https://www.cell.com/cell/pdf/S0092-8674(14)00924-6.pdf

Probing the Stochastic, Motor-Driven Properties of the Cytoplasm Using Force Spectrum Microscopy

Impact of the physical microenvironment on tumor progression and metastasis.

Cells alter their mechanical properties in response to their local microenvironment; this plays a role in determining cell function and can even influence stem cell fate. Here, we identify a robust and unified relationship between cell stiffness and cell volume. As a cell spreads on a substrate, its volume decreases, while its stiffness concomitantly increases. We find that both cortical and cytoplasmic cell stiffness scale with volume for numerous perturbations, including varying substrate stiffness, cell spread area, and external osmotic pressure. The reduction of cell volume is a result of water efflux, which leads to a corresponding increase in intracellular molecular crowding. Furthermore, we find that changes in cell volume, and hence stiffness, alter stem-cell differentiation, regardless of the method by which these are induced. These observations reveal a surprising, previously unidentified relationship between cell stiffness and cell volume that strongly influences cell biology.

https://www.pnas.org/content/pnas/114/41/E8618.full.pdf

Cell volume change through water efflux impacts cell stiffness and stem cell fate.

The nuclear lamina is a fundamental constituent of metazoan nuclei. It is composed mainly of lamins, which are intermediate filament proteins that assemble into a filamentous meshwork, bridging the nuclear envelope and chromatin. Besides providing structural stability to the nucleus, the lamina is involved in many nuclear activities, including chromatin organization, transcription and replication. However, the structural organization of the nuclear lamina is poorly understood. Here we use cryo-electron tomography to obtain a detailed view of the organization of the lamin meshwork within the lamina. Data analysis of individual lamin filaments resolves a globular-decorated fibre appearance and shows that A- and B-type lamins assemble into tetrameric filaments of 3.5 nm thickness. Thus, lamins exhibit a structure that is remarkably different from the other canonical cytoskeletal elements. Our findings define the architecture of the nuclear lamin meshworks at molecular resolution, providing insights into their role in scaffolding the nuclear lamina.

/pdf/the-molecular-architecture-of-lamins-in-somatic-cells-385d0wl9u7.pdf

The molecular architecture of lamins in somatic cells

Injury to the central nervous system (CNS) alters the molecular and cellular composition of neural tissue and leads to glial scarring, which inhibits the regrowth of damaged axons. Mammalian glial scars supposedly form a chemical and mechanical barrier to neuronal regeneration. While tremendous effort has been devoted to identifying molecular characteristics of the scar, very little is known about its mechanical properties. Here we characterize spatiotemporal changes of the elastic stiffness of the injured rat neocortex and spinal cord at 1.5 and three weeks post-injury using atomic force microscopy. In contrast to scars in other mammalian tissues, CNS tissue significantly softens after injury. Expression levels of glial intermediate filaments (GFAP, vimentin) and extracellular matrix components (laminin, collagen IV) correlate with tissue softening. As tissue stiffness is a regulator of neuronal growth, our results may help to understand why mammalian neurons do not regenerate after injury.

/pdf/the-soft-mechanical-signature-of-glial-scars-in-the-central-1lp2w4epu9.pdf

The soft mechanical signature of glial scars in the central nervous system.

https://weitzlab.seas.harvard.edu/files/weitzlab/files/2013_ming_vimentin_0.pdf

The Role of Vimentin Intermediate Filaments in Cortical and Cytoplasmic Mechanics

Root phenotyping provides critical information to plant breeders for developing varieties with improved drought tolerance, greater root biomass, and greater nutrient use efficiency. Phenotyping roots in the natural environment is important for understanding the effect of the soil environment on root genotypic expressions. The goal of this work was to design and test a field‐scale mobile low‐field magnetic resonance imaging (LF‐MRI) Rhizotron that produces actionable root phenotyping data. We demonstrated this novel technology for root visualization and quantification using a LF‐MRI Rhizotron operating at 47 mT with two soil types. The LF‐MRI Rhizotron weights 453 kg, with a height of 90 cm, a diameter of 28 cm and an imaging field of view of 28 cm × 28 cm. The unit was operated in a Belk clay (Entic Hapluderts) and Weswood silt loam (Udifluventic Halustepts) generating 2‐D and 3‐D image data sets. The 2‐D image data had a collection time of 16.5 min per image at an image resolution of 2.2 mm per pixel. The 3‐D data had a collection time of 13 h per image with a 2.2 × 2.2 × 2.2 mm voxel resolution. Low‐field magnetic resonance imaging worked well for visualizing roots in moderate to high clay soils, demonstrating the potential for this technology; however, the broad application of this platform is hampered due to the prohibitively long scanning time to obtain 3‐D images. By increasing the field strength, and therefore the signal‐to‐noise ratio, faster scan times can enable a more useful system for root phenotyping.

Design and demonstration of a low‐field magnetic resonance imaging rhizotron for in‐field imaging of energy sorghum roots

Bioenergy sorghum is a highly productive drought tolerant C4 grass that accumulates 80% of its harvestable biomass in ~4 m length stems. Stem internode growth is regulated by development, shading, and hormones that modulate cell proliferation in intercalary meristems (IMs). In this study, sorghum stem IMs were localized above the pulvinus at the base of elongating internodes using MRI, microscopy, and transcriptome analysis. A change in cell morphology/organization occurred at the junction between the pulvinus and internode where LATERAL ORGAN BOUNDARIES (SbLOB), a boundary layer gene, was expressed. Inactivation of an AGCVIII kinase in DDYM (dw2) resulted in decreased SbLOB expression, disrupted IM localization and reduced internode cell proliferation. Transcriptome analysis identified ~1,000 genes involved in cell proliferation, hormone signaling and other functions selectively up-regulated in the IM compared to a non-meristematic stem tissue. This cohort of genes is expressed in apical dome stem tissues prior to localization of the IM at the base of elongating internodes. Gene regulatory network analysis identified connections between genes involved in hormone-signaling and cell proliferation. The results indicate that gibberellic acid induces accumulation of growth regulatory factors (GRFs) known to interact with ANGUSTIFOLIA (SbAN3), a master regulator of cell proliferation. GRF:AN3 was predicted to induce SbARF3/ETT expression and regulate SbAN3 expression in an auxin dependent manner. GRFs and ARFs regulate genes involved in cytokinin- and brassinosteroid signaling and cell proliferation. The results provide a molecular framework for understanding how hormone signaling regulates the expression of genes involved in cell proliferation in the stem IM.

https://onlinelibrary.wiley.com/doi/pdfdirect/10.1111/tpj.15960

Bioenergy sorghum stem growth regulation: Intercalary meristem localization, development and gene regulatory network analysis.

A simple spatial filter for 2D projection MR imaging is introduced. It works in the third (unresolved) direction to eliminate uniform or slowly varying interfering background signals. A constant amplitude gradient pulse in the unresolved direction is applied at the same time as the usual phase encode gradient during 2D acquisition. The filter is demonstrated for root imaging in soil, where background soil water signals can be troublesome. The filter suppresses the soil water signal while preserving the desired signal of plant roots. Fundamental to the operation of the filter is that the roots are sparse in the image domain, meaning there are relatively few pixels with multiple roots present. The performance of the through-plane filter is demonstrated and compares favorably to more conventional in-plane spatial filtering. 

Hilary Fabich

Papers

The Role of Vimentin Intermediate Filaments in Cortical and Cytoplasmic Mechanics

Design and demonstration of a low‐field magnetic resonance imaging rhizotron for in‐field imaging of energy sorghum roots

Bioenergy sorghum stem growth regulation: Intercalary meristem localization, development and gene regulatory network analysis.

Simple through-plane spatial filter for 2D MRI projections.