Understanding Alzheimer’s Disease through Biophysics Research

September 21 is World Alzheimer’s Day. Alzheimer’s disease is the leading cause of dementia, from which 47 million people worldwide suffer. It affects memory, thinking, orientation, comprehension, calculation, learning capacity, language, and judgement. 

In recognition of World Alzheimer’s Day, we spoke with two Biophysical Society members whose research aims to improve understanding of the mechanisms behind Alzheimer’s and other neurodegenerative diseases.


Liz Rhoades, University of Pennsylvania

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What is the connection between your research and Alzheimer’s disease?

We study the protein tau, which is a microtubule associated protein.  In Alzheimer’s and other neurodegenerative disorders, tau forms fibrillar aggregates that are deposited in brain tissue.  There are six isoforms of tau found in adult humans and alteration in the amounts of the isoforms are linked to disease development. Interactions of tau with microtubules are normally regulated by phosphorylation and tau aggregates derived from patient tissues are hyperphosphorylated, providing a link between loss of native tau function and disease as well.

Why is your research important to those concerned about Alzheimer’s?

We are working to understand basic aspects of tau function because we of the insight it provides to loss of function in disease.  Despite rather intensive study, molecular details of tau function are still lacking.  This is at least in part due to the fact that tau is large intrinsically disordered protein and thus it is challenging to characterize its structural features, particularly when associated with tubulin (the soluble building blocks of microtubules) and microtubules. For example, a few years ago, we observed that tau binds to soluble tubulin, a feature that had not previously received much attention. Our results suggests that it binds with similar affinity to tubulin as it does to microtubules which suggests that understanding how mutation impacts its interactions with tubulin is as important as characterizing how It interacts with microtubules. This is important because therapeutic strategies may very well involve targeting interactions between tau and its functional binding partners – we need to know who those partners are and the relevant features of the interaction!

Rhoades

The image here is from a paper that was published in PNAS last fall. It shows tau binding to two soluble tubulin dimers, and is based on our single molecule FRET measurements. It highlights how tau retains a primarily disordered state while binding and initiating tubulin polymerization. 

How did you get into this area of research?

We had been looking at tau aggregation in the lab for a couple of years, and then I had a few students – two graduate students and an undergraduate –  who were very interested in working with tubulin.  They were they ones who really pushed to get things up and running.  Anyone who has ever done a tubulin purification in their lab knows that this is not a trivial undertaking!

How long have you been working on it?

6 or 7 years

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

In the past we received funding from NSF (MCB)  and currently we receive funding from NIH (NIA).

Have you had any surprise findings thus far?

I think most findings I get excited about are surprises, but there are three that I can think of as particularly surprising.  The first was that tau binding to soluble tubulin had been largely overlooked in previous studies. We started working with soluble tubulin because it was easier for us to use in single molecule experiments (one of our primary tools) and only as we began to get interested results, do we recognize that there really was not a deep literature on tau-tubulin. The second was the tau point mutants linked to different neurodegenerative disorders bound more tightly to tubulin than the wild-type tau.  Our expectation based on the tau-microtubule literature was the mutation should decrease the binding affinity.  We are still working to understand the impact of this on tau function.  The third is that a region which flank the microtubule binding region has a high affinity for tubulin and allows for tau to bind to multiple tubulin dimers simultaneous to form a ‘fuzzy complex’.  This work was published in Biophysical Journal this summer.

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

I think some of it is probably methodology – we are using single molecule FRET and FCS to investigate tau-tubulin and working to make useful measurements in relatively complex, heterogeneous systems.

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

Understanding tau’s interactions with native binding partners may provide new targets for therapeutics.  I think anyone who has a family member or loved one who suffers from Alzheimer’s or another neurodegenerative disorder would find that interesting.


Dieter Willbold, Heinrich-Heine-Universität Düsseldorf and Forschungszentrum Jülich

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What is the connection between your research and Alzheimer’s disease?

My interest is focused on three-dimensional structures and dynamics of medically relevant proteins at atomic resolution and their interactions with native and artificial ligands. Autophagy and neurodegenerative diseases, which by the way do have a clear connection with each other, fall within my main interest areas. I want to understand protein aggregation in time and space at high resolution. And, I want to develop strategies and compounds that allow intervention and prevention. And the protein I am working on now for a very long time as a researcher is the Alzheimer’s disease (AD) related amyloid-beta protein (Aβ).

Why is your research important to those concerned about Alzheimer’s?

We do a lot of in-depth basic science on aggregation of Aβ, tau, alpha-synuclein, and many other proteins. We also design compounds for novel disease intervention strategies, and develop novel biomarkers and assays to measure them appropriately and most sensitively. All three, basic science, drug design and biomarker development, are based on biophysical principles and physico-chemical “thinking” and heavily rely on respective methods, such as NMR, x-ray crystallography, cryo-electron microscopy, ultracentrifugation, surface plasmon resonance, micro-calorimetry, TIRF microscopy, AFM, micro-thermophoresis, biolayer interference, and all kinds of spectroscopies.

Willbold

Figure 1: Cross section through the Aβ fibril illustrating the stepwise overlapping arrangement of the Aβ proteins. (Copyright: Forschungszentrum Jülich / HHU Düsseldorf / Gunnar Schröder). See also: http://www.fz-juelich.de/SharedDocs/Pressemitteilungen/UK/EN/2017/17-09-08-alzheimer-fibrillen.html .

How did you get into this area of research?

Already during my PhD project, which was mainly on the 3D structure determination of the transactivator protein (Tat) of the equine homologue of the HIV virus, I was engaged in structural studies of the amyloid-beta protein (Aβ) by NMR spectroscopy with some of the results published in 1995 with Paul Rösch being my supervisor and mentor. Ever since then, I was thinking of potential therapeutic intervention strategies. Since 1999, when I was heading my own junior research group in Jena, I had the necessary resources to at least start research on intervention strategies.

Soon after, I became involved in projects on prion diseases and prion protein (PrP) aggregation, when I accepted my first professorship at the Heinrich Heine University Düsseldorf in very close collaboration with Detlev Riesner. The common themes in these protein misfolding or protein aggregation diseases became quite clear. In my view, any intervention strategy – rather than a prevention strategy – needed to target toxic aggregates and get rid of them, rather than to reduce the formation of the monomeric species. As a biophysicist, I thought it would be a good idea to shift equilibria between monomers and aggregates away from the toxic aggregates (or oligomers as they are called today). To do this, we looked for compounds that bind to Aβ monomers with the free binding energy being used to lower the free energy level of monomers thus shifting the thermodynamic equilibrium towards Aβ monomers. The wording we use nowadays is that such a compound stabilizes Aβ in an aggregation-incompetent conformation. Because this is also happening with Aβ monomer units within Aβ oligomers, such a compound is also able to damage and destroy already pre-formed Aβ oligomers leading ultimately to their elimination. To identify a useful lead compound, we used mirror image phage display selection, a tool that allows selection of a compound from huge peptide libraries, but yielding a fully D-enantiomeric peptide, that does not have the disadvantages of normal L-peptides, which are very easily degraded and immunogenic. Our lead compound with the name “D3” (D-peptide from the third selection trial) showed really nice properties in vitro and in vivo. When we then wanted to elucidate the mechanism of action, it was essential to establish a whole zoo of methods and assays, which brought me even deeper into the field of protein aggregation in general and Alzheimer’s in particular. I just wanted to elucidate and pinpoint the mechanism of action and to reveal structural details of any interaction of Aβ with itself and with ligands.

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

Yes, now we do. In the beginning, I was not successful at securing additional money from funding agencies, e.g., the DFG. The project received great review reports about the underlying idea, but the panels ultimately decided that the project was too risky. Therefore, I used most of the institutional resources (which was not much in those years) for the project. Only in 2007, the Volkswagen-Stiftung funded a side project. Since 2013, I did and still do receive significant funding from the Helmholtz-Gemeinschaft, the federal ministry BMBF, the EU, and also from the Michael J Fox Foundation, the Weston Brain Institute, Alzheimer’s Research UK, as well as the Alzheimer’s Association. We have also been part of a JPND network.

Have you had any surprise findings thus far?

Yes indeed — many! Just to describe some: our lead compound D3 worked effectively not only in vitro, but also in several animal models. This has been a successful long-term collaboration with my dear colleagues Thomas van Groen, Inga Kadish and Antje Willuweit. D3 improved cognition in several models and decelerated neurodegeneration in an additional animal model that we have received from my dear collaborator Uli Demuth. We established an assay called QIAD that allows us to quantify Aβ oligomer elimination efficiency (https://www.ncbi.nlm.nih.gov/pubmed/26394756). We found that D3 efficiently eliminates Aβ oligomers, but many compounds that have already been in the clinics and failed are not able to do this. By following aggregation of N-terminally truncated and pyro-glutamate-modified Aβ (pEAβ) by NMR and CD spectroscopy, we found intermediates with helical secondary structure during aggregation.

When we tried to follow Aβ aggregation by SANS and analytical ultracentrifugation (AUC), we did not find any intermediates between monomers and penta- or hexamers (https://www.ncbi.nlm.nih.gov/pubmed/28559586). Thus, Aβ seed formation may be a reaction of very high order. In our recent research, (7th Sep 2017, https://www.ncbi.nlm.nih.gov/pubmed/28882996) we published a high resolution cryo-EM structure of Aβ fibrils. This structure provided many surprising findings in one hit including: all 42 residues of Aβ(1-42) are part of the fibril structure, there is no C2 symmetry between the two proto-filaments of the fibril, both ends of the fibril are different, each Aβ monomer subunit contacts many other subunits and six Aβ monomers form the minimal fibril unit. See also the respective report in alzforum.org: http://www.alzforum.org/news/research-news/amyloid-v-fibril-structure-bares-all .

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

I think that from the perspective of the public, two questions are relevant and interesting: Can we visualize highly complex things? Especially as structural biologists, we indeed can. Just look at the beautiful picture of the Aβ fibril at atomic resolution below. The second question is, of course: Can we contribute to efforts for improving the quality of life, for example by developing therapeutic strategies and drug candidates? Yes, we can also do (or at least try to do) this. It is, however, a huge undertaking that needs substantial funding and teamwork with many experts and specialists that you would not contact for basic science.

Just today (18th Sep 2017), we founded a company named Priavoid that will take an optimized derivative of D3 into clinical studies and hopefully to the market someday. In parallel, because Aβ oligomer elimination is our most favored mechanism of action, we have developed a technology called sFIDA (surface-based fluorescence intensity distribution analysis), which is able to quantify Aβ oligomers in body liquids like CSF and blood at single particle sensitivity (https://www.ncbi.nlm.nih.gov/pubmed/27823959). The development of this technology was and is important to ultimately show target engagement of our Aβ oligomer eliminating compounds. sFIDA will also be useful for early diagnosis of any protein misfolding disease, to recruit the “right” patients for clinical studies and to follow treatment success, if there is one. Thus, all in all, I think we have developed interesting results for the public domain.

Is there anything else you would like to add?

During the initial stages of the above described project to develop a novel therapeutic strategy for AD and to identify suitable compounds, there was only myself and my PhD student, Katja Wiesehan. Currently, there are many colleagues and coworkers that are the most experienced experts in their fields. It is such a beautiful experience to work and think with all of them and all the junior researchers, and to finally get things done. Please, have look at them and their groups in Düsseldorf (http://www.ipb.hhu.de/en.html) and Jülich (http://www.fz-juelich.de/ics/ics-6/EN/) with outstations in Grenoble and Hamburg. Finally, I shall not forget the many, many collaborators, of which I named only a few above.

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Temperature-Adapted Zebrafish Membranes Show Their Stripes

BPJ_113_6.c1.inddOrganisms that do not maintain a constant body temperature must have some mechanism to adapt their physiological functions in order to survive at a range of temperatures.  For individual cells within the organism, the cellular membrane serves as a platform for cellular signaling and cell-cell interaction. The organization and physical properties of plasma membrane lipids are sensitive to changes in temperature. When giant plasma membrane vesicles (GPMVs)— derived from cells grown in culture— are cooled slightly below growth temperature, they separate into distinct ordered and disordered liquid phases. These phases can be observed through fluorescence imaging of a phase-selective dye. ZF4 cells, derived from zebrafish, can be adapted to grow at a range of temperatures and GPMVs derived from these cells were used in our study to examine how these cells change the makeup of their plasma membranes to adapt to changes in temperature.

The cover image of the September 19th issue of Biophysical Journal focuses on an imagined “zebrafish,” which was composed of a fluorescence image of a ZF4-derived GPMV with stripe-like phase separated domains, combined with photographs of a zebrafish and a zebra.  These three images were blended together in Photoshop to create our chimeric zebrafish. In this cover image, we wanted to highlight the idea that lipid organization of the plasma membrane could be central to the physiology and function of the organism as a whole. For this reason, we used the fluorescence image of the phase-separated GPMV as the focal point of our “zebrafish,” and used the common visual motif of stripes to draw a comparison between the organization at the sub-cellular level of plasma membrane lipids to the organization at the level of the whole organism. This zebrafish swims through a sea of phase-separated GPMVs, shown in the background, again highlighting the theme of lipid organization.

Projects exploring how the physical properties of the plasma membrane impact membrane organization and function are ongoing in the Veatch Laboratory. This interest applies to a variety of biological processes, from immunoreceptor function to general anesthesia.

– Margaret Burns, Kathleen C. Wisser, Jing Wu, Ilya Levental, Sarah Veatch

Become a BPS Student Leader: Set Up One of the Inaugural Biophysical Society Student Chapters

Student Chapters blog

The Biophysical Society is excited to launch the BPS Student Chapter program this fall, with the first Chapters to be recognized starting in the Spring semester. This program aims to build active student chapters around the globe, increase student membership and participation within the Society, and promote biophysics as a discipline across college campuses through activities organized by the chapters. Students who become officers or participate in the chapters will have an opportunity to take an active leadership role within their institutions and the Biophysical Society, with special opportunities to participate in activities at future Society meetings.

Chapters may be formed within a single institution, or regional chapters may be developed among multiple, neighboring institutions. Recognized chapters will be reimbursed up to $200 by the Society to assist with getting started.

Chapters wishing to be recognized starting in the spring semester of 2018 must submit the Endorsement and Petition Form, Chapter Bylaws, and the Chapter Information Sheet to the Society Office via email to dmcnulty@biophysics.org by November 1, 2017, for consideration. Applicants will be notified in mid-December regarding the status of their recognition.

For more information and a complete list of instructions on forming an official BPS Student Chapter visit www.biophysics.org/StudentChapters.

Highlighting Biophysics Research During Sickle Cell Awareness Month

September is National Sickle Cell Awareness Month in the United States. Sickle cell disease is an inherited blood disorder that affects approximately 100,000 Americans and millions worldwide. It is particularly common among people whose ancestors come from Sub-Saharan Africa, South America, Cuba, Central America, Saudi Arabia, India, and Mediterranean countries such as Turkey, Greece, and Italy.

To recognize the awareness month, we spoke with BPS member George Em Karniadakis, Brown University, and his collaborators Xuejin Li, Brown University, and Ming Dao, MIT, about their research related to sickle cell disease. Their research was also featured on the cover of the July 11, 2017, issue of Biophysical Journal.


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What is the connection between your research and sickle cell disease?

Sickle cell disease (SCD) is the first identified molecular disease affecting more than 270,000 new patients each year. Our interests are in modeling multiscale biological systems using new mathematical and computational tools that we develop in our teams at Brown University and MIT in conjunction with carefully selected microfluidic experiments at MIT. We have an ongoing NIH-funded joint project that focuses on developing such validated predictive models for the sickle cell disease (SCD). In this project, with close collaboration between clinicians, experimentalists, applied mathematicians and physical chemists, we have been  developing new predictive patient-specific models of SCD, linking sub-cellular, cellular, and vessel-level phenomena spanning across four orders of magnitude in spatio-temporal scales. So far we have developed a validated patient-specific and data-driven multiscale modeling approach to probe the biophysical mechanisms involved in SCD from hemoglobin polymerization to vaso-occlusion.

Why is your research important to those concerned about sickle cell disease?

SCD is one of the most common genetic blood disorders that can cause several types of chronic and acute complications such as vaso-occlusive crises (VOC), hemolytic anemia, and sequestration crisis. It is also the first identified molecular disease (as early as 1947 by Linus Pauling), and the underlying molecular cause of the disease has been understood for more than half a century. However, progress in developing treatments to prevent painful VOC and associated symptoms has been slow. Consequently, we have been developing a “first-principles” multiscale approach that can handle the disparity of molecular, mesoscopic and macroscopic phenomena involved in SCD simultaneously. Such simulations could potentially answer questions concerning the links among sickle hemoglobin (HbS) polymerization, cell sickling, blood flow alteration, and eventually VOC. We hope, in turn, that these models will help in assessing effective drug strategies to combat the clinical symptoms of this genetic blood disorder.

Karniadakis

Figure 1. Dynamic behavior of individual sickle RBCs flowing in microfluidic channel. Inside the yellow circles are trapped sickle RBCs at the microgates, and inside the white circles are deformable RBCs, which are capable of circumnavigating trapped cells ahead of them by choosing a serpentine path (indicated by the white arrows).

How did you get into this area of research?

We have been working on multiscale modeling of blood disorders for more than 10 years.  In the very beginning, we were interested in developing new computational paradigms in multiscale simulations, which would enable us to perform multiscale realistic simulation of blood flow in the brain of a patient with an aneurysm. We then realized that the mesoscopic modeling of red blood cells (RBCs) and hemorheology in general seems to be the most effective approach for modeling malaria and other hematologic disorders. Then, we shifted our attention to the particle-based modeling of blood flow by employing the dissipative particle dynamics (DPD) method, which can seamlessly represent the RBC membrane, cytoskeleton, cytosol, surrounding plasma, and even the parasite in the malaria-infected RBCs. We developed multiscale RBC models and employed them to predict mechanical and rheological properties of RBCs and quantify molecular-level mechanical forces involved in bilayer–cytoskeletal dissociation in blood disorder. In 2012, we started to work on SCD, after realizing that no multiscale simulation studies of SCD had been conducted before – our work is the first of its kind!

How long have you been working on it?

As we mentioned above, we have been working in this field for more than 10 years.

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

Yes, we receive support from NHLBI, the institute within NIH focusing on blood disorders based on the interagency funding initiative pioneered by Dr. Grace Peng. For those who are interested in this multiscale consortium they can visit: https://www.imagwiki.nibib.nih.gov/

Have you had any surprise findings thus far?

Plenty! For example, at the vessel scale, using computer models, we have discovered that it is the soft and sticky type of RBCs that initiate the blockage process and lead to sickle cell crises and not the rigid sickle cells! This is the first study to identify a specific biophysical mechanism through which vaso-occlusion takes place. At the cellular scale, we have developed a tiny microfluidic device that can analyze the behavior of blood from SCD patients. Informed from the microfluidic experiments conducted by Dr. Ming Dao’s group at MIT, we have developed a unique patient-specific predictive model of sickle RBCs to characterize the complex behavior of sickle RBCs in narrow capillary-like microenvironment. At the sub-cellular (molecular) scale, we have developed a particle HbS model for studying the growth dynamics of polymer fibers (recent cover of Biophysical Journal). The simulations provide new details of how SCD manifests inside RBCs, which could help other medical researchers in developing new treatments.

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

It is known that the primary cause of the clinical phenotype of SCD is the intracellular polymerization of sickle hemoglobin (HbS) resulting in sickling of RBCs in deoxygenated conditions. However, the clinical expression of SCD is heterogeneous, making it hard to predict the risk of VOC, and resulting in a serious challenge for disease management. Our data-driven stochastic multiscale models, based on particle methods, can be used to explore and understand the dynamics of collective processes associated with vaso-occlusion that links together sub-cellular, cellular, and vessel phenomena. A similar computational framework can be applied to study blood flow in other hematologic disorders, including malaria, hereditary spherocytosis and elliptocytosis, as well as other blood pathological conditions in patients with diabetes mellitus or AIDS.  For example, in ongoing work we have quantified the biophysical characteristics of RBCs in type-2 diabetes mellitus (T2DM), from which the simulation results and their comparison with currently available experimental data are helpful in identifying a specific parametric model that best describes the main hallmarks of T2DM RBCs.  Perhaps, the most important extension is to connect such multiscale models to all the “omics” technologies (genomics, proteomics, metabolomics, etc.) to implement the vision of precision medicine advocated both in the U.S. and around the world.

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

Our studies provide new insight into what causes painful episodes in people with SCD. Using the computational models we could probe different mechanisms and validate diverse hypotheses regarding vaso-occlusion.  For example, we have shown that the rigid crescent-shaped RBCs —the hallmark of SCD — do not cause these blockages on their own. Instead, softer, deformable RBCs are known as cells that start the process by sticking to arteriolar and capillary walls. The rigid crescent-shaped cells then stack up behind these softer cells, creating a sort of a traffic jam.

Currently, hydroxyurea (HU) is the only approved medication in widespread use for the treatment of SCA, and it is thought to work by promoting the production of fetal hemoglobin, which can reduce sickling rate. Using the computational models, we can now run simulations that include fetal hemoglobin, which could help in establishing better dosage guidelines or in identifying a subgroup of patients who would benefit from this treatment or proposing a different type of treatment for others.

In addition, based on our own experience and knowledge, we also presented a short review in SIAM NEWs,   which provides the broader public with a general idea of computational modeling of blood disorders, including SCD. Here is the link to the review: https://sinews.siam.org/Details-Page/in-silico-medicine-multiscale-modeling-of-hematological-disorders.

Do you have a cool image you want to share with the blog post related to this research?

Yes, we have a cool image to share (figure 1). This image shows the different dynamic behavior between individual normal RBCs and sickle ones in microfluidic flow. Normal RBCs are round and flexible, and easily change shape to move through even the smallest blood vessels. Under deoxygenation, RBCs undergo sickling can be hard, sticky, and abnormally shaped, so they tend to get stuck at the microgates and block the blood flow. Once the adjacent microgates in the flow direction (from right to left) are blocked, the deformable RBCs (one is highlighted in white circle) appear to take a preferred path, i.e., they twist and turn along a serpentine path (as indicated by the white arrows) once they spot trapped sickle cells (one is highlighted in yellow circle) ahead of them.

On the State of Professional Opportunities for Women in Biophysics

GKP 2014At last year’s BPS meeting, while talking with several of you about how the Committee for Professional Opportunities for Women (CPOW) can better serve the BPS membership, I learned — much to my surprise — that the perception of gender equality and fairness in biophysics varies widely among our colleagues. At one extreme, some expressed disappointment that “not much has changed” since CPOW was formed in 1972; at the other, some declared “mission accomplished.” I suspect that like me, many of you will disagree with both statements, but I cannot guess where on the spectrum a consensus, if there is one, may lie.

To investigate these perceptions, CPOW will host a blog series where members can express their views on the subject by briefly answering these four questions: In your opinion,

  1. What is the current state of gender equality in science and biophysics?
  2. What is the value of having equality and true inclusiveness?
  3. What is one area that needs attention; and
  4. What is the one thing that can be done right away?

We kick off this initiative by publishing below answers from our fearless BPS Past President Suzanne Scarlata. You are encouraged to read and comment on these blog posts, and to volunteer your own answers by emailing them to Laura Phelan at lphelan@biophysics.org.

Thank you for your engagement. I look forward to hearing from you,

—Gabriela K. Popescu, CPOW Chair


sfscarlata

Suzanne Scarlata, Worcester Polytechnic Institute

What is the current state of gender equality in science and biophysics?

Compared to where we were 20 years ago, we’ve made a great deal of progress. Women now populate key positions in companies, universities, and scientific organizations. While we are still underrepresented especially in top positions, our numbers are growing and the trend is going up. However, we are far from shattering the glass ceiling.

Women have a better support system than in years past. In previous years when only a few senior women were around, women had to rely on father figures for advice in making their way through the system, which, of course, could limit the content of conversations. Now there are more women mentors both locally and through groups like the BPS that can bring together women to share their thoughts.

For the most part, I feel that time is on our side. Most colleagues my age and younger are fairly unbiased and this percentage is increasing every decade. Just a few years ago, I attended a meeting where I was the only female speaker. One of organizers was openly misogynistic which seemed to bother my male colleagues even more than me.

What is the value of having equality and true inclusiveness?

It goes without saying that having true inclusiveness and equality is invaluable. Everyone should be able to have the opportunity to work at their full potential and be appreciated and respected for what they do.

What is one area that needs attention?

Scientifically, we need to continue to promote ourselves (unfortunately, most of us are really bad at self-promotion) and our female colleagues by suggesting them for talks, for positions on editorial boards, and other leadership positions. We need to cite their articles when appropriate and give women the credit they deserve.

Importantly, we need to continually question whether we are treating our students, post-docs, and peers with encouragement and respect. The other day, a female undergraduate biochemistry major with a high GPA told me that her male advisor thought that she should focus on a career in writing and not science once she graduates. I had different advice!

What is the one thing that can be done right away?

While some countries have experienced recent setbacks regarding gender bias, we need to be persistent in encouraging equality both in and out of the lab. Nonscientists may not be aware of the many opportunities there are for their daughters in science, or aware of the problems they might encounter. We need to encourage women at all levels so that our numbers will grow.

When Ruth Bader Ginsburg (one of the nine members of the United States Supreme Court) was asked when she thinks there will be enough women on the court, she replied, “And my answer is when there are nine.”

For Microtubule Sliding, One Arm Is Better Than Two

BPJ_113_5.c1.inddThe versatile and dynamic network of the cytoskeleton scaffold would be stagnant and lifeless if not for the tiny nanoscopic machines called molecular motors. Kinesin motors, in particular, have captured the imagination of biologists and physicists because of their ability to transform ATP into anthropomorphic walking patterns on polar microtubule filaments, which make up a significant portion of the cytoskeleton. Recent experiments have shown that kinesin motors can crosslink adjacent microtubules and facilitate sliding between them resulting in cytoplasmic streaming in Drosophila cells. This facilitates faster distribution of molecules and organelles, and determines cell-shape.

But how do motors bring about microtubule sliding? How does the collective motion of microtubules depend on the movement of motor arms? In our work, we answer these questions by studying the effect of dimeric (one active arm, one anchored arm) and tetrameric (two active arms) kinesin motors on the dynamics of confined microtubules. Through our computer simulations we find that single-armed kinesins bring about much faster dynamics in specific regions of the confinement, compared to their two-armed counterpart. This goes against the intuitive idea that more arms pull more.

The cover image for the September 5th issue of the Biophysical Journal is our rendering of filament organization for two different motor types and the effects of these differences in the large-scale structure and dynamics of confined microtubules. The green shapes on the left represent the active motor heads that walk on polar microtubules. These are depicted as a linear array of dark-blue and yellow circles. The red blob depicts the anchor belonging to the single-armed, dimeric motor. Motor arms walk in specific directions on microtubules, and stretch, producing a sliding stress between microtubules.

The structures shown in the circular confinement on top consist of sluggish filament packages formed by tetrameric motors. The arrows at the bottom represent the highly dynamic microtubule arrangement formed by dimeric motors. Here, we also depicted the trajectories that three selected microtubules have taken. The cover image was crafted to highlight the large-scale biophysical implications of seemingly trivial and counterintuitive details in biology. Through this work we emphasize the vastly different cytoskeletal dynamics due to dimeric and tetrameric motors. By way of the trajectories, we capture the active layer of microtubules close to the circular confinement we observed for the single-armed motor systems.

– Arvind Ravichandran, Gerard Vliegenthart, Guglielmo Saggiorato, Gerhard Gompper, Thorsten Auth

Thematic Meeting, Berlin 2017: Session III

Welcome back to my post-conference note. Let’s jump right into the third session on “Interpreting Experiments Through Molecular Simulations“ including talks by Ceclilia Clementi, Michael Feig and Arianna Fornili on Saturday afternoon and with Shang-Te Danny Hsu, Jana Selent and Massimiliano Bonomi on Sunday morning.

The challenges of computational biophysics aiming to bridge molecular and cellular studies is among others due to the characterization of macromolecular systems by sets of different timescales, separated by large gaps. Celilia Clementi tackles this challenge by combining coherent state analysis and Markov State Modeling. Furthermore, she introduced a theoretical framework for the optimal combination of simulation and experiments in the definition of simplified coarse-grained Hamiltonian protein models. As those simplified models loose information, the combination with experimental data makes them more realistic and provides larger time-scales. She further illustrated the method and mentioned as application the coarse-graine model of FIP35.

Next, Michael Feig investigated protein dynamics and stability with crowders in simulations and experiments. He states protein destabilization in Villin due to crowding, in order to explain the differing results from NMR vs. MD. With Villin under dilute conditions, he observed oligomer formation in MD, whereas transient oligomers form on timescales longer than rotational translation diffusion. Rotational diffusion is slowed down by additional factors. In addition to his study on Villin, he could identify transitions between conformations connecting all states of Bacterial genomic DNA, based on targeted MD.

The last talk of this Saturday was given by Arianna Fornili about identification of rescue sites for protein function. As rescue mutations can be mimicked by drugs, their locations is of interest for drug design wherefore she developed a method. Double Force Scanning (DFS) mimics mutations by external forces, using an elastic network model to represent protein dynamics. In oder to model structural perturbations, linear response theory is used. In detail, she used the Fibonacci lattice for uniform distribution for force vectors. The performance of DFS was tested on p53, predicting 80% non-rescue sides correctly, and on an evolutionary dataset, where 79% of evolutionary rescue sites where predicted correctly.

Subsequently after Ariannas talk and a short coffee break, the first poster session was started. Although it was very crowed (which might have been a live experiments from the organizers, fitting to the last session), posters of high interests have been presented, leaving the presenters no break for a small sip of water. Afterwards, the program was open for all those aiming to explore the city of Berlin. On Sunday morning, the third session was continued with talks from Shang-Te Danny Hsu, Jana Selent and Massimiliano Bonomi.

The first session of this Sunday started with a rather biological topic on the structural basis of substrate recognition and chaperone activity of ribosome-associated trigger factor (TF) regulated by monomer-dimer-equilibrium from Shang-Te Danny Hsu. As the structure contains highly dynamic regions, those parts could not be easily resolved. This was also confirmed by solid state NMR studies, showing ribosome binding-induced conformational change in the ribosome binding domain of TF (TF-RBD). Shang-Te revealed TF substrate specificity by peptide array analysis, originating from recognition of averaged property rather than an exact sequence. Produced SAXS data showed a misfit between those and the crystal structure of TF. Further studies using pulse dipolar ESR spectroscopy revealed multiple dimer configurations of TF, which could be identified by chemical cross-linking. Furthermore, he modeled a dynamical system using several different techniques such as crystallography, NMR, SAXS. Comment: good luck for your PhD student 😉

Next, Jana Selent presented her work on the functional dynamics of the distal C-tail of arrestin. As introduced by her, phosphorylation of the GPCR C-tail triggers the arrestin pre-complex, including a partial C-tail displacement of arrestin. To investigate the role of the distal C-tail, she performed all-atom MD simulations and site-directed mutagenesis studies. She described its conformational space with preference to bind to positively charged residues. Tryptophan-induced dynamic quenching was increased for some residues. Furthermore, she investigated the mechanism of IP6-induced displacement, where IP6 displaces residue 393 to 400 but not further down upon binding to GPCR. As second topic, she investigated the C-edge loop of arrestin. By MD simulations, she showed that C-edge loop of pre-activated arrestin was able to penetrate the membrane, in contrast to activated arrestin. This was confirmed by quenching mutagenesis experiments. Finally, she postulates a step-wise binding process from unbound GPCR over an arrestin-GPCR pre-complex including the displacement of the distal C-tail and the penetration of the C-edge into the membrane, followed by the high affinity complex.

The last talk for this session was hold by Massimiliano Bonomi on integrative structural and dynamical biology with PLUMED-ISDB. As computational and experimental technique have their challenges or errors, a hybrid or integrative method could provide a more realistic view. By providing the module PLUMED-ISDB, he presents a way to investigate heterogeneous systems by including experimental data with a priori information. It uses a Bayesian inference method which accounts for data noise and averaged ensembles. He applied this metainference approach to cryo-EM data, able to explain the data better with a lower resolution as a mixture of dynamics and noise.

At last, he addresses challenges to the community, to those I could not agree more and therefore will end my post with his wishes: More distribution of ensembles of the community like structural models, model populations and protocols; the establishment of robust methods for ensemble comparison and validation; and a way to facilitate comparison of different ensemble modeling approaches by sharing methodologies.

Cheers,

Johanna