal (percentage of Giantin signal within or beyond 10 μm of nuclear envelope). Data are mean ± SEM, ANOVA with Dunnett’s post hoc, n > 50 cells per group from three independent experiments.

Article Snippet: Antibodies: Antibodies against actin (Sigma, MAB1501, 1:5000), AP1M1 (ABclonal, A10129, 1:1000), α/β-tubulin (Cell Signaling Technology (CST), 2148S 1:5000), V5 (Invitrogen, R960-25, 1:2000), FLAG (for WB: Sigma‒Aldrich, F7425, 1:5000 and for IP: Sigma‒Aldrich, A2220, 50% slurry), HA (for WB: CST 3724S, 1:4000; ABclonal AE026, 1:4000; Invitrogen MA1-12429, 1:4000; for IF: Santa Cruz sc-7392, 1:100, Biolegend #901513, 1:800), GFP (Invitrogen, MA5-15256, 1:2000), furin clone MON-152 (Enzo Life Sciences, ALX-803-017, 1:100), Giantin (kindly provided by Dr. A. Linstedt, CMU, 1:750), PACS1 (for IP: Ab 702, 1:100 and for WB: Ab703 , 1:1000), PACS2 (Ab 193 , 1:1000), cytoplasmic dynein heavy chain (Sigma ABT266, 1:1000), Dynein intermediate chain 74.1 (Invitrogen, MA1-070, 1:1000), p150 Glued (CST, 69399, 1:1000), HOOK1 (Proteintech, 10871-1-AP, 1:1000), MIRO1 (also known as RHOT1, ABclonal, A5838, 1:1000), KIF5B (CST, 18148S, 1:1000), BICD2 (Sigma, SAB2702337, 1:1000), LAMTOR1 (CST 8975 T, 1:100), Lis1 (Invitrogen, PA5-20419, 1:1000), DNA-PKcs (CST 4602S, 1:1000), Actinin (CST 3134S, 1:1000), α-tubulin (CST 3873S, 1:1000), Acetyl-α-tubulin (CST 5335, 1:1000), HDAC6 (CST 7558S, 1:1000), Rabbit IgG, whole molecule (Jackson ImmunoResearch Lab., 011-000-003, 1:100).

Techniques: Immunoprecipitation, Western Blot, Control, Staining, Confocal Microscopy

Journal: Communications Biology

Article Title: PACS1 syndrome mutation disrupts dynein-mediated cargo transport via HDAC6 and BICD2

doi: 10.1038/s42003-026-09924-0

Figure Lengend Snippet: a Left: Control (160) and patient (159) cells expressing Lis1-HA, fixed 48 h later, stained for Giantin (Golgi, red), HA (green; pseudocolored magenta), and DAPI (blue). Cell outlines, white. Scale bar, 20 µm. Lis1–HA⁺ cells (*). Scale bar, 20 µm. Right : Quantification of Golgi dispersal (percentage of Giantin signal located within or beyond 10 µm of nuclear envelope). Data are mean ± SEM, two-way ANOVA with Tukey’s post hoc, n > 40 cells per group from three independent experiments. b Top: Control (160) and patient (159) cells treated overnight with 5 µM tubacin or DMSO, fixed, stained for lysosomes (LAMTOR, red), α-tubulin (green), and DAPI (blue). Cell outlines, white. Scale bar, 20 µm. Bottom: Quantification of LAMTOR dispersal (percentage of signal located within or beyond 10 µm of nuclear envelope). Data are mean ± SEM, two-way ANOVA with Tukey’s post hoc, n > 100 cells per group from three independent experiments. c Left : Patient (159) cells expressing Lis1-HA stained as in ( b ); Cell outlines, white. Lis1–HA⁺ cells (*). Scale bar, 20 µm. Right : Quantification of LAMTOR dispersal (percentage of signal located within or beyond 10 µm of nuclear envelope). Data are mean ± SEM, two-way ANOVA with Tukey’s post hoc, n > 70 cells per group from three independent experiments. d U2OS cells expressing PEX3–eYFP–FKBP and BICD2 CC1/2–FRB with Lis1, plus either PACS1 or PACS1 R203W , treated with 1 µM rapalog to induce peroxisome transport. Live imaging (2 fps; six 3-min movies) was used to quantify peroxisome motility. Left: Representative kymographs. Arrowheads, rapalog-induced peroxisome motility in the presence of PACS1 R203W or PACS1 R203W +Lis1. y-axis, time (180 s); x-axis, distance. Scale bar, 2 µm. Top right : Mean peroxisome velocities per cell. Bottom right : Percentage of motile eYFP⁺ peroxisomes per cell over the 18-min interval. Data are mean ± SEM (two-way ANOVA with Tukey’s post hoc); n = 15 / 5547, 17 / 6777, 18 /7916, and 16 / 6438 (cells / total eYFP + peroxisomes) for PACS1, PACS1 + Lis1, PACS1 R203W , and PACS1 R203W + Lis1, respectively, from three independent experiments.

Article Snippet: Antibodies: Antibodies against actin (Sigma, MAB1501, 1:5000), AP1M1 (ABclonal, A10129, 1:1000), α/β-tubulin (Cell Signaling Technology (CST), 2148S 1:5000), V5 (Invitrogen, R960-25, 1:2000), FLAG (for WB: Sigma‒Aldrich, F7425, 1:5000 and for IP: Sigma‒Aldrich, A2220, 50% slurry), HA (for WB: CST 3724S, 1:4000; ABclonal AE026, 1:4000; Invitrogen MA1-12429, 1:4000; for IF: Santa Cruz sc-7392, 1:100, Biolegend #901513, 1:800), GFP (Invitrogen, MA5-15256, 1:2000), furin clone MON-152 (Enzo Life Sciences, ALX-803-017, 1:100), Giantin (kindly provided by Dr. A. Linstedt, CMU, 1:750), PACS1 (for IP: Ab 702, 1:100 and for WB: Ab703 , 1:1000), PACS2 (Ab 193 , 1:1000), cytoplasmic dynein heavy chain (Sigma ABT266, 1:1000), Dynein intermediate chain 74.1 (Invitrogen, MA1-070, 1:1000), p150 Glued (CST, 69399, 1:1000), HOOK1 (Proteintech, 10871-1-AP, 1:1000), MIRO1 (also known as RHOT1, ABclonal, A5838, 1:1000), KIF5B (CST, 18148S, 1:1000), BICD2 (Sigma, SAB2702337, 1:1000), LAMTOR1 (CST 8975 T, 1:100), Lis1 (Invitrogen, PA5-20419, 1:1000), DNA-PKcs (CST 4602S, 1:1000), Actinin (CST 3134S, 1:1000), α-tubulin (CST 3873S, 1:1000), Acetyl-α-tubulin (CST 5335, 1:1000), HDAC6 (CST 7558S, 1:1000), Rabbit IgG, whole molecule (Jackson ImmunoResearch Lab., 011-000-003, 1:100).

Techniques: Control, Expressing, Staining, Imaging

Journal: Communications Biology

Article Title: PACS1 syndrome mutation disrupts dynein-mediated cargo transport via HDAC6 and BICD2

doi: 10.1038/s42003-026-09924-0

Figure Lengend Snippet: a Top: PACS1 supports dynein-dependent endosome-to-TGN trafficking by (i) binding DHC1, adaptors (e.g., AP-1), and acidic cluster/helical cargo motifs (e.g., furin, Nef, Figs. – , , , ) and (ii) interacting with HDAC6 and microtubules to regulate α-tubulin acetylation and vesicle transport velocity (Figs. – , , , ). Dynein adaptors required for Golgi reassembly (e.g., Golgin160, HOOK3) and other PACS1 effectors (AP-3, GGA3, CK2), not shown. Middle: Pacs1 knockout, PACS1 knockdown, or PACS1 Dynmut disrupts retrograde furin trafficking without disturbing Golgi integrity ( , and Fig. ). Loss of PACS1 increases α-tubulin acetylation in cells and in vivo, consistent with its role as an HDAC6 effector . Bottom: PACS1 R203W binds HDAC6, increasing HDAC6 activity, reducing α-tubulin acetylation (Fig. S and ref. ); PACS1 R203W -HDAC6 complexes with BICD2 to suppress dynein activation and transport velocity, disrupting multiple dynein-dependent pathways, including those normally independent of BICD2 (Golgi assembly, lysosome positioning; Figs. , – ). HDAC6 inhibition (tubacin) or Lis1 overexpression counteracts PACS1 R203W –HDAC6–BICD2, thereby restoring dynein motility and organelle positioning (Figs. and ). b AlphaFold2 model of PACS1 FBR, color-coded to show traffic face (β-strands 1, 3, and 7; blue/magenta) and cargo face (β-strands 2, 4, 5, and 6; dark salmon). Surfaces involved in binding DHC1, AP-1/AP-3, GGA3, CK2β, HIV-1 Nef, HDACs 2/3/6, and CK2-phosphorylated furin cytoplasmic domain are indicated (see Discussion and ref. ). Created with BioRender.com.

Article Snippet: Antibodies: Antibodies against actin (Sigma, MAB1501, 1:5000), AP1M1 (ABclonal, A10129, 1:1000), α/β-tubulin (Cell Signaling Technology (CST), 2148S 1:5000), V5 (Invitrogen, R960-25, 1:2000), FLAG (for WB: Sigma‒Aldrich, F7425, 1:5000 and for IP: Sigma‒Aldrich, A2220, 50% slurry), HA (for WB: CST 3724S, 1:4000; ABclonal AE026, 1:4000; Invitrogen MA1-12429, 1:4000; for IF: Santa Cruz sc-7392, 1:100, Biolegend #901513, 1:800), GFP (Invitrogen, MA5-15256, 1:2000), furin clone MON-152 (Enzo Life Sciences, ALX-803-017, 1:100), Giantin (kindly provided by Dr. A. Linstedt, CMU, 1:750), PACS1 (for IP: Ab 702, 1:100 and for WB: Ab703 , 1:1000), PACS2 (Ab 193 , 1:1000), cytoplasmic dynein heavy chain (Sigma ABT266, 1:1000), Dynein intermediate chain 74.1 (Invitrogen, MA1-070, 1:1000), p150 Glued (CST, 69399, 1:1000), HOOK1 (Proteintech, 10871-1-AP, 1:1000), MIRO1 (also known as RHOT1, ABclonal, A5838, 1:1000), KIF5B (CST, 18148S, 1:1000), BICD2 (Sigma, SAB2702337, 1:1000), LAMTOR1 (CST 8975 T, 1:100), Lis1 (Invitrogen, PA5-20419, 1:1000), DNA-PKcs (CST 4602S, 1:1000), Actinin (CST 3134S, 1:1000), α-tubulin (CST 3873S, 1:1000), Acetyl-α-tubulin (CST 5335, 1:1000), HDAC6 (CST 7558S, 1:1000), Rabbit IgG, whole molecule (Jackson ImmunoResearch Lab., 011-000-003, 1:100).

Techniques: Binding Assay, Knock-Out, Knockdown, In Vivo, Activity Assay, Activation Assay, Inhibition, Over Expression

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Spatio-temporal expression profiles of ( A ) ACTB, ( B ) ACTN4, ( C ) INF2, and ( D ) MYL6 retrieved from BrainSpan. The darker the blue color, the higher the protein expression level in the brain region.

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques: Expressing

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Validity verification of DEDRGs. ( A ) Validation of DEDRGs by Western blotting. ( B ) Statistical plots of SLC7A11, ACTB, ACTN4, INF2, and MYL6. Compared with the saline group, ns = no significance, * p < 0.05, ** p < 0.01. n = 3. ( C ) Location of ACTN4 and INF2 proteins in cells from the HPA database: green represents the target protein, red represents microtubules, yellow represents the endoplasmic reticulum, and blue represents the nucleus (scale bar, 20 µm).

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques: Biomarker Discovery, Western Blot, Saline

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Box plot of differential expression of DEDRGs between normal and tumor samples. ( A ) The differential expression of ACTB in pan-cancer. ( B ) The differential expression of ACTN4 in pan-cancer. ( C ) The differential expression of INF2 in pan-cancer. ( D ) The differential expression of MYL6 in pan-cancer. Compared with the normal samples, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques: Quantitative Proteomics

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Pan-cancer prognostic analysis of DEDRGs using univariate Cox regression, including ( A ) ACTB, ( B ) ACTN4, ( C ) INF2, and ( D ) MYL6.

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques:

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Survival analysis of DEDRG expression in pan-cancer. ( A – H ) Survival curves of ACTB in GBMLGG, LGG, MESO, KIRC, UVM, HNSC, LIHC, LUAD. ( I – N ) Survival curves of ACTN4 in GBMLGG, LGG, MESO, PAAD, LUAD, KIRC. ( O – R ) Survival curves of INF2 in LIHC, HNSC, GBM, LAML. ( S – X ) Survival curves of MYL6 in GBMLGG, LGG, ACC, UVM, LAML, SARC.

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques: Expressing

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Pan-cancer immune infiltration analysis: ( A ) Immunoinfiltration analysis of ACTB in GBMLGG, LGG, MESO, KIRC, UVM, HNSC, LIHC, LUAD, and GBM. ( B ) Immunoinfiltration analysis of ACTN4 in GBMLGG, LGG, MESO, PAAD, LUAD, and KIRC. ( C ) Immunoinfiltration analysis of INF2 in LIHC, HNSC, GBM, and LAML. ( D ) Immunoinfiltration analysis of MYL6 in GBMLGG, ACC, LGG, UVM, LAML, and SARC. The correlation coefficient being positive indicates a positive correlation between two variables; a negative correlation coefficient indicates a negative correlation between two variables. The absolute value of the correlation coefficient represents the degree of correlation: 0–0.3 indicates weak or no correlation; 0.3–0.5 indicates weak correlation; 0.5–0.8 indicates moderate correlation; 0.8–1 indicates strong correlation. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques:

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Single-cell type analysis of DEDRGs, including ACTB, ACTN4, INF2, and MYL6, mainly from glandular epithelial cells, squamous epithelial cells, specialized epithelial cells, endocrine cells, neuronal cells, glial cells, germ cells, trophoblast cells, endothelial cells, muscle cells, adipocytes, pigment cells, mesenchymal cells, undifferentiated cells, and blood and immune cells.

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques:

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Gene mutation analysis of DEDRGs. ( A ) The pan-cancer mutation status of ACTB was determined using the cBioPortal tool. ( B ) ACTB base mutation frequency. ( C ) The pan-cancer mutation status of ACTN4 was determined using the cBioPortal tool. ( D ) ACTN4 base mutation frequency. ( E ) The pan-cancer mutation status of INF2 was determined using the cBioPortal tool. ( F ) INF2 base mutation frequency. ( G ) The pan-cancer mutation status of MYL6 was determined using the cBioPortal tool. ( H ) MYL6 base mutation frequency.

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques: Mutagenesis

Journal: Current Issues in Molecular Biology

Article Title: Disulfidptosis: A New Target for Parkinson’s Disease and Cancer

doi: 10.3390/cimb46090600

Figure Lengend Snippet: Tumor pathological staining of ACTB in glioma, renal cancer, head and neck cancer, liver cancer, and lung cancer; ACTN4 in glioma, pancreatic cancer, lung cancer, and renal cancer; INF2 in liver cancer, head and neck cancer, and glioma; MYL6 in glioma (scale bar, 20 µm).

Article Snippet: The membrane was blocked with 5% nonfat milk in TBST for 1 h at room temperature and probed overnight at 4 °C with primary antibodies: beta Actin (1:3000, AF7018, Affinity, Melbourne, Australia), ACTN4 (1:1000, DF8000, Affinity, Melbourne, Australia), INF2 (1:1500, 20466-1-AP, Proteintech, Chicago, IL, USA), MYL6 (1:1000, DF4718, Affinity, Melbourne, Australia), and GAPDH (1:3000, Affinity, Melbourne, Australia) in TBST containing 1% nonfat milk.

Techniques: Staining

Journal: The FASEB Journal

Article Title: Overexpression of enhanced yellow fluorescent protein fused with Channelrhodopsin‐2 causes contractile dysfunction in skeletal muscle

doi: 10.1096/fj.202401664RR

Figure Lengend Snippet: Western blot antibody information.

Article Snippet: ACTN3 (ProteinTech, #24378‐1‐AP) 1:1000 , Goat anti‐rabbit (ProteinTech #SA00001‐2) 1:6000.

Techniques: Western Blot

Journal: The FASEB Journal

Article Title: Overexpression of enhanced yellow fluorescent protein fused with Channelrhodopsin‐2 causes contractile dysfunction in skeletal muscle

doi: 10.1096/fj.202401664RR

Figure Lengend Snippet: Presence of EYFP led to reduced HCN2 and FBP2 protein levels in gastrocnemius skeletal muscle. Significantly reduced protein‐level expressions of (A) HCN2 in the ChR2‐EYFP muscle and (B) FBP2 in both ChR2‐EYFP and ChR2‐only muscles were observed, compared with ChR2‐only and WT muscles, respectively. (C) ACTN3 protein expression was significantly higher in ChR2‐only muscle compared with WT and ChR2‐EYFP muscles. Error bars denote means ± SD. WT group included n = 1 ChR2‐EYFP fl/fl and n = 2 ChR2‐only fl/fl mice (all cre‐negative).

Article Snippet: ACTN3 (ProteinTech, #24378‐1‐AP) 1:1000 , Goat anti‐rabbit (ProteinTech #SA00001‐2) 1:6000.

Techniques: Muscles, Expressing