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The Causative Gene in Chanarian Dorfman Syndrome Regulates Lipid Droplet Homeostasis in C. elegans.

Xie M, Roy R - PLoS Genet. (2015)

Bottom Line: We found that the compromise of one of the three C. elegans orthologues of human cgi-58 significantly improves the survival of AMPK-deficient dauers.We also provide evidence that C. elegans CGI-58 acts as a co-activator of ATGL-1, while it also functions cooperatively to maintain regular lipid droplet structure.Surprisingly, we show that it also acts independently of ATGL-1 to restrict lipid droplet coalescence by altering the surface abundance and composition of long chain (C20) polyunsaturated fatty acids (PUFAs).

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, McGill University, Penfield, Montreal, Quebec, Canada.

ABSTRACT
AMP-activated kinase (AMPK) is a key regulator of many cellular mechanisms required for adjustment to various stresses induced by the changing environment. In C. elegans dauer larvae AMPK- mutants expire prematurely due to hyperactive Adipose Triglyceride Lipase (ATGL-1) followed by rapid depletion of triglyceride stores. We found that the compromise of one of the three C. elegans orthologues of human cgi-58 significantly improves the survival of AMPK-deficient dauers. We also provide evidence that C. elegans CGI-58 acts as a co-activator of ATGL-1, while it also functions cooperatively to maintain regular lipid droplet structure. Surprisingly, we show that it also acts independently of ATGL-1 to restrict lipid droplet coalescence by altering the surface abundance and composition of long chain (C20) polyunsaturated fatty acids (PUFAs). Our data reveal a novel structural role of CGI-58 in maintaining lipid droplet homeostasis through its effects on droplet composition, morphology and lipid hydrolysis; a conserved function that may account for some of the ATGL-1-independent features unique to Chanarin-Dorfman Syndrome.

No MeSH data available.


Related in: MedlinePlus

CGI-58 regulates lipid droplet fusion.(A)cgi-58 restricts fusion events among lipid droplets in both control and AMPK mutant dauer larvae. Red C1-BODIPY-C12 labeled lipid droplets were imaged in real time (15 min) in control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Scale bar = 6 μm. All strains carry daf-2(e1370) in (A)-(D). (B)cgi-58, but not atgl-1 limits the exchange of lipid content among lipid droplets isolated from control, CGI-58, AMPK and AMPK; CGI-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Lipid exchange was determined between isolated Green and Red C1-BODIPY-C12 labeled lipid droplets mixed in vitro for 30 minutes, following which fluorescence of individual droplets was analysed. Longer incubation times were performed and similar results were obtained. The abundance of lipid droplets/ml reaction volume was adjusted to similar levels prior to quantification. LD1 and LD2 represent two individual differentially labelled lipid droplets. The line graph at the right side of each set of images represents the fluorescent intensity of Green and Red C1-BODIPY-C12 labeled lipid droplets along the white arrow. Scale bar = 2 μm. (C) Quantification of the lipid droplets containing both green and red labelled lipid contents (both red and green fluorescent intensities are greater than 100) derived from the droplets described in (B) determined for 1000 lipid droplets analyzed for each respective genotype. ** indicates statistical significance (P<0.01). (D) Lipid content was more rapidly replenished in droplets obtained from animals that lack CGI-58. The dotted line in the graphs of the second (cgi-58) and fourth (aak(0); cgi-58) panel represent the maximum recovered intensity observed in the graph of the first (control daf-2) and third (aak(0)) panel. FRAP analysis was performed on control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 dauer day 0 animals. Raw imaging data were captured and processed using Metamorph image acquisition software. Scale bar = 4 μm.
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pgen.1005284.g005: CGI-58 regulates lipid droplet fusion.(A)cgi-58 restricts fusion events among lipid droplets in both control and AMPK mutant dauer larvae. Red C1-BODIPY-C12 labeled lipid droplets were imaged in real time (15 min) in control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Scale bar = 6 μm. All strains carry daf-2(e1370) in (A)-(D). (B)cgi-58, but not atgl-1 limits the exchange of lipid content among lipid droplets isolated from control, CGI-58, AMPK and AMPK; CGI-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Lipid exchange was determined between isolated Green and Red C1-BODIPY-C12 labeled lipid droplets mixed in vitro for 30 minutes, following which fluorescence of individual droplets was analysed. Longer incubation times were performed and similar results were obtained. The abundance of lipid droplets/ml reaction volume was adjusted to similar levels prior to quantification. LD1 and LD2 represent two individual differentially labelled lipid droplets. The line graph at the right side of each set of images represents the fluorescent intensity of Green and Red C1-BODIPY-C12 labeled lipid droplets along the white arrow. Scale bar = 2 μm. (C) Quantification of the lipid droplets containing both green and red labelled lipid contents (both red and green fluorescent intensities are greater than 100) derived from the droplets described in (B) determined for 1000 lipid droplets analyzed for each respective genotype. ** indicates statistical significance (P<0.01). (D) Lipid content was more rapidly replenished in droplets obtained from animals that lack CGI-58. The dotted line in the graphs of the second (cgi-58) and fourth (aak(0); cgi-58) panel represent the maximum recovered intensity observed in the graph of the first (control daf-2) and third (aak(0)) panel. FRAP analysis was performed on control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 dauer day 0 animals. Raw imaging data were captured and processed using Metamorph image acquisition software. Scale bar = 4 μm.

Mentions: It is possible that the increased frequency of lipid droplet encounters might only occur in isolated droplets and have little relevance to droplet behavior in vivo. To address this we monitored lipid droplet dynamics in live dauer larvae in real time. By imaging live animals over a period of 15 min we observed an enhanced frequency of encounters that occurred among individual lipid droplets in CGI-58-deficient control daf-2 and AMPK mutant animals (Fig 5A and S1–S4 Movies).


The Causative Gene in Chanarian Dorfman Syndrome Regulates Lipid Droplet Homeostasis in C. elegans.

Xie M, Roy R - PLoS Genet. (2015)

CGI-58 regulates lipid droplet fusion.(A)cgi-58 restricts fusion events among lipid droplets in both control and AMPK mutant dauer larvae. Red C1-BODIPY-C12 labeled lipid droplets were imaged in real time (15 min) in control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Scale bar = 6 μm. All strains carry daf-2(e1370) in (A)-(D). (B)cgi-58, but not atgl-1 limits the exchange of lipid content among lipid droplets isolated from control, CGI-58, AMPK and AMPK; CGI-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Lipid exchange was determined between isolated Green and Red C1-BODIPY-C12 labeled lipid droplets mixed in vitro for 30 minutes, following which fluorescence of individual droplets was analysed. Longer incubation times were performed and similar results were obtained. The abundance of lipid droplets/ml reaction volume was adjusted to similar levels prior to quantification. LD1 and LD2 represent two individual differentially labelled lipid droplets. The line graph at the right side of each set of images represents the fluorescent intensity of Green and Red C1-BODIPY-C12 labeled lipid droplets along the white arrow. Scale bar = 2 μm. (C) Quantification of the lipid droplets containing both green and red labelled lipid contents (both red and green fluorescent intensities are greater than 100) derived from the droplets described in (B) determined for 1000 lipid droplets analyzed for each respective genotype. ** indicates statistical significance (P<0.01). (D) Lipid content was more rapidly replenished in droplets obtained from animals that lack CGI-58. The dotted line in the graphs of the second (cgi-58) and fourth (aak(0); cgi-58) panel represent the maximum recovered intensity observed in the graph of the first (control daf-2) and third (aak(0)) panel. FRAP analysis was performed on control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 dauer day 0 animals. Raw imaging data were captured and processed using Metamorph image acquisition software. Scale bar = 4 μm.
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Related In: Results  -  Collection

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Show All Figures
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pgen.1005284.g005: CGI-58 regulates lipid droplet fusion.(A)cgi-58 restricts fusion events among lipid droplets in both control and AMPK mutant dauer larvae. Red C1-BODIPY-C12 labeled lipid droplets were imaged in real time (15 min) in control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Scale bar = 6 μm. All strains carry daf-2(e1370) in (A)-(D). (B)cgi-58, but not atgl-1 limits the exchange of lipid content among lipid droplets isolated from control, CGI-58, AMPK and AMPK; CGI-58 mutant dauer larvae 32 hours after shifting to restrictive temperature. Lipid exchange was determined between isolated Green and Red C1-BODIPY-C12 labeled lipid droplets mixed in vitro for 30 minutes, following which fluorescence of individual droplets was analysed. Longer incubation times were performed and similar results were obtained. The abundance of lipid droplets/ml reaction volume was adjusted to similar levels prior to quantification. LD1 and LD2 represent two individual differentially labelled lipid droplets. The line graph at the right side of each set of images represents the fluorescent intensity of Green and Red C1-BODIPY-C12 labeled lipid droplets along the white arrow. Scale bar = 2 μm. (C) Quantification of the lipid droplets containing both green and red labelled lipid contents (both red and green fluorescent intensities are greater than 100) derived from the droplets described in (B) determined for 1000 lipid droplets analyzed for each respective genotype. ** indicates statistical significance (P<0.01). (D) Lipid content was more rapidly replenished in droplets obtained from animals that lack CGI-58. The dotted line in the graphs of the second (cgi-58) and fourth (aak(0); cgi-58) panel represent the maximum recovered intensity observed in the graph of the first (control daf-2) and third (aak(0)) panel. FRAP analysis was performed on control daf-2, daf-2; cgi-58, daf-2; aak(0) or daf-2; aak(0); cgi-58 dauer day 0 animals. Raw imaging data were captured and processed using Metamorph image acquisition software. Scale bar = 4 μm.
Mentions: It is possible that the increased frequency of lipid droplet encounters might only occur in isolated droplets and have little relevance to droplet behavior in vivo. To address this we monitored lipid droplet dynamics in live dauer larvae in real time. By imaging live animals over a period of 15 min we observed an enhanced frequency of encounters that occurred among individual lipid droplets in CGI-58-deficient control daf-2 and AMPK mutant animals (Fig 5A and S1–S4 Movies).

Bottom Line: We found that the compromise of one of the three C. elegans orthologues of human cgi-58 significantly improves the survival of AMPK-deficient dauers.We also provide evidence that C. elegans CGI-58 acts as a co-activator of ATGL-1, while it also functions cooperatively to maintain regular lipid droplet structure.Surprisingly, we show that it also acts independently of ATGL-1 to restrict lipid droplet coalescence by altering the surface abundance and composition of long chain (C20) polyunsaturated fatty acids (PUFAs).

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, McGill University, Penfield, Montreal, Quebec, Canada.

ABSTRACT
AMP-activated kinase (AMPK) is a key regulator of many cellular mechanisms required for adjustment to various stresses induced by the changing environment. In C. elegans dauer larvae AMPK- mutants expire prematurely due to hyperactive Adipose Triglyceride Lipase (ATGL-1) followed by rapid depletion of triglyceride stores. We found that the compromise of one of the three C. elegans orthologues of human cgi-58 significantly improves the survival of AMPK-deficient dauers. We also provide evidence that C. elegans CGI-58 acts as a co-activator of ATGL-1, while it also functions cooperatively to maintain regular lipid droplet structure. Surprisingly, we show that it also acts independently of ATGL-1 to restrict lipid droplet coalescence by altering the surface abundance and composition of long chain (C20) polyunsaturated fatty acids (PUFAs). Our data reveal a novel structural role of CGI-58 in maintaining lipid droplet homeostasis through its effects on droplet composition, morphology and lipid hydrolysis; a conserved function that may account for some of the ATGL-1-independent features unique to Chanarin-Dorfman Syndrome.

No MeSH data available.


Related in: MedlinePlus