XB-ART-47936Nat Cell Biol October 1, 2013; 15 (10): 1253-9.
A nuclear F-actin scaffold stabilizes ribonucleoprotein droplets against gravity in large cells.
The size of a typical eukaryotic cell is of the order of ∼10 μm. However, some cell types grow to very large sizes, including oocytes (immature eggs) of organisms from humans to starfish. For example, oocytes of the frog Xenopus laevis grow to a diameter ≥1 mm. They have a correspondingly large nucleus (germinal vesicle) of ∼450 μm in diameter, which is similar to smaller somatic nuclei, but contains a significantly higher concentration of actin. The form and structure of this nuclear actin remain controversial, and its potential mechanical role within these large nuclei is unknown. Here, we use a microrheology and quantitative imaging approach to show that germinal vesicles contain an elastic F-actin scaffold that mechanically stabilizes these large nuclei against gravitational forces, which are usually considered negligible within cells. We find that on actin disruption, ribonucleoprotein droplets, including nucleoli and histone locus bodies, undergo gravitational sedimentation and fusion. We develop a model that reveals how gravity becomes an increasingly potent force as cells and their nuclei grow larger than ∼10 μm, explaining the requirement for a stabilizing nuclear F-actin scaffold in large Xenopus oocytes. All life forms are subject to gravity, and our results may have broad implications for cell growth and size control.
PubMed ID: 23995731
PMC ID: PMC3789854
Article link: Nat Cell Biol
Genes referenced: actl6a actn1 coil fscn1 grap2 igf2bp3 lmnb3 npm1 xpo6
Article Images: [+] show captions
|Figure 3. Mechanics, anchoring and structural regulation of nuclear actina, Compressive forces were applied to the GV using a microneedle, demonstrating a coupled elastic response of the actin meshwork (Lifeact::GFP, green) and lamin cortex (RFP::Lamin B3, red). Arrowhead shows increased intensity, suggesting actin polymerization occurs in response to force. b, Tensile forces were applied to the GV using a microneedle, showing a similar coupled elastic response. c, Two nuclear bodies trapped in a dense native actin meshwork. d, Tropomyosin injection leads to a compacted actin meshwork, and the nuclear bodies are often deformed. e, Fascin injection leads to bundling of the actin meshwork. For c–e, nucleoli are labeled with NPM1::RFP (red), and actin is labeled with Lifeact::GFP (green). d, Bar graph showing the MSD of large (R=1 μm) beads at a lag time of 5 sec, under various conditions: untreated (n=35 z-positions from 14 GVs, 3,011 particles identified), apyrase (n=13 z-positions from 4 GVs, 617 particles identified, p-value = 0.22), tropomyosin (n=24 z-positions from 12 GVs, 1,278 particles identified, p-value = 0.16), alpha-actinin (n=18 z-positions from 5 GVs, 1,254 particles identified, p-value = 0.05), and fascin (n=39 z-positions from 18 GVs, 2,123 particles identified, p-value = 0.34). Error bars = s.e.m. Scale bar is 10 μm in all images.|
|Figure 5. Cell size, organelle scaling, and gravitya, Linear scaling was observed between nuclear diameter, LGV, and cell diameter, Lcell from Stage I, IV, V, and VI oocytes and estimated from the literature (Supplementary Note). b, Linear scaling found between average nucleolar radius and nuclear diameter based on measurements in Stage I, IV, V and VI oocytes and estimated from literature (Supplementary Note). Nucleoli were measured from late stage oocytes (n=8 GVs), intermediate stage oocytes (n=17 GVs) and early stage oocytes (n=17 GVs). Error bars = s.d. Red solid line is the weighted best-fit line of the data. c, Cell growth during oogenesis in X. laevis. Each of the six stages is shown. Scale bar = 1 mm. d, State diagram of ℓsed vs. LGV. Black solid line separates region where gravity is negligible (white, ℓsed > LGV) from region where gravity dominates (red, ℓsed < LGV). Measurements (closed circles) and approximations (open circles) (Supplementary Note) are plotted for different nuclear bodies (black: nucleoli, green: HLBs/CBs, blue: small RNP complexes). For nuclear diameters > 10 μm, gravity becomes increasingly dominant for both HLBs and nucleoli, requiring a stabilizing F-actin scaffold.|