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

Biophysics and Bleeding Disorders

March is Bleeding Disorders Awareness Month in the US. More than three million Americans who have hemophilia, von Willebrand disease, and other rare bleeding disorders. These conditions prevent blood from clotting the way it should, which can lead to prolonged bleeding after injury, surgery, or physical trauma. We spoke with Biophysical Society member Valerie Tutwiler, an American Heart Association graduate research fellow in the lab of John Weisel at the University of Pennsylvania, about her hemostasis and thrombosis research.

What is the connection between your research and bleeding disorders?


This recent cover of Biophysical Journal shows Tutwiler, Wang, Litvinov, Weisel, and Shenoy’s image of a colorized scanning electron microscope image of a coronary artery thrombus extracted from a heart attack patient.

Blood clotting or hemostasis is the process that stems bleeding. On one hand if you have insufficient clotting this can result in prolonged bleeding, on the other hand a hypercoagulable state can result in thrombosis. Thrombi can result in the obstruction of blood flow, which can cause heart attacks and strokes. My thesis research pertains largely to studying one portion of the coagulation process blood clot contraction, or the volume shrinkage of the clot, which has been implicated to play a role in hemostasis and the restoration of blood flow past otherwise obstructive thrombi.

Why is your research important to those concerned about bleeding disorders?

While there is much known about the various aspects of blood clotting relatively little is known about the process of clot contraction despite the clinical implications of its importance in the formation of a strong hemostatic seal and the restoration of blood flow past otherwise obstructive thrombi. The study of clot contraction is a highly interdisciplinary problem and as a result can be of interest to researchers from many different fields. Platelets are active contractile cells, which interact with an extracellular matrix of fibrin, a naturally occurring polymer with unique mechanical properties. The fibrin matrix can be imbedded with other blood cells, such as red blood cells, as well. From a biophysical standpoint the mechanisms of clot contraction have not been well understood. To better elucidate this process, we performed a systematic study on how the molecular and cellular composition of the blood influences the rate and extent of clot contraction along with the mechanical properties of the contracting clot using a novel application of an optical tracking system.

Additionally, to further explore the mechanical nature of the clot contraction process we developed a mathematical model that couples active platelets with a passive viscoelastic matrix made up of fibrin and red blood cells. The model predicts the process of clot contraction and explains some of the experimental observations of clot size, structure and mechanical forces. Interestingly, we found that clot contraction is altered in thrombotic states such as ischemic stroke patients. Collectively, these findings show that the study of clot contraction has the potential to inform the development of diagnostics and therapeutics.

How did you get into this area of research?

Since beginning research I have been interested in applying engineering techniques to answer biological questions. I became interested in hemostasis and thrombosis research while completing my first co-op experience in undergrad.

How long have you been working on it?

I began doing hemotology research during my undergraduate career. However, I started studying clot contraction specifically when I started my PhD research.

Do you receive public funding for this work? If so, from what agency?

I am currently funded by the American Heart Association as a pre-doctoral fellow, although we also receive funding from the National Institute of Health and National Science Foundation.

Have you had any surprise findings thus far?

We were surprised to find such a striking decrease in the extent of clot contraction in ischemic stroke patients compared to healthy subjects. Correlations with stroke severity suggest that clot contraction may be a potential pathogenic factor in ischemic stroke. These findings have led us to expand our study to other pathological conditions as well.

What is particularly interesting about the work from the perspective of other researchers?

Due to the conservation of the basic principles of contractile proteins and motility, the information learned from the development of a mathematical model of active contractile cells interacting with a viscoelastic matrix can be applied to a variety of different processes.

What is particularly interesting about the work from the perspective of the public?

Bleeding and thrombotic conditions remain leading causes of death and disability worldwide. Gaining a more thorough understanding of the processes involved in hemostasis and thrombosis will lead to the development of more effective diagnostic tools and more targeted therapeutics.

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

Pi helps us describe almost everything, not just circles.

Most people know of π, or ‘pi’, as the number they learned in high school that has to do with circles: it is the ratio of a circle’s diameter to its circumference (π=C/d), the area of the circle is πr2 (especially hilarious because pie are round, not squared), etc. Some of us even remember it as an irrational number, meaning you cannot write it down as a simple fraction, and maybe some people, certainly not me, still have it memorized as starting with 3.14159265. What is less appreciated, however, is that this number has utility far beyond allowing us to calculate the area of a circle.

In biophysics, and in science in general, we use statistics to compare our data with our hypotheses. Many of the phenomena we measure fall along (or can be manipulated to fall along) a normal distribution. A normal distribution is a common continuous probability distribution characterized by the familiar “bell curve” shape, or Gaussian, which corresponds to the Gaussian distribution shown in the image below. When the mean, μ, is zero and the variance, σ2, is one, this function (the blue curve) is e^(-x2) and the area under the curve is the square root of pi! When the mean and variance are other values, the curve can be described more fully with the equation:

Where a = 1 / (σ (2π)1/2) a , b = μ, and c = σ.

pi day graph


Normalized Gaussian curves with expected value μ and variance σ2. The corresponding parameters are a = 1 / (σ (2π)1/2) a , b = μ, and c = σ.


How was the Gaussian distribution first determined, you may ask? While pi itself is thought to be first measured by the ancient Babylonians between 1900-1680 B.C., the Gaussian distribution originated in the 18th century when Abraham de Moivre started calculating gambling odds extremely precisely. De Moivre studied a very simple system at first: flipping a coin. He would calculate the probability of getting a certain number of heads from a certain number of coin flips. He found that as the number of events (coin flips) increased, the more his probability distribution approached a smooth curve. Thus he went about finding a mathematical expression for this curve, which resulted in the “normal curve”.

Independently, two mathematicians Adrain and Gauss in 1808 and 1809, respectively, developed the formula for the normal distribution and showed that errors observed in astronomical data fell along this distribution. Small errors in measurements occurred more frequently than large ones. The distribution was also independently discovered by Laplace, who elegantly showed how pi enters into the Gaussian distribution (which is summarized nicely here: Laplace also introduced the Central Limit Theorem, which proves that with a large enough number of samples the mean will be normally distributed, regardless of the underlying original distribution. This is why the normal distribution ends up popping up in so many places.

In biophysics, every time we think about mean and variance, calculate a p value (which assumes a normal distribution), do image processing, or try to understand the probabilities of a particular event, we owe a debt to pi. Not only do we use the Gaussian for statistics, but we also often use it in fields where we need to apply a potential or some external force either experimentally or in simulation. Basically, pi underlies all of the fundamental biological process we study on a daily basis. Thanks pi!

By Sonya Hanson, postdoc at Memorial Sloan Kettering Cancer Center


References: (Including public domain figure)


How to Write a Biophysics Article Worthy of Publication: Part 3- From Submission to Acceptance

William O. Hancock
Pennsylvania State University
Member, Biophysical Society Publications Committee

The first part of this series covered writing a first draft of a manuscript, and the second part covered the honing and polishing needed to bring the manuscript to the point where it is ready to submit to a journal.  The topic of this final article is navigating the process of submitting, revising, and getting your manuscript accepted for publication.

Choosing a journal

Because this piece is written with the Biophysical Journal in mind, your manuscript has hopefully developed into an appropriate submission to that journal.  From the journal website:

The mission of Biophysical Journal (BJ) is to publish the highest quality work that elucidates important biological, chemical, or physical mechanisms and provides quantitative insight into fundamental problems at the molecular, cellular, and systems, and whole-organism levels. Articles published in the Journal should be of general interest to quantitative biologists, regardless of their research specialty.

If your manuscript has evolved away from this definition, then you may want to choose another journal.  A good guide is to consider what journals are commonly read by colleagues in your field and fields relevant to your work.  Don’t be overly swayed by impact factors, and avoid predatory journals.  Consider the makeup of the Editorial Board who will be deciding on whether your manuscript is sent to review, and consider the business model of the journal.  Society-based journals (such as Biophysical Journal) carry the weight of the Society, usually have a history, and are generally run by scientists for scientists.

Before submitting your manuscript (and during the process of writing drafts and polishing your figures), consult the Guide for Authors and follow formatting, word count, and figure guidelines.  This will speed the submission and review of your manuscript, it increases the chance of acceptance, and it will save you time during later revision steps.

Most journals accept pre-submission inquiries to assess the suitability of the manuscript for the journal (and some journals require them).  This process involves sending your title and abstract together with a short letter to the editor, and it saves time for everyone involved.

Navigating the review process
picture-3The process of submitting a manuscript involves a number of decision points that are shown in the figure at right.  Upon initial submission, an editor will decide if the manuscript should be reviewed or be rejected (triaged) at this initial submission stage.  Considerations include suitability of the topic for the journal, novelty of the work, completeness of the work, and perceived impact.  Although it can be discouraging, this initial triage is another important time saver for everyone involved.  Avoiding rejection at this juncture can be helped by a pre-submission inquiry to determine suitability, and by a convincing cover letter.

Cover letter

One element that is sometimes underappreciated by authors is the cover letter, which provides the author a platform to persuade the editor of the importance of the work and its suitability for the journal.  The editor will generally be asking two questions:  (1) Is this work significant?  (2) Do the results justify the conclusions?  In the letter, it is important to distill the key findings into a few sentences.  However, more importantly, you want to place the work in the larger context of your field, and of the larger field of biophysics, cell biology, structural biology, or whatever your specialty may be.  This larger perspective is what the editor is thinking about — what is the impact of this manuscript, and will publishing it advance the mission of the journal?  Therefore, it can help to point out important recently published work by yourself and others that relates to the manuscript.  It is also good to remind the editor of the larger impact of the work on medicine, basic science, or technology.  Some of this persuasion means plucking text from the Introduction or Discussion of the manuscript, but it also requires stepping out to more of a 30,000 foot perspective and persuading the editor in a way not unlike a grant application.  Be specific and persuasive without being grandiose.

What makes an effective review?

Now that your manuscript has made it to peer review, it will be read by two or more reviewers who are considered experts in the subject of your manuscript.  The primary goal of the reviewers is to ask:  Do the results justify the conclusions?  A good review should provide substantive feedback that enables the editor to make an informed decision on the manuscript and the authors to revise and improve the manuscript.  Reviews generally begin with a brief summary of the findings and their relevance to the field, and may include the following:

  • A critical evaluation of the experiments, highlighting any flaws in experimental design, questionable interpretation of data, and any internal consistencies.
  • Highlighting previously published work (with references) that either contradict the work or may make the current experiments redundant.
  • Reasonable requests for further experiments, particularly control experiments but also obvious (important) experiments that the authors may have neglected.
  • Request for further analysis, reanalysis, or alternative presentation of experimental data, including adding or clarifying statistics.
  • A critique of the text and figures highlighting areas of confusion, excessive verbosity, or flawed logic.

A good review will be civil, will avoid vague complaints, and will not harp unnecessarily on small details that may not be related to the principal point of the manuscript.  The authors and editor are helped most by specificity and forthrightness in the evaluation of the manuscript.

Revising and responding to reviews

When the editor receives the reviews back, they then make a decision either to accept the manuscript as is (which is rare), reject the manuscript, or ask for major or minor revisions.  At this point, the author has to make a decision. Rejections can be appealed in select cases, but this avenue should be used sparingly and should have strong justification.  If the appeal is denied, then the authors should incorporate suggestions from reviewers before resubmitting to another journal, because it is likely that other reviewers will have the same complaints.

If minor revisions are requested, the authors can generally address the comments by editing the text, improving the figures, or making other modifications that don’t take much time.  In this case, the authors should attend to these tasks immediately and resubmit the revision.  In the case of major revisions, the authors have other decisions to make.  In some cases, the revisions and additional experiments requested are so extensive that it essentially requires rewriting the manuscript.  Depending on constraints, the best avenue may be to make minor modifications and submit it to a more specialized or lower profile journal.  If the decision is to revise and resubmit, then the authors must make a battle plan that involves some combination of further experiments, reanalysis of data, and revising the text and figures.  Often a limit of 90 or 120 days for resubmission is given (though deadlines can usually be extended by a reasonable request); this timeline provides a scale of the amount of new work that is expected.

When resubmitting a manuscript, the authors should also submit both a marked copy that highlights changes, and a point-by-point response to the reviewer comments.  It is expected that authors make a good faith effort to make edits and carry out further analysis and experiments.  A letter that tries to simply rebut every suggested experiment will not generate good will with the editor or reviewers.  That being said, it is reasonable to carry out some of the experiments suggested by reviewers and rebut suggested experiments that are onerous or extraneous.  Editors and reviewers will be more inclined to accept an explanation for not doing an experiment if you have followed their directive on other suggested work.  In some cases, data addressing a reviewer concern can be presented in the response to reviewers letter and not included in the text of the revised manuscript.

Upon resubmission, the editor may decide to accept the manuscript, or they may send it back out for review. At this point, the manuscript will be re-evaluated by one or more of the original reviewers.  In some cases, a new reviewer may be added to address a particular aspect of the manuscript.  If a major revision is requested and the authors have not carried out the requested experiments or sufficiently revised the work, the manuscript may be rejected at this point.  If the revisions were extensive and the reviewers still have complaints, then the manuscript may be sent back to the author for another round of revisions.  While this action is necessary in some cases, the extra work and time can be avoided by authors responding fully to critiques on their first revision and by reviewers detailing all of their concerns on their initial review and abstaining from making new critiques of aspects of the manuscript that were not commented on during the first round.

Publishing your paper

Hopefully this process will culminate with your manuscript being accepted for publication.  Congratulations!  But before you can move on to your next paper, there are a number of details to take care of.  First, it is imperative that the final revision that was submitted is error free.  It is worth taking the time now to be sure that the version that the journal has in hand has all figure numbers correct, all references in order, and other small details in place.  This is also the last time you will be able to edit the Supplemental Information, so be sure that document is properly formatted and is complete.  You will be sent page proofs for final checking, but it is best to have everything ironed out before the manuscript goes to proof stage, so that the final stage only involves checking for typesetting errors, figure placement, and related small details.

Over this three-part series, we have gone from data in a lab notebook to a published paper.  This process takes a lot of work, and although it gets easier the more you do it, publishing a paper is always a considerable effort.  However, peer-reviewed publications are the currency of science, and so the effort is necessary and worth it, and reaching this milestone is cause for celebration.  And, after the celebration dies down, then get back to the lab and do it again…


The author thanks Beth Staehle for assistance and advice, Olaf Anderson for many of the ideas that went into this work, and members of the Biophysical Society Publications Committee for many helpful suggestions.  He also thanks his mentors Joe Howard and Al Gordon, as well as his 8th grade grammar teacher, Jim Ernst, for teaching him how to write.  W.O.H. is supported by the NIGMS.

Helpful online resources

In addition to the references presented in Part 2 of this series, there are a number of more general resources online to help improve your scientific communication.

  • An excellent online writing resource with tutorials that focus on science writing fundamentals

  • Helpful eBook on writing scientific papers from Nature Education

  • A useful style guide, particularly for questions on grammar

  1. Gopen and J. Swan. The Science of Scientific Writing. American Scientist, November-December 1990.
  • An in-depth article that focuses on the readers’ perspective and breaks down sentence and paragraph structure for maximum communication


Michael Alley, The Craft of Scientific Writing, 3rd Edition, Springer, 1995.

Michael Jay Katz, From Research to Manuscript:  A Guide to Scientific Writing.  Springer Netherlands, 2009.

Interested in learning more about writing a good biophysics article?  Want a chance to ask questions?  Dr. Hancock will present a webinar on this subject today, March 10, at 1:00 PM ET.  Register at




The Science Behind the Image Contest Winners: Fluorescent Muscles in Living C. elegans

The BPS Art of Science Image Contest took place again this year, during the 61st Annual Meeting in New Orleans. The image that won third place was submitted by Ryan Littlefield, assistant professor, Department of Biology, University of South Alabama. Littlefield took some time to provide information about the image and the science it represents.


How did you compose this image?

Usually these C. elegans worms move around quite vigorously.  I added muscimol to prevent muscle contraction.  I picked three different types of worms that appear red only, green only, and red and green (which appear mostly yellow) and mixed them together on a thin pad of agarose.  The worms in the image all happened to clump together, resulting in a nice demonstration of the different color patterns.  I collected Z-stacks for each of the fields of view on an Andor spinning disk confocal microscope.  Using ImageJ software, I then made projections for the images that included the body wall muscle of the worms and pair-wise stitching of about six different projections.

What do you love about this image? 

The juxtaposition of all three types of transgenic worms being next to each other was very striking. The image includes all the different regions of the worms in various orientations, shows many of the different muscle types in these worms, and shows how the muscle cells fit together.

What do you want viewers to see or think about when they view this image?

The striated myofibrils in these worms are beautifully organized along their lengths, and it naturally raises the question of how this organization is achieved. In addition, the different muscle types show the viewer that there is a lot of diversity along the length of these 1mm worms.

How does this image reflect your scientific research?

I am interested in how actin and myosin become organized into functional, contractile bundles. In particular, I am interested in how actin filament lengths are specified in striated muscle.  In these worms, I modified an isoform of muscle myosin and tropomodulin with fluorescent proteins (mCherry and GFP, respectively) to determine how thin filaments are organized within these muscles.

What are some real-world applications of your research?

Both the uniformity of and specific lengths of actin filaments are important components of muscle physiology. Misregulation of actin filament lengths may be important factor in diseases including cardio- and skeletal and myopathies.  In addition, muscle damage from extended spaceflight, sarcopenia from aging, and acute muscle injuries may be slowed or prevented by interventions that prevent actin filament lengths from changing.

How does your research apply to those who are not working in your specific field?

Striated myofibrils are a dramatic example of a dynamic, self-organizing biological system that is attuned to a specific function (contraction).  Similar mechanisms for functional self-organization may also be used for other contractile actomyosin bundles, such as stress fibers and contractile fibers in smooth muscle, and for other dynamic cytoskeletal systems, such as flagella and microtubules in the mitotic spindle.