Mesoscopic Adaptive Resolution Scheme toward understanding of interactions between sickle cell fibers

BPJ_113_1.c1.inddSickle cell disease (SCD) is a molecular disease that affects hemoglobin. To understand the altered morphologies and mechanical properties of sickle red blood cells in SCD, it is important to investigate polymerization of the mutated form of hemoglobin (HbS) and subsequent interaction with the red blood cell (RBC) membrane.

The cover image for the July 11 issue of the Biophysical Journal is an artistic rendering of several RBCs and a white blood cell in the blood flow. Some RBCs are sickle-like shape. The largest RBC in the bottom left corner is shown in the protein level. The red, blue, and green particles represent the lipid particles, spectrin proteins, and actin junctions in the RBC membrane, respectively. Inside the RBC,  a single HbS fiber consists of 14 chains of HbS tetramers arranged in a 7 double-stranded configuration. The 14 chains of HbS tetramers are twisted about a common axis in a rope-like fashion. In the image, one small yellow particle is one HbS molecule, and  56 small particles are coarse-grained as one large yellow particle.

As shown in the image, the proposed hybrid HbS fiber model seamlessly couples these two HbS fiber models at different length scales by applying a mesoscopic adaptive resolution scheme (MARS). The stiff HbS fibers interact with the RBC membrane and distort the RBC to the sickle shape. Sickle RBC morphologies are determined by the number of HbS fiber domains and the structure of each domain inside the cells. In return, the RBC membrane also suppresses the growth of the HbS fiber. In addition to irregular shapes, sickle RBCs are characterized by increased cell rigidity due to intracellular fiber structures, resulting in blood flow impairment and vaso-occlusive crises in the microcirculation.

This cover was inspired by the pressing need to understand the integrated process of HbS nucleation and polymerization, and subsequent alterations of cell morphology, which is a multi-scale process ranging from nanometers to micrometers. In order to accurately describe this process, the hybrid HbS fiber model is employed to capture the dynamic process of polymerization of HbS fibers, while maintaining the mechanical properties of polymerized HbS fibers, thus providing a means of bridging the subcellular and cellular phenomena in sickle cell disease.

—- Lu Lu, He Li, Xin Bian, Xuejin Li, and George Em Karniadakis

Frozen Single Cell under Raman Spectroscopy

BPJ_112_12.c1.inddCryopreservation is the technology used to stabilize cells for a variety of applications, including diagnosis and treatment of disease. Because we don’t completely understand the mechanisms of freezing damage, poor or inadequate methods of preservation have limited our ability to use cells for cell therapy. Also, observations of cell responses could not be correlated to viability on a cell-by-cell basis using conventional low-temperature microscopy techniques. This study establishes our ability to measure the viability of individual frozen cells based on the correlation of cytochrome c distribution, a signal that can be detected using Raman spectroscopy, with trypan blue staining. With Raman spectroscopy, we are able to observe cells during freezing, and identify specific chemical and morphological changes inside the cell that result in life or death.

The cover image for the June 20 issue of the Biophysical Journal is an artistic rendering of frozen cells surrounded by extracellular ice and unfrozen solution. The background image is Lake Michigan in cold winter. Floating ice is separated by unfrozen water. The distribution pattern of the floating ice and unfrozen water is just like a frozen sample: ice crystals are separated by unfrozen concentrated solution. Schematic diagrams of frozen cells were imbedded in the background image to mimic a real frozen cell sample. In the diagram, the blue area represents unfrozen solution, the white area represents extracellular ice, the red line represents a region of cell membrane in close proximity to extracellular ice, and the black line represents a region of cell membrane in unfrozen solution between adjacent extracellular ice crystals. The schematic diagrams were precisely positioned in the background image such that the unfrozen solution in the diagram was co-located with unfrozen water in the background image and the extracellular ice crystals in the diagram were co-located with the floating ice in the background image. Our studies found that interactions between the cell membrane and extracellular ice resulted in intracellular ice formation (IIF), and increasing the distance between extracellular ice and cell membrane decreased the incidence of IIF.

Raman spectroscopy has enhanced our understanding of freezing damage. These studies can enable the development of new and improved cell preservation protocols and eventually improve the growth of cellular therapies and our ability to treat patients.

– Guanglin Yu, Yan Rou Yap, Katie Pollock, Allison Hubel

A Dynamic Biochemomechanical Model of Geometry-Confined Cell Spreading

BPJ_112_11.c1.inddCell spreading is involved in many physiological and pathological processes. In confluent multicellular systems, the dynamic evolution of an individual cell is influenced by its neighbors. It has been recognized that microsystems (e.g., microchambers) with defined geometry can affect the spatiotemporal dynamics of cells. However, it remains unclear how cells sense and respond to geometric confinement at the subcellular level. We answer this question by establishing a dynamic biochemomechanical model of geometry-confined cell spreading. This model reveals that the positioning of the cell-division plane is strongly affected by its boundary confinement.

The cover image for the June 6 issue of the Biophysical Journal illustrates the dynamic configurational evolution of a cell (blue) spreading in an L-shaped microchamber (silver). Its nucleus and microtubules are represented by the green sphere and emanated lines, respectively. The cell flattens and forms lamellipodia on the substrate but cannot step over the side walls of the chamber. In the initial spreading stage, the cell takes a round shape and spreads isotropically on the substrate before contacting the chamber boundary. The nucleus is positioned at its mass center (remote). Once the cell membrane contacts the chamber boundary, it may slide along or be fixed on the latter, depending on the force equilibrium condition (middle). For a cell undergoing anisotropic spreading, the length-dependent microtubule forces can drive the nucleus to move. Finally, the spreading cell reaches a steady-state configuration, which dictates the nuclear deformation and the cell-division plane (near).

The cover image was inspired by the cell spreading dynamics model which integrates biological, chemical, and mechanical mechanisms based on experimental observations. More details of the model can be found in our Biophysical Journal paper. This interdisciplinary work helps understand how microenvironments affect the spreading dynamics and division of cells. The findings also have potential applications in regulating cell division and designing cell-based sensors.

– Zi-Long Zhao, Zong-Yuan Liu, Jing Du, Guang-Kui Xu, Xi-Qiao Feng

A Shape Shifting Surface Layer

BPJ_112_9.c1.inddSurface layers (S-layers) are shells of protein that surround many microbes. Most S-layers are made of one or two proteins that self-assemble into a very thin protein crystal. Crystalline S-layers serve many functions in prokaryotes, including protection and shape determination. In some cases, crystalline S-layers enable pathogenic bacteria to infect humans, either by making the bacterium sticky, or by helping it avoid detection by our immune system. Surprisingly, the S-layer protein from the bacterium Caulobacter crescentus is not always crystalline, and can form two different structures on the surface of the cell.

Caulobacter crescentus is a crescent-shaped bacterium that can be found in many freshwater environments. The cover image for the May 9th issue of the Biophysical Journal is an artistic rendering of Caulobacter crescentus cells swimming in their natural habitat. The left side of the image depicts the surface of a single Caulobacter cell. Caulobacter’s S-layer protein can assume two forms, shown in green and blue. The green form is crystallized S-layer protein, which makes a hexagonal pattern on the surface of the cell. The blue form is an amorphous aggregate of the S-layer protein. That is, the protein is jumbled up and can’t make a repeating pattern.

Most previous work on S-layers has suggested that S-layers are crystalline out of necessity. However, our work indicates that in at least one case, this is not true. This inspired the creation of this cover image, which shows a surface layer that consists of two structural states: crystalline and amorphous. With this study and image, we hope to inspire further investigation into the structural flexibility, rather than the crystallinity, of S-layers.

Artist: Greg Stewart/SLAC

– Fatemeh Jabbarpour, Paul Bargar, John Nomellini, Po-Nan Li, Thomas Lane, Thomas Weiss, John Smit, Lucy Shapiro, Soichi Wakatsuki, Jonathan Herrmann

Modeling Unravels the Upper Limit of Mitotic Spindle Size

BPJ_112_7.c1.inddWhen talking about organelle size, most people believe the bigger the cell, the larger the organelle should be. In fact, this is not true, at least for mitotic spindles. Recent studies showed that the mitotic spindle size scales with the cell size in small cells, but approaches an upper limit in large cells. However, how the spindle size is sensed and regulated still eludes scientists.

The cover image for the April 11 issue of the Biophysical Journal shows a configuration sampled by a three-dimensional computational simulation guided by a general model for mitotic spindles. The model explicitly shows microtubules (colorful rods), centrosomes (green sphere), and chromosomes (pink bulks). Microtubules can be nucleated from the centrosomes, grow outward, and show the dynamic instability. When microtubules encounter the cortex or chromosome arms, they can generate pushing forces (red rods) due to the polymerization of microtubules. Various molecular motors on cortex and chromosomes, including dyneins (yellow dots) and kinesins (green dots), can bind to microtubules and generate pushing forces (green rods) or pulling forces (blue rods). Therefore, the centrosomes and chromosomes can move under these forces. In this way, the mitotic spindle can be self-assembled to form a bipolar structure with certain size, positioned to the cell center, and orientated to the long axis. Meanwhile, the chromosomes can be attached correctly and aligned on the equatorial plate.

This computational model is very useful for studying the size regulation of mitotic spindles. The spindle size is usually defined as the pole-to-pole distance, so that the problem of spindle size regulation is dependent on the positioning of two poles. The position of each pole is determined by the mechanical equilibrium between the cortical force and the chromosome force on the spindle pole.  For each pole, the chromosomes and the cortex are geometrically asymmetric. In small cells, the geometric asymmetry is small and the pole is nearly positioned to at the center of each half cell so that the spindle size scales with the cell size. However, in large cells, because few microtubules can reach the cell boundary, the geometric asymmetry is large and the spindle size is only determined by the chromosomes; that is, the spindle size approaches the upper limit. Therefore, this work revealed a novel and essential physical mechanism of the spindle size regulation.

There are certainly many other factors that influence spindle size but only quantitatively; they cannot explain the existence of the size limit of spindles.

This computational model provides a very powerful and robust tool. It can be combined with existing biochemical techniques to explore many important and interesting phenomena, including the positioning and orientation of mitotic spindles, the spontaneous oscillation of chromosomes, the mechanical response of spindles under various forces, and many other relevant questions.

– Jingchen Li and Hongyuan Jiang

Understand the Regulation of Learning and Memory Formation from a Molecular Prospective

BPJ_112_6.c1.inddWhen most people talk about calcium (Ca2+), they think about building bones and muscle contraction. In fact, calcium is also essential for learning and memory formation. Molecular basis for learning and memory formation has aroused attention since 1980s.  So what does calcium do with learning and memory formation? The calcium-modulating protein calmodulin (CaM) coordinates the activation of a family of Ca2+-regulated proteins, which are crucial for synaptic plasticity associated with learning and memory in neurons. These proteins include neurogranin (Ng) and CaM-dependent kinase II (CaMKII). In a resting cell, CaM is mostly reserved by Ng and free of Ca2+, whereas in a stimulated cell, CaM is able to bind Ca2+ and activate CaMII, which plays a pivotal role in learning and memory formation for both long-term potentiation and mechanisms for the modulation of synaptic efficacy.

The cover image for the March 28 issue of the Biophysical Journal shows the crystal structure of CaM-CaMKII peptide and the structure of CaM-Ng from coarse-grained molecular simulations. CaM molecules are ribbons in silver, calcium ions are represented by yellow beads, CaMKII peptide is in green surface representation, and Ng peptide is in red surface representation. One CaM-Ng peptide complex is near, where the Ng is aligned with a “pry” (pink); the other is far, indicating rich level of Ng. The images were rendered using the software Visual Molecular Dynamics developed by University of Illinois at Urbana-Champaign with the built-in Tachyon ray tracer. The illustration of a neuron in hippocampus is taken from Shelley Halpain, UC San Diego. Dendrites are green, dendritic spines red, and DNA (in cell nucleus) blue. The illustration of a human brain contains red dots to indicate active parts of the cerebral cortex.

Our computational study provides the very first detailed description at atomistic level ofhow binding of CaM with two distinct targets, Ng and CaMKII, influences the release of Ca2+ from CaM, as a molecular underpinning of CaM-dependent Ca2+ signaling in neurons. We believe this study bridges the molecular regulations in atomistic detail and the understanding of cellular process cascade of learning and memory formation.

– Pengzhi Zhang, Swarnendu Tripathi, Hoa Trinh, Margaret S. Cheung

PhosphoHero is in Charge of Neurofilaments’ Order

BPJ_112_5.c1.inddGels are neither solids nor liquids but rather a network of deformable and crosslinked polymers. Therefore, it is not surprising that the mechanical properties of synthetic gels are controlled by the degree of cross-linking, achieved, for example, by photopolymerization or the addition of chemical agents. One of the best examples of mechanically supporting bio-gels is the cytoskeleton, where crosslinked polymers (actin, microtubules and intermediate filaments) form a viscoelastic network. For microtubules and actin networks, analogues biological cross-linkers (associated proteins) have been identified. Nonetheless, in some important cases, the biophysical crosslinking mechanism or the existence of associated crosslinking proteins have not been identified.

Neurofilaments (NF) are neuronal specific intermediate filaments that form spaced filamentous networks in the long axon projections. Each neurofilament resembles a bottlebrush: a semi-flexible filament decorated with protruding floppy (intrinsically disordered) long carboxyl terminal tails. The tails engage in extensive crosslinking interactions, which have been the focus of many studies.

In addition to their lack of secondary rigid structure, NF tails contain many “phosphorylation sites”. These sites are specific amino-acid sequences recognized by enzymes that can add or remove charged phosphate groups, known as phosphorylation and dephosphorylation, respectively.

Our cover image for the March 14 issue of the Biophysical Journal illustrates an NF gel made of well aligned bottlebrushes at the front, and un-oriented ones at the back. The superhero (PhosphoHero) artistically illustrates the roles of NF phosphorylation. On the one hand, PhosphoHero increases the cross-linking between the filaments via the generation of ionic bridging between opposite charged residues. This in turn aligns the red filaments in nematic liquid crystalline order, as depicted by the crossed polarized NF hydrogel microscopy in the background. On the other hand, phosphorylation also increases the tails’ net negative charge, and consequently its compression response. Thus, phosphorylation acts as a regulatory knob to control the structure, orientation and mechanical properties of the cellular scaffold, the cytoskeleton.

Future studies into the role of intrinsically disordered proteins, and in particular their tunable phosphorylation states and their role in long-range alignment should be full of further surprises. Intrinsically disordered proteins were evolutionally selected to hold functional, although sometimes atypical properties, characteristic to superheroes.

The cover was hand-drawn and then digitally colored in Photoshop by Eliran Malka.

– Eti Malka-Gibor, Micha Kornreich, Adi Laser-Azogui, Ofer Doron, Irena Zingerman-Koladko, Jan Harapin, Ohad Medalia, Roy Beck