Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: a , Volcano plot showing the PP or PC, fast-to-fed log 2 (fold change (FC)) and the –log 10 ( P ) for identified proteins. Colour coding highlighting significance on the basis of P value and fold change. Proteomics data were analysed using the Limma R package (version 3.40.6). b , Lipid pathways in hepatocytes. OXPHOS, oxidative phosphorylation. c – e , Heatmaps of proteins involved in lipid metabolic pathways ( c ), mitochondrial dynamics and structure ( d ), mitochondria–LD interactions and LD formation ( e ) across PP and PC sorted hepatocytes from fed and fasted mice. Heat map values were derived from the means of three independent experiments and normalized to the total protein expression in livers from fed mice. f , Western blot analysis of PLIN5 levels under different dietary conditions. Data are presented as mean ± s.d. from n = 5 independent experiments. Statistical significance was calculated using a two-tailed unpaired Student’s t -test. Schematic in b created in BioRender ; Porat-Shliom, N. https://BioRender.com/3ps9u0r (2026).

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Phospho-proteomics, Derivative Assay, Expressing, Western Blot, Two Tailed Test

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: (A) Localization of endogenous PLIN5 in liver sections from fed and fasted Dendra2 mice. Scale bar 2 μm. (B) Confocal images of liver sections from mtDendra2 (green) transgenic mice fed control, WD, or overnight fasted or WD-fed-overnight fasted. LDs were labeled with Lipidtox (magenta) and actin with phalloidin (yellow). Colocalization between mitochondria and LD is shown in white. Scale bar 3 μm. (C) Mitochondria-LD colocalization was quantified as the overlapping pixels normalized to the area under different dietary conditions. Data presented as mean ± SD from n = 5 independent experiments. Statistical significance was assessed using one-way ANOVA with a Bonferroni correction for multiple comparisons. WD: western diet; LD: lipid droplets.

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Transgenic Assay, Control, Labeling, Western Blot

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: a , Experimental design schematic. b , Representative confocal images of periportal (PP), mid-lobular (M) and pericentral (PC) hepatocytes from mtDendra2 (green) mice fed CNTR and overexpressing PLIN5 variants. Actin is labelled with phalloidin (yellow), and LDs are labelled with LipidTox (magenta). c , scPhenomics of LD features, including total area, density and count, across the PP–PC axis from mice overexpressing PLIN5 variants. d , scPhenomics of mitochondrial features, including area and circularity, across the PP–PC axis from mice overexpressing PLIN5 variants. scPhenomics data are presented as mean ± s.d. from 12 PP–PC axes analysed from two mice. e , Heat map of mitochondria and LDs overlapping pixels across the PP–PC axis (R1–R12) from mice overexpressing PLIN5 variants. Icons in a created in BioRender ; Porat-Shliom, N. https://BioRender.com/mzkgcrm (2026).

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques:

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: (A) Representative confocal images of liver sections from control (CNTR) diet-fed mice overexpressing PLIN5 variants. Three mice per PLIN5 variant were analyzed. Images show mitochondria (green), LDs (magenta), and actin (yellow). Scale bar 20 μm. PV: portal vein; CV: central vein. WT: wild type.

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Control, Variant Assay

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: a , Representative confocal images of periportal (PP), mid-lobular (M) and pericentral (PC) hepatocytes from mtDendra2 (green) mice fed a WD and overexpressing PLIN5 variants. Actin is labelled with phalloidin (yellow), and LDs are labelled with LipidTox (magenta). b , Heat map of mitochondria and LDs overlapping pixels across the PP–PC axis (R1–R12) from WD-fed mice overexpressing PLIN5 variants. c , d , Triglyceride ( c ) and FFAs ( d ) were measured in the livers of mice overexpressing PLIN5 variants, fed either CNTR or WD. Statistical significance was calculated with two-way ANOVA and Tukey’s test to correct for multiple comparisons. Data presented as mean ± s.d. from n = 10 independent experiments. e – g , Malondialdehyde (MDA) ( e ), NADP/NADPH ( f ) and reduced glutathione (GSH) ( g ) were measured in the livers of mice overexpressing PLIN5 variants, fed either CNTR or WD. The dotted line represents WD equals mean CNTR. Values are presented as mean WD/CNTR diet ratios ± s.d. from n = 10 independent experiments. For each assay, a linear model was fit with diet (CNTR versus WD) and genotype as factors. The WD effect for each variant was compared to the Null group. In each variant group, WD was compared with the mean CNTR diet to test whether it differed from 1 (* q < 0.15; ** q < 0.01; *** q < 0.01). Moderated t -statistics and associated P values were obtained using empirical Bayes shrinkage (eBayes). A multiple testing correction was applied across assays using the Benjamini–Hochberg method, and adjusted P values ( q values) were calculated using two-way ANOVA.

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Variant Assay

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: (A) Representative confocal images of liver sections from Western Diet (WD)-fed mice overexpressing PLIN5 variants. Three mice per PLIN5 variant were analyzed. Images show mitochondria (green), LDs (magenta), and actin (yellow). Scale bar 20 μm. (B) Weight gain in mice overexpressing PLIN5 variants and fed CNTR or WD (n = 10 mice per diet). Serum FA (C) , cholesterol (D) , and glucose (E) levels in mice overexpressing PLIN5 variants and fed CNTR or WD (n = 5 per diet). Data presented as mean ± SD, statistical significance was calculated with two-way ANOVA and Tukey’s test to correct for multiple comparisons. PV: portal vein; CV: central vein; WT: wild type. CNTR: control; WD: western diet; FA: fatty acid.

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Western Blot, Variant Assay, Control

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: (A) Relative expression of mRNA levels of lipogenesis enzymes Fatty acid synthase (Fasn), Acetyl-CoA carboxylase 1 (Acaca), ATP citrate lyase (Acly), Stearoyl-CoA desaturase-1 (Scd1) in mice overexpressing PLIN5 variants fed a control diet (CNTR), or 4-week WD-fed. Data presented as mean ± SD from n = 5 independent experiments. Statistical significance was calculated using a two-way ANOVA and Tukey’s test to correct for multiple comparisons. (B) Fatty acid synthase (FASN) expression levels in mice overexpressing PLIN5 variants and fed CNTR or WD (n = 10 mice per diet). (C) Relative phosphorylated Acetyl-CoA carboxylase 1 (ACC1) and phosphorylated hormone-sensitive lipase (HSL) in mice overexpressing PLIN5 variants fed a control diet (CNTR), or 4-week WD-fed. Data presented as mean ± SD from n = 10 independent experiments. Statistical significance was calculated using a two-way ANOVA and Tukey’s test to correct for multiple comparisons. Representative Western blots are shown. DNL: de novo lipid synthesis; PV: portal vein; CV: central vein; WT: wild type. CNTR: control; WD: western diet.

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Expressing, Control, Western Blot

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: (A) Representative confocal images of liver sections from mtDendra2 mice fed WD for 4, 8, and 12 weeks. Mitochondria (green), LDs (magenta), and actin (yellow). Scale bar 5 μm. (B) Heat map of mitochondria and LDs overlapping pixels across the PP-PC axis (R1-R12) and prolonged WD feeding. (C) Relative Plin5 mRNA expression in mice fed WD for 4,8, and 12 weeks. (D) Relative PLIN5 mRNA expression in mice fed a control (CNTR) diet or Western Diet (WD) for 4, 8, and 12 weeks. Data presented as mean ± SD from n = 3 independent experiments. Statistical significance was calculated using a two-way ANOVA and Tukey’s test to correct for multiple comparisons. PV: portal vein; CV: central vein. WT: wild type.

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Expressing, Control, Western Blot

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: a , Histopathology was used to group 12 healthy samples on the basis of lipid content. Oil red O staining of a representative individual from each group. Scale bar, 500 μm. b , Immunofluorescence staining of mitochondria (green), LDs (magenta), actin (yellow) and GS (white) in a liver section from a representative individual with minimal or mild steatosis. Scale bar, 100 μm. c , Volume rendering of mitochondria (green), LDs (magenta) and actin (yellow) in a representative PP and PC hepatocytes from patients with minimal or mild steatosis. Colocalizing pixels are shown in white. d – f , Quantification of LD count ( d ), LD volume ( e ) and mitochondria–LD colocalization ( f ), in n = 16 cells from 2 people with minimal steatosis and n = 12 cells from 3 people with mild steatosis, presented as mean ± s.d. Statistical significance was calculated using a one-way ANOVA. g , Plin5 expression was assessed in liver samples from seven people without steatosis, two with minimal steatosis and three with mild steatosis. Data presented as mean ± s.d. Statistical significance was calculated using a one-way ANOVA. Statistical significance was calculated using a t -test.

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Histopathology, Staining, Immunofluorescence, Expressing

Journal: Nature Metabolism

Article Title: PLIN5 phosphorylation orchestrates mitochondria lipid-droplet coupling to control hepatic lipid flux and steatosis

doi: 10.1038/s42255-026-01476-1

Figure Lengend Snippet: Proposed model of mitochondria–LD contacts regulation by PLIN5. PLIN5 induces mitochondria–LD contacts in a phosphorylation-dependent manner. PLIN5-S155A induces contact sites promoting the esterification of FFA to form TG and storage in LDs, which is protective from oxidative stress owing to increased dietary lipids. In PLIN5-S155E, mitochondria and LDs are rarely associated; LDs are smaller, and antioxidant capacity is reduced. In long-term WD feeding, the abundance of mitochondria–LD contacts increases, suggesting that the assembly of these contacts might be part of an adaptive response. Created in BioRender; Porat-Shliom, N. https://BioRender.com/jnbti4d (2026).

Article Snippet: Singleplex reactions (5 μl) containing a FAM-MGB expression assay for Plin5 (Mm00508854_m1) or Tbp control (Mm01277041_m1) (Thermo Fisher) were performed using cDNA synthesized from 8 ng RNA and 1× Fast Advanced Master Mix (Thermo-without Amp Erase UNG) were performed in triplicate.

Techniques: Phospho-proteomics

Journal: Scientific Reports

Article Title: Perilipin 5 fine-tunes lipid oxidation to metabolic demand and protects against lipotoxicity in skeletal muscle

doi: 10.1038/srep38310

Figure Lengend Snippet: Representative blots ( A ) and quantification of PLIN5 ( B ) and ATGL ( C ) protein content in different mouse skeletal muscles (n = 5) (EDL: extensor digitorum longus , TA: tibialis anterior , Sol: soleus ). **p < 0.01, ***p < 0.001 versus EDL. ( D ) Quantification of PLIN5 protein content in vastus lateralis muscle of healthy lean and endurance-trained volunteers (n = 11 per group). Correlations between muscle PLIN5 protein and ( E ) cytochrome oxidase activity, and ( F ) glucose disposal rate (n = 33). *p < 0.05 versus lean.

Article Snippet: For overexpression experiments, adenoviruses expressing in tandem GFP and human PLIN5 (hPLIN5) were used (Vector Biolabs, Philadelphia, PA).

Techniques: Muscles, Activity Assay

Journal: Scientific Reports

Article Title: Perilipin 5 fine-tunes lipid oxidation to metabolic demand and protects against lipotoxicity in skeletal muscle

doi: 10.1038/srep38310

Figure Lengend Snippet: ( A ) Representative blot and quantification of PLIN5 protein content in control (Ad-GFP) and PLIN5-overexpressing myotubes (Ad-PLIN5) (n = 3). Pulse-Chase studies using [1- 14 C] oleate were performed to determine ( B ) FA release into the culture medium (Ad-GFP = 59 ± 1.8 nmol/3 h/mg protein), ( C ) FA oxidation (Ad-GFP = 2.22 ± 0.13 nmol/3 h/mg protein), and the rate of incorporation of radiolabeled oleate into ( D ) TAG, ( E ) DAG (T0 Ad-GFP = 3.14 ± 0.14 nmol/3 h/mg protein) and ( F ) intracellular FA content (T0 Ad-GFP = 0.42 ± 0.03 nmol/3 h/mg protein) in control (Ad-GFP) and PLIN5-overexpressing myotubes (Ad-PLIN5). ( G ) Glycogen synthesis and ( H ) glucose oxidation were measured in control myotubes (Ad-GFP) and myotubes overexpressing PLIN5 (Ad-PLIN5) using [U- 14 C] glucose. ( I ) PDK4 gene expression was measured in control (Ad-GFP) and PLIN5-overexpressing myotubes (Ad-PLIN5). (n = 6) *p < 0.05, **p < 0.01 ***p < 0.001 versus Ad-GFP.

Article Snippet: For overexpression experiments, adenoviruses expressing in tandem GFP and human PLIN5 (hPLIN5) were used (Vector Biolabs, Philadelphia, PA).

Techniques: Control, Pulse Chase, Gene Expression

Journal: Scientific Reports

Article Title: Perilipin 5 fine-tunes lipid oxidation to metabolic demand and protects against lipotoxicity in skeletal muscle

doi: 10.1038/srep38310

Figure Lengend Snippet: Pulse-Chase studies using [1- 14 C] oleate were performed to determine the rate of ( A,D ) incorporation of radiolabeled oleate into TAG and ( B,E ) oleate oxidation in control myotubes (Ad-GFP) and myotubes overexpressing PLIN5 (Ad-PLIN5) either during ( A–C ) forskolin (FK) (Ad-GFP CONT = 1.66 ± 0.25 nmol/3 h/mg protein) or ( D–E ) electrical pulse (EPS) stimulation (Ad-GFP CONT = 6.33 ± 2.05 nmol/24 h/mg protein) (n = 6). Values are expressed in % of Ad-GFP Control ( A,B,D,E ) and in fold change over control in ( C and F ). *p < 0.05, **p < 0.01 ***p < 0.001 versus Ad-GFP.

Article Snippet: For overexpression experiments, adenoviruses expressing in tandem GFP and human PLIN5 (hPLIN5) were used (Vector Biolabs, Philadelphia, PA).

Techniques: Pulse Chase, Control

Journal: Scientific Reports

Article Title: Perilipin 5 fine-tunes lipid oxidation to metabolic demand and protects against lipotoxicity in skeletal muscle

doi: 10.1038/srep38310

Figure Lengend Snippet: ( A ) Glycogen synthesis was measured in control myotubes (Ad-GFP) and myotubes overexpressing PLIN5 (Ad-PLIN5) using [U- 14 C] glucose in absence or presence of 100 nM insulin, in control cells and in cells treated with 300 μM of palmitic acid for 24 h (n = 9). ( B ) Total diacylglycerols (DAG) and ( C ) Ceramide (CER) d18:1/16:0 content were measured in control myotubes (Ad-GFP) and myotubes overexpressing PLIN5 (Ad-PLIN5) (n = 4). *p < 0.05, **p < 0.01 versus Ad-GFP.

Article Snippet: For overexpression experiments, adenoviruses expressing in tandem GFP and human PLIN5 (hPLIN5) were used (Vector Biolabs, Philadelphia, PA).

Techniques: Control

Journal: Scientific Reports

Article Title: Perilipin 5 fine-tunes lipid oxidation to metabolic demand and protects against lipotoxicity in skeletal muscle

doi: 10.1038/srep38310

Figure Lengend Snippet: PLIN5 ( A ) gene expression and ( B ) protein content measured in control (shNT) and PLIN5 silenced (shPLIN5) mouse tibialis anterior muscle (n = 6). Palmitate ( C ) and glucose ( D ) oxidation rate were measured using respectively [U- 14 C] glucose or [1- 14 C] palmitate in control (shNT) and PLIN5 silenced (shPLIN5) muscle homogenates. Palmitate oxidation (i.e. CO2), acid soluble metabolites accumulation (i.e. ASMs) and total oxidation (i.e. the sum of CO2 release and ASMs accumulation) were measured (n = 6). ( E ) Insulin-stimulated glucose uptake was determined in control (shNT) and PLIN5 knockdown (shPLIN5) muscles. (n = 7). *p < 0.05, ***p < 0.001 versus shNT.

Article Snippet: For overexpression experiments, adenoviruses expressing in tandem GFP and human PLIN5 (hPLIN5) were used (Vector Biolabs, Philadelphia, PA).

Techniques: Gene Expression, Control, Knockdown, Muscles

Journal: Scientific Reports

Article Title: Perilipin 5 fine-tunes lipid oxidation to metabolic demand and protects against lipotoxicity in skeletal muscle

doi: 10.1038/srep38310

Figure Lengend Snippet: PLIN5 ( A ) gene expression and ( B ) protein content measured in control (shNT) and PLIN5 silenced (shPLIN5) mouse tibialis anterior muscle (n = 6). ( C ) Insulin-stimulated glucose uptake, ( D ) total ceramide (CER) and ( E ) total diacylglycerols (DAG) content were determined in control (shNT) and PLIN5 knockdown (shPLIN5) muscles. (n = 7). ( F ) Insulin-stimulated Akt phosphorylation on Ser473 and Thr308 residues was measured in control (shNT) and PLIN5 silenced (shPLIN5) muscle (n = 4). *p < 0.05, **p < 0.01, ***p < 0.001 versus shNT.

Article Snippet: For overexpression experiments, adenoviruses expressing in tandem GFP and human PLIN5 (hPLIN5) were used (Vector Biolabs, Philadelphia, PA).

Techniques: Gene Expression, Control, Knockdown, Muscles, Phospho-proteomics

Journal: Scientific Reports

Article Title: Perilipin 5 fine-tunes lipid oxidation to metabolic demand and protects against lipotoxicity in skeletal muscle

doi: 10.1038/srep38310

Figure Lengend Snippet: In the resting state, PLIN5 protects LD from lipolytic attack by lipases. An increase in PLIN5 content (red arrows) slows down lipolysis and FA oxidation, favoring a switch towards glucose utilization. During lipolytic stimulation (i.e. PKA activation or contraction), PLIN5 enhances FA oxidation, thereby increasing CO 2 production. It has been suggested that PLIN5 could provide a physical linkage between LD and mitochondria. We can hypothesize that this relocation has metabolic consequences by facilitating FA channeling from LD to mitochondria, thus allowing a more efficient coupling between IMTG lipolysis and FA oxidation upon increased metabolic demand. Finally, the up-regulation of PLIN5 with high-fat feeding is insufficient to protect from LD-mediated CER accumulation. FA: Fatty Acids; IMTG: Intramyocellular Triacylglycerols; CER: Ceramides.

Article Snippet: For overexpression experiments, adenoviruses expressing in tandem GFP and human PLIN5 (hPLIN5) were used (Vector Biolabs, Philadelphia, PA).

Techniques: Activation Assay