What’s Going on With the Federal Budget?

budget3It’s Thanksgiving Week and Congress is in recess.  Perhaps the last you heard about the federal budget for FY 2016 was that there was a bipartisan deal at the end of October.  Sounded like a good outcome.  That is true, but that deal didn’t actually provide funding for the coming year; it just increased the amount of money that could be spent.  Congress has until December 11 to figure out how it is going to divide up those additional dollars and pass a bill to fund the government for the coming year.  So why haven’t you heard much?

After the October budget deal, Congress began working behind closed doors on how to appropriate the additional dollars. The appropriations chairmen let their subcommittees know how much money they had to divide up among the programs for which they were responsible. These numbers were not made public. The subcommittees were supposed to send their proposals back to the Congressional leadership by November 20.  It is rumored that the conversations have not only focused on dollar amounts for each programs, but also on what policy riders will be included in the final bill.  Policy riders are directives that require certain actions or disallow certain actions by federal agencies.  The Democrats prefer a spending bill without riders; Republicans are pushing to include riders that reflect their priorities.  An example of a potential policy rider that affects scientists would be one that would require the National Science Foundation to certify that all funded grants represent research that is the national interest by making the U.S. more secure or improving the economy.   (This rider was in a spending bill approved by the House earlier this year, and could end up part of the ominbus bill currently being worked on.)

The rumors are that Congress will release an omnibus bill funding all federal agencies and programs on December 1, at which time we will be able to see how the agencies we care about have fared.  It is expected that the next ten days will be spent working out the riders and final numbers.

What has BPS been up to?

While the Hill has not been forthcoming with information during the past month, the Society has remained active in advocating for science funding in the final bill.  When the budget deal was reached, the Society sent a thank you letter to the White House and Congressional Leaders.  The Society has also sent communications to the Hill as a member of several coalitions in which it participates.  Many of these groups are also working on FY 2017 funding; a letter as sent by a coalition of coalitions, in support of raising science funding 5.2% across the board in 2017.

What can you do?

BPS has also been encouraging members to get involved.  A call for members to write to their Senators and Representative to thank them for the budget deal and advocate for science in FY 2016 went out to all U.S. members in early November. Thus far, 54 advocates have sent 166 letters.  If you haven’t written yet you can do so here.

Enjoy the quiet of Thanksgiving Week and stay tuned for more budget news in early December!


Ellipsoid Localization Microscopy

Cover 109-10 FinalMulti-layered protein coats are used for environmental protection, sensing, and interaction by many micro-organisms, including spore-forming bacteria and viruses. It is potentially very useful to measure the order of protein layers in these microbes, because this may indicate the function of different proteins — for example, which proteins form the outermost layers that protect the spore from lytic enzymes, and which hold the structure together?

Electron microscopy studies have established the order of some coat proteins, but the nature of the process is invasive, time-consuming and expensive. Conventional optical microscopy of fluorescent fusion proteins is non-invasive and highly practical, but lacks the resolution to distinguish adjacent protein layers. We have developed a computational technique that allows the ordering and size of coat protein layers to be determined from fluorescence images captured on a traditional wide-field fluorescence microscope.

The cover picture shows a wide-field fluorescence image, cut through to reveal a superresolved reconstruction produced by our ellipsoid localization microscopy method. We first model the spore coats as fluorescent ellipsoidal shells, and simulate the image that would be recorded by a microscope. The parameters of the model, such as position, average radius, aspect ratio, and shell thickness, are iteratively fit to the real image data, allowing us to determine the dimensions of protein shells to a precision better than 10 nm. This identifies the order and geometry of concentric protein layers and produces results consistent with previous electron microscopy studies, where available. Furthermore, our model also includes a parameter for the tendency of proteins to localize more heavily towards the poles, enabling a measurement of structural anisotropy. These parameters can then be fed back into our image generation pipeline, with the effect of optical blurring removed, producing a super-resolved reconstruction of the fluorescent protein shells. The cut-through line in the cover picture visually demonstrates that the sizes and orientations of shells are found correctly.

This ellipsoidal localization microscopy is our first development in a wider class of fluorescent shell localization techniques being developed in the Department of Chemical Engineering and Biotechnology, University of Cambridge. You can find more information about our research here.

  • Julia Manetsberger, James Manton, Miklos Erdelyi, Henry Lin, David Rees, Graham Christie, Eric Rees

Alzheimer’s Disease and Biophysics

November is Alzheimer’s Disease Awareness Month in the US. An estimated 5.3 million Americans suffer from the disease, which is particularly prevalent among people age 65 and older. We recently spoke with BPS member Chuck Sanders, Vanderbilt University, Departments of Biochemistry and Medicine, Center for Structural Biology, about his research related to Alzheimer’s disease, which has also affected his own family.

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

While not without controversy, the production of the amyloid-beta polypeptide is still generally thought to be central to the development of Alzheimer’s disease.  My lab studies the immediate precursor of amyloid-beta, the 99 residue transmembrane C-terminal domain of the amyloid precursor protein (C99).

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

Developing an understanding of the structure and molecular interactions of C99 may provide the basis for developing strategies to prevent its cleavage by gamma-secretase to generate amyloid-beta.  This would be expected to help prevent or even treat Alzheimer’s disease.

How did you get into this area of research and how long have you been working on it?

I am embarrassed to say so, but we originally chose C99 as a target for structural studies when we tried to express and purify over 20 disease-linked human membrane proteins small enough for NMR analysis.  C99 was chosen because it was one of the only proteins that could be well-expressed in E. coli.  I note that after we started studying C99 my father was diagnosed with Alzheimer’s disease, so this project has taken on a very personal quality to me.

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

Yes, we are very grateful to NIGMS for an ongoing RO1 grant that supports this project.  We also appreciate it that we also have had some previous (see grant) support for it from the Alzheimer’s Association.  I should add that we also recently obtained a grant from the BrightFocus foundation to start a new Alzheimer’s project on the TREM2 protein, but that is a different story.

Have you had any surprise findings thus far? 

Four!   First, the structure of C99 is more complicated and interesting than we expected.  It has a kinked transmembrane helix with flanking amphipathic helices as both termini.  Secondly, it binds cholesterol to form a stoichiometric complex and does so with physiologically-relevant affinity.  Third, the cholesterol binding site is built around GXXXG sequence motifs, which usually are thought to be associated with membrane protein oligomerization.  Fourth, its structure is remarkably tolerant of even major changes in membrane lipid composition.

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

alzheimers blog image

Binding of cholesterol to the kinked transmembrane domain of C99 with a conformational rearrangement of a surface-associated N-terminal helix and loop to optimize hydrogen bonding of the cholesterol head group in the membrane (here represent as a yellow slab). Not show in this picture is the disordered N-terminus or a long disordered loop that connect the end of the transmembrane domain to a surface-associated C-terminal helix (also not shown). Figure from Y. Song et al. (2013) Biochemistry 52, 5051-5054.

There are over 1500 papers on the relationship of cholesterol to Alzheimer’s disease (with elevated neuronal cholesterol being thought to be a pro-AD factor).  Finding that a central protein in the amyloidogenic pathway binds cholesterol avidly leads to a series of testable hypotheses regarding how cholesterol promotes Alzheimer’s

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

One might imagine that our work and the work of others suggests that lowering dietary cholesterol might help to prevent Alzheimer’s disease.  It remains to be seen if this is the case—damage of blood vessels to the brain by cholesterol-linked microinfarctions does seem to contribute to the pathology of Alzheimer’s.  However, with respect to possibly impacting amyloid-beta production it must be pointed out that dietary cholesterol cannot cross the blood-brain-barrier (BBB).  All the cholesterol found in neurons is made in the brain.  One wonders if the long term use (possibly over many years) of cholesterol lowering drugs that are able to cross the BBB might not have some benefit in helping to prevent or delay the onset of Alzheimer’s disease by reducing amyloid-beta production.

Protein-rich Clusters

Cover 109-9 Blog ImageThe cover image represents protein-rich clusters in insulin solution, where they likely serve as precursors to crystal nucleation.  Insulin crystallization is a part of insulin biosynthesis in the mammalian pancreas and an essential step in the manufacture of diabetes medicine.  The image was taken by oblique illumination dark-field microscopy and artificial coloring was applied for artistic effect.

Protein-rich clusters are of interest as nucleation precursors in protein crystallization, which is a part of industrial processes and laboratory procedures and is notoriously difficult.  Oftentimes crystals never form but instead amorphous aggregates, fibrils, dense liquids, and other undesirables appear.  The main kinetic impediment to protein nucleation appears to be the relatively large surface free energy  g of the crystal-solution interface.  In combination with the large size of the protein molecules, the high g leads to a high free energy barrier for nucleation.  In turn, this high barrier imposes high supersaturations at which nucleation may occur, thus expediting the formation of undesired solid phases.  To understand how proteins work around the high surface free energy problem, the two-step mechanism of nucleation was put forth.  According to this mechanism, crystal nuclei assemble within preexisting protein-rich clusters rather than from molecules in the dilute solution.  Owing to the high protein concentration in the clusters, the surface free energy of a crystalline nucleus emerging in them drops off by up to two orders of magnitude from g of a nucleus forming in the solution.  Thus, the protein-rich clusters emerge as crucial prerequisites for protein crystal nucleation.

The protein-rich clusters are much larger than the prediction of a colloid clustering scenario, which assumes structurally intact molecules and evaluates the balance of attractive and repulsive forces between them.  Our group put forth an alternative kinetic mechanism, according to which the clusters consist of a concentrated mixture of protein complexes and monomers.  Our paper in the November 3 issue of Biophysical Journal presents decisive evidence in favor of a kinetic mechanism.  We use the highly basic protein lysozyme at nearly neutral and lower pH as a model and explore the response of the cluster population to the electrostatic forces, which govern numerous biophysical phenomena, including crystallization and fibrillization. The results demonstrate that the Coulomb forces that govern aggregation in biological systems and many other phenomena in nature do not affect the cluster size.  In combination with other cluster behaviors, this response demonstrates that the mesoscopic clusters represent a novel class of protein condensate.  The clusters form by a unique mechanism, which includes the accumulation of transient protein oligomers that are linked by hydrophobic bonds between the peptide backbones exposed to the solvent after partial protein unfolding.  Our findings indicate that fine-tuning of the intra- and inter-molecular water-structuring interactions may be an essential tool to control the cluster population and in this way enhance or suppress protein crystallization and fibrillization.

– Maria Vorontsova, Ho Yin Chan, Vassiliy Lubchenko, Peter Vekilov

Addressing Our Own Implicit Bias

Gabriela Popescu, University of Buffalo and chair of the Biophysical Society’s Committee for Professional Opportunities for Women, reflects on the gaps between our consciously held values and our implicit biases, and how we can work against those biases.

Like many, I believe that I genuinely value diversity, and think of myself as being inclusive and fair, striving to do the right thing (most of the time). Several months ago, I learned otherwise.

After listening to a talk by Dr. Mahzarin Banaji, a leading researcher on implicit bias, I was inspired to check on this belief!!! I took the test at Project Implicit and I was humbled to learn that like the many I aspire to enlighten,Biophysical_0080(2-8-15)_1 I need a good dose of enlightenment myself; like society as a whole, I too am unconsciously doubtful of women’s professional commitment, ability, and talents, and therefore in situations where information is insufficient or inconclusive I tend to err on the side of…exclusion. From Dr. Banaji I also learned that I do this not because deep down I am a mean person, but because through eons of evolution when left unchecked my brain decides for me that it is better to be safe than sorry. What to do? How can I align my behaviors with my consciously chosen values of inclusion and diversity in science, in leadership, and in everyday decisions?

Experts advise taking a minute to write down our commitment to fairness and inclusion, and our personal reasons for this commitment. I tried it, and at least for me it worked! When I remind myself regularly of my intention to be fair and inclusive, I bring these values too into the mix of criteria my brain uses to make decisions, I bring them into the RAM!

To give an example, have you ever, when serving on a committee, read the committee’s charge and statement of purpose at the beginning of its meetings? BPS committees and Council started doing this a few years ago and I find the practice helpful. It brings front and center, into my RAM, the purpose of the committee and clarifies how I, as a member, can add value.

Someone suggested that reading a simple statement of commitment to fairness and inclusion at the beginning of a study section meeting, or an editorial board meeting may help us as reviewers and editors bring into alignment our behaviors, evaluations, and decisions with our espoused and presumably dearly held values of fairness and inclusion. It sounds simple and safe enough to try.

In other words “an inclusive thought a day may keep bias at bay.” What do you – explicitly – think?

Designing the Intracellular Superhighway

Cover 109-8Intracellular transport of vesicles and organelles is essential for maintaining spatial organization within eukaryotic cells. Transport of cargo is typically carried out by a combination of diffusion and active, motor-driven translation along networks of actin and microtubule cytoskeletal filaments. Perturbations to transport can significantly impact cellular viability and can result in disease at the organismal scale. While much work has been done to probe how motor properties affect transport, relatively little is understood about how the architecture of the network influences transport. Network architecture, characterized by the density, lengths, locations, and orientations of filaments, likely influences intracellular transport in much the same way that road connectivity is a critical determinant of vehicular traffic. In our study, we implement simulations in which filament networks with a wide range of architectures are explicitly modeled to investigate the effects of network properties on transport from the nucleus to the cell membrane.

The cover image shows a three-dimensional visualization of intracellular transport inspired by our simulations. The image was rendered in the open-source ray-tracing program POVRAY and shows a random network of cytoskeletal filaments (red/blue cylinders) guiding the transport of cargo vesicles (green/black spheres) through a cell. Filaments have a polarity, represented by the red and blue ends, which dictates the direction of motor traversal along the filaments. The cell nucleus is represented by the large, silver sphere in the center of the image. The cytoplasm, which contains all cellular material outside the nucleus, is represented by the thin metallic disk under the filaments that reflects the two-dimensional network geometry.

Our simulations showed that transport times are minimized when the mass of the cytoskeletal network is localized close to the nucleus. Prior models of transport that used intermittent processes to model ballistic and diffusive modes of transport assumed spatial homogeneity; here, we show that breaking this homogeneity may enhance transport. By explicitly modeling filaments, we reveal that mean transport times in random networks are primarily dictated by total filament mass. However, particular filament arrangements can cause “traps” near the nucleus that result in highly variable transport times. This variability can be mitigated by distributing the network mass over more filaments with shorter lengths and by introducing a buffer region between the nucleus and the transport network. Our results also show that multiple motors actually slow cargo transport and increase variability, suggesting that single motors can more robustly transport cargo than a collection of multiple motors over random networks.

Given that variability in transport times can be very large, it seems likely that characteristics of the transport system, such as network architecture, have been tuned, in some cases, to reduce variability, even at the expense of non-optimal mean transport times. Overall, our results suggest a diverse range of mechanisms by which molecular transport on filaments can be tuned and regulated.  There are  potential applications for enhancing reactions in biomimetic systems through rational transport network design. Our work (details at gopinathanlab.ucmerced.edu and whatislife.stanford.edu) will be of interest to biophysicists and bioengineers, as well as to molecular and cellular biologists working on intracellular transport-related topics.

– David Ando, Nickolay Korabel, Kerwyn Huang, Ajay Gopinathan

Get to Know: Frances Separovic, BPS Secretary

We recently spoke with Biophysical Society Secretary Frances Separovic, University of Melbourne, Australia, about why she loves biophysics, what makes Australia unique, and her surprising life goal.

frances-headshotWhat is your current position & area of research?

I am professor and Head of the School of Chemistry at the University of Melbourne. My primary research area is membrane biophysics and biological solid-state NMR spectroscopy. Our lab studies how peptides and toxins get into cell membranes.

What drew you to a career as a biophysicist?

Working out how things work, and being able to do this at an atomistic level is thrilling.

What do you find unique or special about BPS? Why are you excited to serve as secretary?

The diversity of fields covered by our members and the pervasive enthusiasm for discovery. As secretary, I hope to raise awareness of how biophysics underlies our understanding of biological systems and welcome the opportunity to help bring together the global biophysics community.

Who do you admire  and why?

Richard Feynman, Marie Curie and Nelson Mandela come to mind – their passion, persistence and pursuit to resolve often conflicted principles.

What do you like to do, aside from science?

Travel – although it is usually associated with science. I enjoy reading novels, movies, plays, exhibitions and stand-up comedy.

What makes you most proud about living in Australia?

Its natural beauty, lifestyle, and multiculturalism. I immigrated to Australia as a child and was fortunate to grow up learning from different cultures.

What do you want scientists to know about biophysics/science research in Australia?

The Australian Society for Biophysics will celebrate its 40th Anniversary this year. Although small on a world scale, we kick above our weight and, although on the other side of the globe, we are well connected. We’re also proud of the Braggs who, a hundred years ago, were awarded the Nobel Prize in Physics for X-ray crystallography.


The Nullarbor Plain, spanning the border between South Australia and Western Australia

What is something BPS members would be surprised to learn about you?

I love to drive long distances while listening to loud music – my ambition is to drive a road train [a truck pulling multiple trailers] across the Nullarbor.