Fat or lean? Atlastin links ER to lipid storage

Research Areas:

Cell biology, Endoplasmic reticulum, Lipid droplets, Fluorescence microscopy

Imaging Needs:

Localization of sub-micron features with green and red fluorescence

Imaging System (live C. elegans imaging):
  • GFP: DGAT-2 to label lipid droplets
  • mRuby: ATLN-1 to label alastin
  • mCherry: HDEL to label the ER
  • PerkinElmer Ultraview spinning disk confocal microscope
  • Plan-Apochromat 100×, NA 1.45 oil objective
  • GFP: 488 nm laser excitation, 500–555 nm emission filter
  • mRuby, mCherry: 561 nm laser excitation, 415–475 and 580–650 nm dual-pass emission filter
  • Autofluorescence: 488 nm laser excitation, 415–475 and 580–650 nm dual-pass emission filter
  • Hamamatsu ORCA-R2 CCD camera
  • PerkinElmer Volocity and Bitplane Imaris software
Imaging System (cellular immunofluorescence):
  • Nikon TE2000U inverted microscope
  • Yokogawa spinning-disk confocal
  • 100× Plan Apo NA 1.4 objective lens
  • Hamamatsu ORCA-ER CCD camera
  • MetaMorph software
In Vivo–In Vitro Correlations

What if the same quantitative in vitro cell assay could also be used in vivo? Takakura, et al., detect cellular events at both levels using firefly luciferase bioluminescence and the Hamamatsu ImagEM-1K EM CCD camera. Read now.

THE QUESTION

What determines the size of lipid droplets for fat storage?

Eukaryotes store fats as lipid droplets within cells—membrane-bound capsules of triglycerides and sterol esters. The size of these droplets correlates with nutrient availability. Researchers have discovered enzymes responsible for synthesizing and breaking down the stored lipids, but it remains unclear just how the process is regulated.

THE BARRIERS

Within cells, lipid droplets are closely associated with the endoplasmic reticulum (ER). Electron microscopy indicates the two must be coupled together for a lipid droplet to grow larger. How does the ER influence droplet enlargement?

THE SOLUTION

A Conserved Role for Atlastin GTPases in Regulating Lipid Droplet Size
Klemm RW, Norton JP, Cole RA, Li CS, Park SH, Crane MM, Li L, Jin D, Boye-Doe A, Liu TY, Shibata Y, Lu H, Rapoport TA, Farese RV Jr, Blackstone C, Guo Y, Mak HY
Cell Rep. 2013 May 30; 3(5): 1465–1475. PMCID: PMC3742324.

Klemm, et al,1 screened for genes affecting the shape of lipid droplets in the intestinal cells of C. elegans, with the help of Hamamatsu’s ORCA-R2 and ORCA-ER CCD cameras. Mutations in the gene for atlastin, a member of the dynamin family of GTPases that is important for ER structure, caused reduced lipid droplet size as well as defects in ER morphology. Experiments in Drosophila, C. elegans, and mammalian cells agreed—atlastin mutation or depletion by RNA interference decreased lipid droplet size and overall triglyceride levels, while atlastin overexpression led to giant lipid droplets.

Localization of fluorescently labeled ER and lipid droplets showed an altered distribution of droplets in atlastin-depleted cells coincident with a contracted, distorted ER. The authors conclude that lipid droplet size depends on atlastin-mediated fusion in the ER membrane, by a mechanism that is highly conserved among species.

THE POSSIBILITIES

Klemm, et al. characterized sub-micron lipid droplet and ER morphology in living C. elegans larvae and cultured cells using fluorescence imaging with the ORCA-R2 and ORCA-ER CCD cameras. Find out another way researchers are using Hamamatsu cameras to visualize events within intact, living organisms—read Illuminating Activity—in vitro and in vivo.

References

  1. Klemm, et al. A Conserved Role for Atlastin GTPases in Regulating Lipid Droplet Size. Cell Rep. 2013 May 30; 3(5): 1465–1475. PMCID: PMC3742324.
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