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Fatty acids from membrane lipids become incorporated into lipid bodies during Myxococcus xanthus differentiation.

Bhat S, Boynton TO, Pham D, Shimkets LJ - PLoS ONE (2014)

Bottom Line: Third, peripheral rods, developing cells that do not produce lipid bodies, fail to shorten.The lipid body regulon involves many developmental genes that are not specifically involved in fatty acid synthesis or degradation.MazF RNA interferase and its target, enhancer-binding protein Nla6, appear to negatively regulate cell shortening and TAG accumulation whereas most cell-cell signals activate these processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Georgia, Athens, Georgia, United States of America.

ABSTRACT
Myxococcus xanthus responds to amino acid limitation by producing fruiting bodies containing dormant spores. During development, cells produce triacylglycerides in lipid bodies that become consumed during spore maturation. As the cells are starved to induce development, the production of triglycerides represents a counterintuitive metabolic switch. In this paper, lipid bodies were quantified in wild-type strain DK1622 and 33 developmental mutants at the cellular level by measuring the cross sectional area of the cell stained with the lipophilic dye Nile red. We provide five lines of evidence that triacylglycerides are derived from membrane phospholipids as cells shorten in length and then differentiate into myxospores. First, in wild type cells, lipid bodies appear early in development and their size increases concurrent with an 87% decline in membrane surface area. Second, developmental mutants blocked at different stages of shortening and differentiation accumulated lipid bodies proportionate with their cell length with a Pearson's correlation coefficient of 0.76. Third, peripheral rods, developing cells that do not produce lipid bodies, fail to shorten. Fourth, genes for fatty acid synthesis are down-regulated while genes for fatty acid degradation are up regulated. Finally, direct movement of fatty acids from membrane lipids in growing cells to lipid bodies in developing cells was observed by pulse labeling cells with palmitate. Recycling of lipids released by Programmed Cell Death appears not to be necessary for lipid body production as a fadL mutant was defective in fatty acid uptake but proficient in lipid body production. The lipid body regulon involves many developmental genes that are not specifically involved in fatty acid synthesis or degradation. MazF RNA interferase and its target, enhancer-binding protein Nla6, appear to negatively regulate cell shortening and TAG accumulation whereas most cell-cell signals activate these processes.

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Lipid bodies are derived from membrane phospholipids.Myxococcus cells were grown in the presence of palmitic acid alkyne (PAalk) in A1 minimal medium and stained with Nile red. Click chemistry was used to attach Alexa Fluor 488 azide to PAalk prior to visualization. Both Alex Fluor fatty acid and the Nile red stain are visible in membranes during vegetative growth (DK1622 0 h). The PAalk was removed and cells were allowed to develop on TPM agar (DK1622 24 h). Labeled membrane lipids were incorporated into Nile red stained lipid bodies, seen in both channels and the merged images. In a strain deficient in lipid body production (LS3931), incorporation occurs into membrane lipids (LS3931 0 h) but not lipid bodies (LS3931 24 h). A fadL mutant (LS3125), defective in fatty acid uptake, is unable to incorporate PAalk altogether. Wild-type cells grown in the absence of PAalk then allowed to develop (DK1622a 24 h) exhibit no fluorescence (bottom row, second panel) indicating little bleed through between channels. Scale bar is 10 µm.
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pone-0099622-g004: Lipid bodies are derived from membrane phospholipids.Myxococcus cells were grown in the presence of palmitic acid alkyne (PAalk) in A1 minimal medium and stained with Nile red. Click chemistry was used to attach Alexa Fluor 488 azide to PAalk prior to visualization. Both Alex Fluor fatty acid and the Nile red stain are visible in membranes during vegetative growth (DK1622 0 h). The PAalk was removed and cells were allowed to develop on TPM agar (DK1622 24 h). Labeled membrane lipids were incorporated into Nile red stained lipid bodies, seen in both channels and the merged images. In a strain deficient in lipid body production (LS3931), incorporation occurs into membrane lipids (LS3931 0 h) but not lipid bodies (LS3931 24 h). A fadL mutant (LS3125), defective in fatty acid uptake, is unable to incorporate PAalk altogether. Wild-type cells grown in the absence of PAalk then allowed to develop (DK1622a 24 h) exhibit no fluorescence (bottom row, second panel) indicating little bleed through between channels. Scale bar is 10 µm.

Mentions: In wild-type cells (Figure 4, DK1622), PAalk is detected in the membranes of vegetative cells grown in A1 (0 h) as a thin layer of fluorescence that outlines each cell. The chemical form(s) was not identified, but a likely possibility is phosphatidylethanolamine, which comprises 70% of the phospholipid in Myxococcus[36]. By 24 h of development, PAalk becomes prominent within DK1622 lipid bodies demonstrating that the TAGs are at least partially derived from the fatty acids in membrane lipids. PAalk is also incorporated into membranes of a strain deficient in making lipid bodies (Figure 4, LS3931), but fully formed lipid bodies are rarely observed. No detectable signal is seen in lipid bodies from wild-type cells grown without PAalk fatty acid indicating that there is no bleed through between fluorescence channels (Figure 4, last line).


Fatty acids from membrane lipids become incorporated into lipid bodies during Myxococcus xanthus differentiation.

Bhat S, Boynton TO, Pham D, Shimkets LJ - PLoS ONE (2014)

Lipid bodies are derived from membrane phospholipids.Myxococcus cells were grown in the presence of palmitic acid alkyne (PAalk) in A1 minimal medium and stained with Nile red. Click chemistry was used to attach Alexa Fluor 488 azide to PAalk prior to visualization. Both Alex Fluor fatty acid and the Nile red stain are visible in membranes during vegetative growth (DK1622 0 h). The PAalk was removed and cells were allowed to develop on TPM agar (DK1622 24 h). Labeled membrane lipids were incorporated into Nile red stained lipid bodies, seen in both channels and the merged images. In a strain deficient in lipid body production (LS3931), incorporation occurs into membrane lipids (LS3931 0 h) but not lipid bodies (LS3931 24 h). A fadL mutant (LS3125), defective in fatty acid uptake, is unable to incorporate PAalk altogether. Wild-type cells grown in the absence of PAalk then allowed to develop (DK1622a 24 h) exhibit no fluorescence (bottom row, second panel) indicating little bleed through between channels. Scale bar is 10 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4048283&req=5

pone-0099622-g004: Lipid bodies are derived from membrane phospholipids.Myxococcus cells were grown in the presence of palmitic acid alkyne (PAalk) in A1 minimal medium and stained with Nile red. Click chemistry was used to attach Alexa Fluor 488 azide to PAalk prior to visualization. Both Alex Fluor fatty acid and the Nile red stain are visible in membranes during vegetative growth (DK1622 0 h). The PAalk was removed and cells were allowed to develop on TPM agar (DK1622 24 h). Labeled membrane lipids were incorporated into Nile red stained lipid bodies, seen in both channels and the merged images. In a strain deficient in lipid body production (LS3931), incorporation occurs into membrane lipids (LS3931 0 h) but not lipid bodies (LS3931 24 h). A fadL mutant (LS3125), defective in fatty acid uptake, is unable to incorporate PAalk altogether. Wild-type cells grown in the absence of PAalk then allowed to develop (DK1622a 24 h) exhibit no fluorescence (bottom row, second panel) indicating little bleed through between channels. Scale bar is 10 µm.
Mentions: In wild-type cells (Figure 4, DK1622), PAalk is detected in the membranes of vegetative cells grown in A1 (0 h) as a thin layer of fluorescence that outlines each cell. The chemical form(s) was not identified, but a likely possibility is phosphatidylethanolamine, which comprises 70% of the phospholipid in Myxococcus[36]. By 24 h of development, PAalk becomes prominent within DK1622 lipid bodies demonstrating that the TAGs are at least partially derived from the fatty acids in membrane lipids. PAalk is also incorporated into membranes of a strain deficient in making lipid bodies (Figure 4, LS3931), but fully formed lipid bodies are rarely observed. No detectable signal is seen in lipid bodies from wild-type cells grown without PAalk fatty acid indicating that there is no bleed through between fluorescence channels (Figure 4, last line).

Bottom Line: Third, peripheral rods, developing cells that do not produce lipid bodies, fail to shorten.The lipid body regulon involves many developmental genes that are not specifically involved in fatty acid synthesis or degradation.MazF RNA interferase and its target, enhancer-binding protein Nla6, appear to negatively regulate cell shortening and TAG accumulation whereas most cell-cell signals activate these processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Georgia, Athens, Georgia, United States of America.

ABSTRACT
Myxococcus xanthus responds to amino acid limitation by producing fruiting bodies containing dormant spores. During development, cells produce triacylglycerides in lipid bodies that become consumed during spore maturation. As the cells are starved to induce development, the production of triglycerides represents a counterintuitive metabolic switch. In this paper, lipid bodies were quantified in wild-type strain DK1622 and 33 developmental mutants at the cellular level by measuring the cross sectional area of the cell stained with the lipophilic dye Nile red. We provide five lines of evidence that triacylglycerides are derived from membrane phospholipids as cells shorten in length and then differentiate into myxospores. First, in wild type cells, lipid bodies appear early in development and their size increases concurrent with an 87% decline in membrane surface area. Second, developmental mutants blocked at different stages of shortening and differentiation accumulated lipid bodies proportionate with their cell length with a Pearson's correlation coefficient of 0.76. Third, peripheral rods, developing cells that do not produce lipid bodies, fail to shorten. Fourth, genes for fatty acid synthesis are down-regulated while genes for fatty acid degradation are up regulated. Finally, direct movement of fatty acids from membrane lipids in growing cells to lipid bodies in developing cells was observed by pulse labeling cells with palmitate. Recycling of lipids released by Programmed Cell Death appears not to be necessary for lipid body production as a fadL mutant was defective in fatty acid uptake but proficient in lipid body production. The lipid body regulon involves many developmental genes that are not specifically involved in fatty acid synthesis or degradation. MazF RNA interferase and its target, enhancer-binding protein Nla6, appear to negatively regulate cell shortening and TAG accumulation whereas most cell-cell signals activate these processes.

Show MeSH
Related in: MedlinePlus