Journal: Nature Aging

Article Title: Reduced ULK1 links impaired autophagy and mitophagy to Alzheimer’s disease pathology

doi: 10.1038/s43587-026-01108-z

Figure Lengend Snippet: a , Tau puncta formation (left) and degradation of preformed tau puncta (right) were quantified in HEK293 cells expressing 0N4R P301S Tau-Venus in the presence or absence of ULK1 activator Rac-(BL)-918. n = a total of 50 randomly selected fields per group from 3 biological replicates. b , c , HEK293 P301S tau-Venus cells were transfected with siRNA-ULK1 ( b ), siRNA-ULK2 ( c ) or siRNA-scramble ( b , c ), and transfected cells were incubated with or without 5-μM Rac-(BL)-918 in the presence of 10-nM tau seeds. Graphs in a – c show normalized fluorescence intensity relative to vehicle control ( a ) or in the absence of Rac-(BL)-918 ( b , c ), as indicated. n = a total of 54 randomly selected fields per group from 3 biological replicates. d , Rac-(BL)-918 inhibited the formation of tau puncta dependent on ULK1, FUNDC1, PINK1, Ambra1 and Beclin1. HEK293 P301S tau-Venus cells were transfected with siRNA-scramble or a siRNA candidate, followed by treatment with 10-nM tau seeds and 5-μM Rac-(BL)-918 simultaneously. n = a total of 36–40 randomly selected fields per group from 3 biological replicates. e , Rac-(BL)-918 increased the degradation of preformed tau puncta dependent on ULK1 and Beclin1. HEK293 P301S tau-Venus cells were transfected with siRNA-scramble or a siRNA candidate, followed by subsequent treatment with 10-nM tau seeds and 5-μM Rac-(BL)-918. n = a total of 36–40 randomly selected fields per group from 3 biological replicates. f , Associative memory tests were administered to adult day-2 transgenic C. elegans expressing hTau[P301L] n -sid-1 OV in the presence of siRNA targeting unc-51/ULK1 or control siRNA in the presence of Rac-(BL)-918, where indicated. Data are shown in the format of % CI, where a lower score corresponds to greater response to odorant and better memory function. n = 3 biological sets. g , Associative memory tests were administered to adult day-2 transgenic C. elegans expressing hTau[P301L] n -sid-1 OV in the absence or presence of siRNA to pink-1 or fndc-1/FUNDC1 . Data are shown in the format of % CI, where a lower score corresponds to greater response to odorant and better memory function. n = 5 biological sets. In the box plots, the center line denotes the median, box range indicates the 25th–75th percentile and whiskers denote minimum and maximum values. Unless specified elsewhere, data are mean ± s.e.m. Results represent pooled data from three to five biological replicates. Violin and box plots are centered around the median with interquartile ranges and all data points shown; the shape of the plot reflects the distribution of data. Statistical significance was determined using the Kruskal–Wallis test (non-normal data distribution) or one-way ANOVA (normal data distribution) ( a – e ) or two-way ANOVA followed by Tukey’s multiple comparisons test ( f , g ).

Article Snippet: The siRNA reagents used including siRNA targeting ULK1 (catalog no. SR322391, OriGene), PINK1 (catalog no. SR324912, OriGene), Parkin (catalog no. SR321228, OriGene), FUNDC1 (catalog no. SR315322, OriGene), Ambra1 (catalog no. SR310808, OriGene), BNIP3 (catalog no. SR300461, OriGene), BNIP3L/NIX (catalog no. SR300462, OriGene), GSK3-beta (catalog no. SR301979, OriGene), ULK2 (catalog no. SC-44183, Santa Cruz Biotechnology), Atg5 (catalog no. SR322789, OriGene), Beclin1 (catalog no. SR322490, OriGene), Sirt1 (catalog no. SR323581, OriGene) or scramble control siRNA Oligo Duplexes at 100 nM.

Techniques: Expressing, Transfection, Incubation, Fluorescence, Control, Transgenic Assay

Journal: Nature Aging

Article Title: Reduced ULK1 links impaired autophagy and mitophagy to Alzheimer’s disease pathology

doi: 10.1038/s43587-026-01108-z

Figure Lengend Snippet: ( a ) Effects of ULK1 activators (2, 4 µM of LYN-1604 dihydrochloride; 0.5, 5 µM of (Rac)-BL-918) and inhibitors (5, 10 µM of SBI-0206965; 2.5, 5 µM of XST-14) on Tau seed-induced formation of Tau puncta in HEK293 cells expressing 0N4R P301S Tau-Venus. Data are pooled from 3 biological replicates. (b) Effects of ULK1 activators (2, 4 µM of LYN-1604 dihydrochloride; 0.5, 5 µM of (Rac)-BL-918) and inhibitors (5, 10 µM of SBI-0206965; 2.5, 5 µM of XST-14) on degradation of preformed Tau puncta in the HEK293 cells expression 0N4R P301S Tau-Venus. Data are pooled from 3 biological replicates. ( c ) mt-Keima and YPH-Parkin expressing HeLa cells were used to evaluate the effects on mitophagy induction by treating a positive control CCCP. Representative images are shown. ( d-f ) Quantification of mitophagy events by treating ULK1 activators, Rac-BL-918 (0.5, 5 µM) and LYN-1604 (2, 4 µM), as well as ULK1 inhibitors SBI-0206965 (5, 10 µM) and XST-14 (2.5, 5 µM). One dot represents the average value of one image. Data were pooled from 3 biological replicates. ( g-h ) Representative images ( g ) and quantification ( h ) of mitophagy induction from primary cortical neurons by treating ULK1 activators, Rac-BL-918 (5 µM) and LYN-1604 (4 µM), as well as ULK1 inhibitors SBI-0206965 (5 µM) and XST-14 (5 µM). Mitophagy was detected using a mitophagy detection dye kit. Nuclei were stained with DAPI. Scale bar = 20 μm. ( i ) Representative blots of target proteins (ULK1, ULK2, PINK1, GSK3β, Parkin, Atg5, FUNDC1, AMBRA1, BNIP3, Nix, Beclin1) and loading control (GAPDH or β-tubulin) in HEK293 cells expressing 0N4R P301S Tau-Venus transfected with siRNAs (100 nM, 48 h) or scrambled siRNA. ( j ) Associative memory tests were administered to adult day 2 transgenic C. elegans expressing hTau[P301L]n-sid-1 OV in the presence of ULK1 inhibitors (10, 100 µM of SBI-0206965; 5, 50 µM of XST-14). Attraction to odorant is quantified as Chemotaxis Index (% CI), where lower score corresponds to greater response to odorant/better memory function. Unless specified elsewhere, data are mean ± S.E.M. Statistical analyses performed were one-way ANOVA followed by Dunnett’s multiple comparisons test ( a , b , d-f , h ); two-way ANOVA followed by Tukey’s multiple-comparisons test ( j ).

Article Snippet: The siRNA reagents used including siRNA targeting ULK1 (catalog no. SR322391, OriGene), PINK1 (catalog no. SR324912, OriGene), Parkin (catalog no. SR321228, OriGene), FUNDC1 (catalog no. SR315322, OriGene), Ambra1 (catalog no. SR310808, OriGene), BNIP3 (catalog no. SR300461, OriGene), BNIP3L/NIX (catalog no. SR300462, OriGene), GSK3-beta (catalog no. SR301979, OriGene), ULK2 (catalog no. SC-44183, Santa Cruz Biotechnology), Atg5 (catalog no. SR322789, OriGene), Beclin1 (catalog no. SR322490, OriGene), Sirt1 (catalog no. SR323581, OriGene) or scramble control siRNA Oligo Duplexes at 100 nM.

Techniques: Expressing, Positive Control, Staining, Control, Transfection, Transgenic Assay, Chemotaxis Assay

Journal: Molecular Medicine Reports

Article Title: ULK1 activates NCOA4-mediated ferritinophagy via the Beclin1/VPS34 complex in cardiomyocyte hypertrophy

doi: 10.3892/mmr.2026.13826

Figure Lengend Snippet: ULK1 expression is elevated in hypertrophic cardiac tissues and cardiomyocytes. (A) C57BL/6J mice at 8-weeks-old were subjected to sham or TAC surgery. Representative whole heart images (scale bar, 0.5 cm), M-mode echocardiography, H&E staining (scale bar, 100 µm) and WGA staining (scale bar, 50 µm) of heart tissue. Assessment of (B) HW/BW, (C) LVW/BW and (D) HW/TL (n=6). Statistical graphs for (E) LVEF (%), (F) FS (%), (G) IVS, d (mm) and (H) LVPW, d (mm) (n=6). (I) Cardiomyocyte cross-sectional area acquired from WGA staining. Cells were measured from different microscopic fields of 6 samples in each group. The mRNA levels of (J) BNP and (K) β-MHC in the hearts from sham or TAC-surgery mice (n=6). The mRNA levels of (L) BNP and (M) β-MHC in HL-1 cells treated with Ang II (n=3). (N) Stained cells and (O) quantification of cell surface area of HL-1 cells treated with Ang II. Scale bar, 50 µm (n=3). Western blot analysis and summarized data demonstrating (P) Beclin1 and (Q) VPS34 protein expression in HL-1 cells treated with Ang II (n=3). (R) Western blot analysis and summarized data demonstrating ULK1 protein levels in the hearts obtained after sham or TAC surgery (n=6). (S) Western blot analysis and summarized data demonstrating ULK1 protein levels in the cardiomyocytes stimulated with Ang II (n=6). All data are presented as mean ± SEM. *P < 0.05 and **P<0.01 vs. sham or control group. ULK1, serine/threonine protein kinase ULK1; TAC, transverse aortic constriction; HW, heart weight; BW, body weight; LVW, left ventricular weight; TL, tibia length; LVEF, left ventricular ejection fraction; FS, fractional shortening; d, diastole; IVS, interventricular septal thickness; LVPW, left ventricular posterior wall thickness; CSA, cross-sectional area; BNP , brain natriuretic peptide; β-MHC , β-myosin heavy chain; α-SMA, α smooth muscle actin; Ang II, angiotensin II; VPS34, PI3K catalytic subunit type 3; WGA, wheat germ agglutinin.

Article Snippet: The overexpression plasmids for ULK1 and Beclin1 were constructed by cloning the respective mouse cDNAs into the pcDNA3.1(+) vector backbone (Shanghai GeneChem Co., Ltd.).

Techniques: Expressing, Staining, Western Blot, Control

Journal: Molecular Medicine Reports

Article Title: ULK1 activates NCOA4-mediated ferritinophagy via the Beclin1/VPS34 complex in cardiomyocyte hypertrophy

doi: 10.3892/mmr.2026.13826

Figure Lengend Snippet: Autophagy activation is required for ULK1-induced ferroptosis in HL-1 cells. Western blot analysis and summarized data demonstrating (A) Beclin1, (B) LC3-II and (C) p62 protein levels in HL-1 cells treated with 3-MA. (D) HL-1 cells were infected with mRFP-GFP-LC3-labeled adenoviruses and the formation of autophagosomes (yellow) and autolysosomes (red) were observed in indicated group. Scale bar, 50 µm. (E) Red and yellow dots/cell in indicated group. (F) Cell viability of HL-1 cells was detected using a Cell Counting Kit-8 assay in the 3-MA group. The contents of (G) Fe 2+ , (H) MDA and (I) SOD were determined in 3-MA group. (J) The mRNA level of Ptgs2 . (K) Immunofluorescence staining for 4-HNE in 3-MA group. Scale bar, 50 µm. All data are presented as mean ± SEM. n=3. *P<0.05 and **P<0.01 vs. control group; # P<0.05 and ## P<0.01 vs. ULK1 group. ULK1, serine/threonine protein kinase ULK1; 3-MA, 3-methyladenine; LC3, microtubule-associated protein 1 light chain 3; mRFP, monomeric red fluorescent protein; MDA, malondialdehyde; SOD, superoxide dismutase; Ptgs2 , prostaglandin endoperoxide synthase 2; 4-HNE, 4-hydroxynonenal.

Article Snippet: The overexpression plasmids for ULK1 and Beclin1 were constructed by cloning the respective mouse cDNAs into the pcDNA3.1(+) vector backbone (Shanghai GeneChem Co., Ltd.).

Techniques: Activation Assay, Western Blot, Infection, Labeling, Cell Counting, Immunofluorescence, Staining, Control

Journal: Molecular Medicine Reports

Article Title: ULK1 activates NCOA4-mediated ferritinophagy via the Beclin1/VPS34 complex in cardiomyocyte hypertrophy

doi: 10.3892/mmr.2026.13826

Figure Lengend Snippet: ULK1 activates ferritinophagy dependent on Beclin1/VPS34 complex. (A) Cell viability of HL-1 cells was assessed using a Cell Counting Kit-8 assay in the indicated groups. The levels of (B) Fe 2+ , (C) MDA and (D) SOD were determined. (E) The mRNA levels of Ptgs2 . (F) Immunofluorescence staining for 4-HNE in indicated groups. Scale bar, 50 µm. Western blot analysis and summarized data demonstrating (G) NCOA4 and (H) FTH1 protein levels in indicated group. (I) The colocalization of NCOA4 (red) and LC3-II (green) in HL-1 cells was determined using immunofluorescence staining. Scale bar, 50 µm. All data are presented as mean ± SEM. n=3. *P<0.05 and **P<0.01 vs. control group; # P<0.05 and ## P<0.01 vs. ULK1 group. ULK1, serine/threonine protein kinase ULK1; VPS34, PI3K catalytic subunit type 3; siRNA, small interfering RNA; NC, negative control; MDA, malondialdehyde; SOD, superoxide dismutase; Ptgs2 , prostaglandin endoperoxide synthase 2; 4-HNE, 4-hydroxynonenal; FTH1, ferritin heavy chain 1; NCOA4, nuclear receptor coactivator 4; LC3, microtubule-associated protein 1 light chain 3.

Article Snippet: The overexpression plasmids for ULK1 and Beclin1 were constructed by cloning the respective mouse cDNAs into the pcDNA3.1(+) vector backbone (Shanghai GeneChem Co., Ltd.).

Techniques: Cell Counting, Immunofluorescence, Staining, Western Blot, Control, Small Interfering RNA, Negative Control

Journal: Molecular Medicine Reports

Article Title: ULK1 activates NCOA4-mediated ferritinophagy via the Beclin1/VPS34 complex in cardiomyocyte hypertrophy

doi: 10.3892/mmr.2026.13826

Figure Lengend Snippet: Beclin1/VPS34 complex is associated with NCOA4-mediated ferritinophagy. (A) Cell viability of HL-1 cells was assessed using a Cell Counting Kit-8 assay. The levels of (B) Fe 2+ , (C) MDA and (D) SOD were determined. (E) The mRNA levels of Ptgs2 . (F) Immunofluorescence staining for 4-HNE in indicated group. Scale bar, 50 µm. Western blot analysis and summarized data demonstrating (G) NCOA4 and (H) FTH1 protein levels in the indicated groups. All data are presented as mean ± SEM. n=3. *P<0.05 and **P<0.01 vs. control group; # P<0.05 and ## P<0.01 vs. Beclin1 group. VPS34, PI3K catalytic subunit type 3; NCOA4, nuclear receptor coactivator 4; siRNA, small interfering RNA; NC, negative control; MDA, malondialdehyde; SOD, superoxide dismutase; Ptgs2 , prostaglandin endoperoxide synthase 2; 4-HNE, 4-hydroxynonenal; FTH1, ferritin heavy chain 1.

Article Snippet: The overexpression plasmids for ULK1 and Beclin1 were constructed by cloning the respective mouse cDNAs into the pcDNA3.1(+) vector backbone (Shanghai GeneChem Co., Ltd.).

Techniques: Cell Counting, Immunofluorescence, Staining, Western Blot, Control, Small Interfering RNA, Negative Control

Journal: Molecular Medicine Reports

Article Title: ULK1 activates NCOA4-mediated ferritinophagy via the Beclin1/VPS34 complex in cardiomyocyte hypertrophy

doi: 10.3892/mmr.2026.13826

Figure Lengend Snippet: Mechanisms of ULK1 activating NCOA4-mediated ferritinophagy via Beclin1/VPS34 complex in cardiomyocyte hypertrophy. This figure was created using Figdraw ( www.figdraw.com ). ULK1, serine/threonine protein kinase ULK1; NCOA4, nuclear receptor coactivator 4; VPS34, PI3K catalytic subunit type 3; LC3, microtubule-associated protein 1 light chain 3; TAC, transverse aortic constriction; Ang II, angiotensin II; ROS, reactive oxygen species.

Article Snippet: The overexpression plasmids for ULK1 and Beclin1 were constructed by cloning the respective mouse cDNAs into the pcDNA3.1(+) vector backbone (Shanghai GeneChem Co., Ltd.).

Techniques: