Journal: Journal of Advanced Research

Article Title: Tankyrase activity is essential for asymmetric division and chromosome segregation in oocyte meiosis

doi: 10.1016/j.jare.2025.07.008

Figure Lengend Snippet: Expression and distribution of TNKS in mouse oocyte meiosis. (A) Expression level of TNKS1 and TNKS2 at GV (0 h), GVBD (2 h), MI (8 h), and MII; (12 h) stages in mouse oocytes. (B) Representative images of oocytes from GV to MII; stages stained with anti-TNKS antibody. Black, TNKS. Blue, DNA. Bar = 10 μm. (C) Representative images of oocytes co-stained with TNKS and α-Tubulin. Red, TNKS. Green, α-Tubulin. Blue, DNA. Bar = 20 μm. (D) Representative images of oocytes at the MI stage co-stained with TNKS and α-Tubulin after Nocodazole or Taxol treatment. Red, TNKS. Green, α-Tubulin. Blue, DNA. Bar = 20 μm. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Rabbit anti-α-Tubulin antibody (11224-1-AP), rabbit polyclonal anti-GAPDH antibody (10494-1-AP), rabbit anti-GRP78 antibody (11587-1-AP), rabbit anti-TNKS1 antibody (18030-1-AP), rabbit anti-Ran antibody (10469-1-AP), and rabbit anti-Rab11a antibody (67902-1-Ig), rabbit anti-Securin antibody (18040-1-AP), rabbit anti-CDC25C antibody (66912-1-Ig), rabbit anti-Formin2 antibody (11259-1-AP), rabbit anti-BubR1 (11504-2-AP) were from Proteintech.

Techniques: Expressing, Staining

Journal: Journal of Advanced Research

Article Title: Tankyrase activity is essential for asymmetric division and chromosome segregation in oocyte meiosis

doi: 10.1016/j.jare.2025.07.008

Figure Lengend Snippet: TNKS regulates ARP2/Rab11a/Fascin for chromosome migration in mouse oocytes. (A) Representative images and relative intensity of cytoplasmic actin filaments in MI stage oocytes from the control (n = 39) and JW55 treatment (n = 38) groups. Green, α-Tubulin. Red, F-Actin. Blue, DNA. Bar = 10 μm. ***, P < 0.001. (B) Representative images and relative intensity of cortical actin filaments in MI stage oocytes from the control (n = 44) and JW55 treatment (n = 40) groups. Black, F-Actin. Blue, DNA. Bar = 20 μm. ***, P < 0.001. (C) GO enrichment analysis of TNKS-associated proteins based on mass spectrometry data. (D) Protein-protein interaction network analysis of actin-related proteins identified by mass spectrometry upon STRING database. Different node color indicated core and non-core nodes, while edge thickness edge represented interaction strength. (E) Band intensity analysis of N-WASP, Fascin, ARP2, and Ran in the MI stage oocytes from control and JW55 treatment groups. *, P < 0.05. (F) Co-IP analysis with an anti-TNKS antibody. The immunoblots were probed with antibodies against Fascin, Ran, and pS19-Myosin II;. (G) Representative images and relative intensity of Ran in MI stage oocytes from the control (n = 50) and JW55 treatment (n = 47) groups. Red, Ran. Blue, DNA. Bar = 10 μm. **, P < 0.01. (H) Representative images of ARP2 in MI or TI stage oocyte from the control and JW55 treatment groups. Chromosome positions were categorized into center, center-one quadrant, and one quadrant of the oocyte. The arrow highlighted the accumulation of ARP2 in the cortex overlying the chromosomes. Red, ARP2. Black, DNA. Bar = 20 μm. (I) The percentage of asymmetric cortical ARP2 distribution in the control (n = 42) and JW55 treatment (n = 43) oocytes. Accumulation of ARP2 in the cortex overlying the chromosome was defined as asymmetric. **, P < 0.01. (J) The percentage of chromosome position patterns in the control (n = 42) and JW55 treatment (n = 43) oocytes. *, P < 0.05. **, P < 0.01. ***, P < 0.001. (K) Representative images and relative intensity of Fascin in MI stage oocytes from the control (n = 61) and JW55 treatment (n = 51) groups. Green, Fascin. Blue, DNA. Bar = 20 μm. ***, P < 0.001. (L) Representative images and relative intensity of Rab11a in MI stage oocytes from the control (n = 54) and JW55 (n = 49) treatment groups. Red, Ran. Blue, DNA. Bar = 10 μm. *, P < 0.05. (M) Co-IP analysis with an anti-TNKS antibody. The immunoblots were probed with an anti-Rab11a antibody. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Rabbit anti-α-Tubulin antibody (11224-1-AP), rabbit polyclonal anti-GAPDH antibody (10494-1-AP), rabbit anti-GRP78 antibody (11587-1-AP), rabbit anti-TNKS1 antibody (18030-1-AP), rabbit anti-Ran antibody (10469-1-AP), and rabbit anti-Rab11a antibody (67902-1-Ig), rabbit anti-Securin antibody (18040-1-AP), rabbit anti-CDC25C antibody (66912-1-Ig), rabbit anti-Formin2 antibody (11259-1-AP), rabbit anti-BubR1 (11504-2-AP) were from Proteintech.

Techniques: Migration, Control, Mass Spectrometry, Co-Immunoprecipitation Assay, Western Blot

Journal: Journal of Advanced Research

Article Title: Tankyrase activity is essential for asymmetric division and chromosome segregation in oocyte meiosis

doi: 10.1016/j.jare.2025.07.008

Figure Lengend Snippet: TNKS regulates p-PLK1 and microtubule stability for spindle assembly in mouse oocyte. (A) Representative images and percentage of abnormal spindle in MI stage oocyte from the control (n = 47) and JW55 treatment (n = 42) groups. Green, α-Tubulin. Blue, DNA. Bar = 20 μm. *, P < 0.05. (B) Representative images and percentage of abnormal spindle in MII; stage oocyte from the control (n = 57) and JW55 treatment (n = 45) groups. Green, α-Tubulin. Blue, DNA. Bar = 20 μm. **, P < 0.01. (C) Quantitative analysis of chromosome alignment in MI stage oocyte from the control (n = 50) and JW55 treatment (n = 34) groups. We defined the length on both sides of chromosomes as L and the diameter of oocytes as D. The ratio of L/D was markedly increased in the JW55-treated oocytes compared to the control groups. White, DNA. Bar = 20 μm. *, P < 0.05. (D) Representative images and relative intensity of H3S10ph in MI stage oocyte from the control (n = 56) and JW55 treatment (n = 58) groups. Red, H3S10ph. Blue, DNA. Bar = 5 μm. *, P < 0.05. (E) Representative images of γ-Tubulin distribution in MI or MII; stage oocytes from the control and JW55 treatment groups. Magenta, γ-Tubulin. Green, α-Tubulin. Blue, DNA. Bar = 20 μm. (F) The percentage of abnormal γ-Tubulin distribution in MI/MII; stage oocytes from the control (n = 36/58) and JW55 treatment (n = 43/42) groups. *, P < 0.05. **, P < 0.01. (G) Band intensity analysis of p-PLK1 in MI stage oocytes from the control and JW55 treatment groups. *, P < 0.05. (H) Protein-protein interaction network analysis of microtubule-related proteins identified by mass spectrometry of TNKS and PLK1 upon STRING database. Different node color indicated core and non-core nodes, while edge thickness edge represented interaction strength. (I) Co-IP analysis with an anti-TNKS antibody. The immunoblots were probed with an anti‐p-PLK1 antibody. (J) Western blot results of Ac-Tubulin expression in MI stage oocytes from the control and JW55 treatment groups. (K) Relative intensity of Ac-Tubulin in the MI stage oocytes from the control and JW55 treatment groups. *, P < 0.05. (L) Representative images and relative intensity of Ac-Tubulin in MI stage oocytes from the control (n = 50) and JW55 treatment (n = 47) groups. Red, Ac-Tubulin. Blue, DNA. Bar = 10 μm. ***, P < 0.001. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Rabbit anti-α-Tubulin antibody (11224-1-AP), rabbit polyclonal anti-GAPDH antibody (10494-1-AP), rabbit anti-GRP78 antibody (11587-1-AP), rabbit anti-TNKS1 antibody (18030-1-AP), rabbit anti-Ran antibody (10469-1-AP), and rabbit anti-Rab11a antibody (67902-1-Ig), rabbit anti-Securin antibody (18040-1-AP), rabbit anti-CDC25C antibody (66912-1-Ig), rabbit anti-Formin2 antibody (11259-1-AP), rabbit anti-BubR1 (11504-2-AP) were from Proteintech.

Techniques: Control, Mass Spectrometry, Co-Immunoprecipitation Assay, Western Blot, Expressing

Journal: Cell Discovery

Article Title: Loss-of-function variants in ODAD1 disrupt ODA docking and induce actin cytoskeletal remodeling in primary ciliary dyskinesia

doi: 10.1038/s41421-026-00875-8

Figure Lengend Snippet: a Schematic workflow of the HSVM analysis, including nasal swab collection, medium transfer, and imaging. b Ciliary beat pattern (CBP) of nasal epithelial cells from a healthy control and patients harboring ODAD1 variants. c Kymography analysis of ciliary beating in samples from healthy controls and patients carrying the variants. d Quantification of the CBF of nasal epithelial cells from a healthy control and patients carrying ODAD1 variants. Each data point represents the CBF measured from one ciliary bundle. The data are derived from two independent samples per individual. The total number ( n ) of bundles analyzed for each group is indicated on the graph. e Schematic diagram of ALI cultures of human nasal epithelial cells. f Western blot analysis of th levels of the ODAD1 protein in ALI cultures from a healthy control and patients carrying ODAD1 variants. g Immunofluorescence staining for acetylated α-tubulin (red) and ODAD1 (cyan) in ALI cultures from healthy controls and patients carrying ODAD1 variants. Scale bar, 5 μm. h Kymography analysis of ciliary beating in ALI cultures from healthy controls and patients carrying ODAD1 variants. i Quantification of CBF in ALI cultures from a healthy control and patients carrying ODAD1 variants. Each data point represents the CBF measured from one ciliary bundle. The data are derived from three independent samples per individual. The total number ( n ) of bundles analyzed for each group is indicated on the graph. f – i ALI cultures were analyzed on Day 24 of differentiation. d , i P values were determined using one-way ANOVA with Tukey’s multiple comparison test and are indicated directly in the figures. Data are presented as means ± SEM.

Article Snippet: The following antibodies were used for immunofluorescence analyses: rabbit anti-ODAD1 (1:200; Atlas Antibodies, cat# HPA042524), mouse anti-acetylated α-tubulin (1:1000; Sigma-Aldrich, cat# T7451), rabbit anti-acetylated α-tubulin (1:1000; Cell Signaling Technology, cat# 5335), mouse anti-V5 (1:1000; Invitrogen, cat# R96025 ), mouse anti-CEP164 (1:50; Santa Cruz Biotechnology, cat# sc-515403), rabbit anti-VANGL1 (1:100; Cusabio, cat# CSB-PA848385ESR1HU), mouse anti-E-cadherin (1:200; Abcam, cat #ab231303), rabbit anti-SPEF2 (1:100; Atlas Antibodies, cat #HPA039606) and rabbit anti-CEP43 (Proteintech, cat# 11343-1-AP) antibodies.

Techniques: Imaging, Control, Derivative Assay, Western Blot, Immunofluorescence, Staining, Comparison

Journal: Cell Discovery

Article Title: Loss-of-function variants in ODAD1 disrupt ODA docking and induce actin cytoskeletal remodeling in primary ciliary dyskinesia

doi: 10.1038/s41421-026-00875-8

Figure Lengend Snippet: a – c SEM images of cilia in ALI cultures. a Representative images of the ciliary morphology. b Cilia number per field. Each data point represents the cilium count from one SEM field of view (3,000× magnification). The data were obtained from 2 control individuals (2 fields per individual; total n = 4 fields), 2 patients homozygous for c.705_706insGCAG (PCD-1: 2 fields; PCD-3: 2 fields; total n = 4 fields), and 2 patients homozygous for c.-41-2A > C (PCD-4: 2 fields; PCD-7: 2 fields; total n = 4 fields). c Quantitative analysis of the ciliary orientation. The percentage of fields with normal and abnormal ciliary orientations is shown. Quantification was based on the analysis of 103 fields from 2 control individuals (41 and 62 fields), 104 fields from 2 patients carrying the c.705_706insGCAG variant (PCD-1: 54 fields; PCD-3: 50 fields), and 85 fields from 2 patients carrying the c.-41-2A > C (PCD-4: 44 fields; PCD-7: 41 fields). d – f Immunofluorescence staining for CEP164 (magenta) and phalloidin (red) in ALI cultures. d Representative images. e Number of basal bodies per area. Each data point represents the basal body count within a standardized area from one randomly acquired confocal field of view. The data were obtained from 2 control individuals (3 fields per individual; total n = 6), 2 patients homozygous for c.705_706insGCAG (PCD-1: 3 fields; PCD-3: 3 fields; total n = 6), and 2 patients homozygous for c.-41-2A > C (PCD-4: 3 fields; PCD-7: 3 fields; total n = 6). f Number of basal bodies per MCC. Each data point represents the basal body count within an individual MCC. The data were obtained from 2 control individuals (9 MCCs per individual; total n = 18), 2 patients homozygous for c.705_706insGCAG (PCD-1: 9 MCCs; PCD-3: 9 MCCs; total n = 18), and 2 patients homozygous for c.-41-2A > C (PCD-4: 7 MCCs; PCD-7: 8 MCCs; total n = 15). g – i Immunofluorescence staining for acetylated α-tubulin (cyan) and phalloidin (red) in ALI cultures. g Representative images. h Percentage of MCCs. Each data point represents the percentage of MCCs within one randomly acquired confocal image. The data were obtained from 2 control individuals (2 and 3 images per individual; total n = 5), 2 patients homozygous for c.705_706i n sGCAG (PCD-1: 3 images; PCD-3: 3 images; total n = 6), and 2 patients homozygous for c.-41-2A > C (PCD-4: 3 images; PCD-7: 3 images; total n = 6). i Area of MCCs per cell. Each data point represe n ts the apical surface area of an individual MCC. The data were obtained from 2 control individuals (24 and 25 MCCs per individual; total n = 49), 2 patients homozygous for c.705_706insGCAG (PCD-1: 18 MCCs; PCD-3: 18 MCCs; total n = 36), and 2 patients homozygous for c.-41-2A > C (PCD-4: 19 MCCs; PCD-7: 19 MCCs; total n = 38). All experiments were performed on Day 24 of ALI differentiation, with the data collected from two independent culture batches. P values were determined using one-way ANOVA with Tukey’s multiple comparison test and are indicated directly in the figures. Data are presented as means ± SEM.

Article Snippet: The following antibodies were used for immunofluorescence analyses: rabbit anti-ODAD1 (1:200; Atlas Antibodies, cat# HPA042524), mouse anti-acetylated α-tubulin (1:1000; Sigma-Aldrich, cat# T7451), rabbit anti-acetylated α-tubulin (1:1000; Cell Signaling Technology, cat# 5335), mouse anti-V5 (1:1000; Invitrogen, cat# R96025 ), mouse anti-CEP164 (1:50; Santa Cruz Biotechnology, cat# sc-515403), rabbit anti-VANGL1 (1:100; Cusabio, cat# CSB-PA848385ESR1HU), mouse anti-E-cadherin (1:200; Abcam, cat #ab231303), rabbit anti-SPEF2 (1:100; Atlas Antibodies, cat #HPA039606) and rabbit anti-CEP43 (Proteintech, cat# 11343-1-AP) antibodies.

Techniques: Control, Variant Assay, Immunofluorescence, Staining, Comparison

Journal: Cell Discovery

Article Title: Loss-of-function variants in ODAD1 disrupt ODA docking and induce actin cytoskeletal remodeling in primary ciliary dyskinesia

doi: 10.1038/s41421-026-00875-8

Figure Lengend Snippet: a Schematic representation of the experimental design for lentiviral-mediated ODAD1 rescue in apical-out airway organoids. b , c Western blot analysis of ODAD1 expression in lentivirus-transduced ODAD1 -variant apical-out airway organoids. d , e Immunofluorescence staining for acetylated α-tubulin (green) and V5 (magenta) in apical-out airway organoids with or without lentiviral ODAD1 expression. f CBF in ODAD1 -variant apical-out airway organoids with or without lentiviral ODAD1 expression. g Proportion of apical-out organoids with cilia motility in the control and lentivirus-transduced groups. h Kymography analysis of ciliary beating in ODAD1-rescued and untreated ODAD1 -variant apical-out organoids. i CBP in ODAD1-rescued and untreated ODAD1 -variant apical-out organoids. j Immunofluorescence staining for acetylated α-tubulin (red) and DNAH9 (cyan) in apical-out airway organoids with or without lentiviral ODAD1 expression. All experiments involved at least three independent biological replicates and were performed on Day 14 of apical-out organoid differentiation. f , g P values were determined using two-tailed Student’s t -test, and data are presented as means ± SEM.

Article Snippet: The following antibodies were used for immunofluorescence analyses: rabbit anti-ODAD1 (1:200; Atlas Antibodies, cat# HPA042524), mouse anti-acetylated α-tubulin (1:1000; Sigma-Aldrich, cat# T7451), rabbit anti-acetylated α-tubulin (1:1000; Cell Signaling Technology, cat# 5335), mouse anti-V5 (1:1000; Invitrogen, cat# R96025 ), mouse anti-CEP164 (1:50; Santa Cruz Biotechnology, cat# sc-515403), rabbit anti-VANGL1 (1:100; Cusabio, cat# CSB-PA848385ESR1HU), mouse anti-E-cadherin (1:200; Abcam, cat #ab231303), rabbit anti-SPEF2 (1:100; Atlas Antibodies, cat #HPA039606) and rabbit anti-CEP43 (Proteintech, cat# 11343-1-AP) antibodies.

Techniques: Western Blot, Expressing, Variant Assay, Immunofluorescence, Staining, Control, Two Tailed Test