Mechanotransduction in Collective Cell Migration and its Synthetic Mimic

Written by: Raman van Wee

Speaker:  Joachim P. Spatz

Department: Biophysical Chemistry

Subject: Mechanotransduction         

Location: Applied Sciences  

Date: 08-09-2017   

 

Text:

Spatz kicked off by showing us several videos of collective movement, both at the population level and at the cellular level. The latter included wound healing and formation of lateral line in zebrafish. Next a video of the motion of an epithelial monolayer of upper skin came by, it was very chaotic and movingly, which actually surprised me, I was expecting a rather static situation. Quantitively speaking groups of 10 cells up to 200 micrometer showed to behave as a collective, tuning the direction of the force to the group. In contrast there are leader cells, which go into a space on their own, followed up by the rest of the group. By reversing videos, behavior of leader cells could be investigated before it became apparent that the cell would become a leader cells. These cells seem to be predetermined, as they are at least 1000 micrometers apart of each other. The system regulates itself as shown by putting several leader cell too close to each other leading to the system eliminating those that are unwanted and thus leading to a stable, well distanced situation of leader cells. Remarkably if a group of followers, following a leader cell, threatens to exceed 10 followers, a new leader is brought forward from the group. I would say this requires extensive communication and coordination to execute well.

The second part of the seminar was about making a synthetic cell. This begins by making droplets having bilayer of membrane. Using sequential pico-injection microfluids, proteins and lipids could be inserted into the future cells. This sequential addition is very important, blending it all together at once doesn’t do it. Although once in the synthetic cell, the different components can not be separated with the eye, they do actually collaborate into larger structures. Interestingly, including myosine leads to the cell rotating around its axis. The need/strength for this rotating became clear when rotation was blocked by attaching a bead. In that case tension would build up until the cell would lose contact with the glass.

Afterwards we had the opportunity to speak to Spatz directly and ask him a few questions. We came to realization that he thinks in one year from now his synthetic cells will have some sort of mitochondria like energy factories. For the duplication part of life he thinks other research previously done can be very beneficial, drastically lowering the time required to integrate it in his cell. Besides, what actually surprised me was that his seminar was primarly based on 2 papers, whereas I expected more.

 

Conclusion:

Collective migration is mechanically regulated by the length up to which cells collectively integrate forces together. The cells follow this rule to select and follow new leaders. In synthetic cells, components have to be inserted one by one although they eventually entangle. Forces in a cell can lead to the cell rotating around its axis.

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Information of the structure of the universe

written by: Raman van Wee

Speaker:      Robbert Dijkgraaf

Department: Physics dept. Princeton

Subject:       The relation between the extremely large & the tiny world

Location:     Applied Sciences

Date: 07-09-2017   

Professor Dijkgraaf told us about the various scales scientists work at. We concluded that life (& Nanobiology) is centered at the middle of this sizescale.
According to Dijkgraaf this makes studying life extra challenging. In the outer sizes a different set of rules and equations hold, but in the middle, where life happens to be, the collide, making it extra hard. I found this both surprising and intriguing. Surprising because it posed quite a contradiction with my
instinctual beliefs, intuitionally I felt the center of scale is the relatively easy part and going out into space or into the world of quarks would be more challenging. It intrigued me as well, because as a Nanobiologist I am working right around this center!
The concepts in the introduction were well imaginable and thus comprehensible, as the seminar continued, it became less and less logical for me. A lot of concepts were new for me and were touched on only briefly, hence I lost parts of the seminar. Just like Dijkgraaf said one could easily get lost in H space while I didn’t even know what is comprised. Despite me not following all the content, it still widened my horizon, as it became apparent how much there is around I still don’t know anything about. A lot of those concepts and theories are also more or less related to my field of study and are therefore worth taking a closer look at some time. Although slightly off topic, when Cees Dekker introduced Dijkgraaf, he told us about some of the many committees, boards or councils Dijkgraaf has working in. I found this interesting as well, besides his own research he is also looking at science from different levels and angles, which I believe is anything but boring.

Conclusion:
All in all life is at the center of the sizescales. Beyond the cellular and organelle level there is yet so much to be discovered. In the future a beautiful goal
would be to implement concepts in the universe scale into the atomic scale and vice versa. Personally I am most interested in some of the quantum related theories and concepts.

How to achieve cellular replication without fail: lessons from bacterial cells.

Speaker: Christine Jacobs-Wagner
Department: Microbial Sciences Institute, Yale West Campus
Subject: How to achieve cellular replication without fail: lessons from bacterial cells.
Location: TU Delft (BN Seminar)
Date: 13-10-2017     

 Author: Nemo Andrea

 The topic of today’s talk was cellular replication, which, in Christine’s opinion, is the ability that separates the living from the non-living. In order to study this process, they study bacterial replication, as bacteria both divide rapidly and do so with high accuracy. The speaker stressed how, if one stops to think about it, achieving such short (20 minutes) division times in a varying environment with in the inherently highly stochastic environment of a living cell, is a truly remarkable feat. Thus, the robustness of bacterial replication and the relative simplicity of prokaryotic systems as compared to eukaryotic model systems make bacteria the candidate of choice for her research.

As there are many topics that can be explored within cellular replication, the speaker decided to focus on a single system that her group had recently done work on. Many bacteria have plasmids, which can contain vital functions for a bacterium, thus requiring efficient segregation of plasmids upon division. While plasmids diffuse throughout the cell and are thus, on average, equally divided among the two halves of a bacterium, suggesting that in the case of cell division no problem should arise, this is not the case. If a plasmid is present in high copy number, the chance of one daughter cell having significantly fewer copies of the plasmid is very small, but if a plasmid has low copy number the probability of one daughter cell ending up with no copies due to low number noise becomes significant. It is thus that bacteria have developed methods to effectively separate the plasmids that are present in low numbers. One could argue that it is not the bacterium driving the evolution of such a system but rather the plasmid itself, as plasmids that effectively do this ensure their survival, but that is really more a matter of perspective than anything else, and beside the point argued in this talk.

The plasmid that the group focused on displayed very curious behavior. The plasmids (in elongated bacteria) are distributed equidistantly along the long axis. The plasmids do diffuse, but stay in roughly the same area over time. It should be apparent that if such a distribution is maintained, the plasmids will be divided equally among daughter cells. As it turns out, this is achieved through a particular variant of the Par system. This variant uses just two proteins: PAR A/B, which are encoded on the plasmid itself. ParB binds to the plasmid, and does this by recognizing specific sequences on the plasmid, ensuring selective binding. ParA, on the other hand, binds to ParB, after which ParB will stimulate the ATPase activity of ParA, which will then unbind from the DNA. ParA also unspecifically binds to the DNA (of the bacterium). They observed (in case of a single plasmid) an oscillation of ParA from one side of the cell to the other, with the plasmid (with ParB) following the ParA signal. They then produced a first model, to test if simple Brownian dynamics of a diffusing plasmid could reproduce such behavior. They found that this was insufficient and concluded from this that a translocating force must be present to create this behavior.

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Figure taken from: DNA-relay mechanism is sufficient to explain ParA-dependent intracellular transport and patterning of single and multiple cargos. ; cropped

The key insight that was missing from the first simulation was the fact that DNA is not static, but moves around in the cell. They determined that the movement of a locus in the DNA can essentially be seen as an elastic harmonic potential, with a force of around 0.04pN. This is a really small force, even by cellular standards. They redid the simulations with this idea incorporated and found that they were able to reproduce the behavior observed in bacteria. The conceptual change is now as follows: The plasmid with ParB is pulled around through its multiple connections with multiple harmonic potentials through ParA. It will then unbind from the ParA in that area and move in a direction (slightly). In the new situation, the plasmid will ‘see’ more ParA in the direction of movement than behind it (which is the previous location, where it made ParA unbind) essentially creating a gradient of forces in the direction of movement. The speed of the plasmid seemed to correspond well with the force of the harmonic potentials. The initial symmetry breaking is caused by the stochastic nature of ATP hydrolysis.

The speaker mentioned another recent model that was developed that may explain similar phenomena, but pointed out that certain parts of this specific system are most likely not reconcilable with that model. The model presented today was also extremely elegant requiring just two components; one that binds to chromosomes, and another that modulates that first component’s lifetime. I quite enjoyed the talk, as Christine was an excellent presenter. While I am generally not easily convinced by simulations, this model seems to make sense both mechanistically and matches real world experiments. We also had a brief session afterwards to ask some questions, which was cut short due to unforeseen circumstances. I do appreciate her taking the time to sit down with us and answer some questions – even the ones not related to the talk.

What you can learn from computer modelling

Speaker: Christine Jacobs-Wagner
Department: Casimir Research School
Subject: How to achieve cellular replication without fail: lessons from bacterial cells
Location: Delft
Date: October 13, 2017

Although Christine Jacobs-Wagner studied biochemistry in college, she would describe herself more as a cell biologist. According to her, one is not defined by education, but rather by the questions they pose themselves. That’s why her lab is very multidisciplinary, which made her seminar really interesting. I liked the way she didn’t just come to Delft to lecture us about what’s she’s done, but rather to get feedback from the scientists here, who might have a stronger modelling background than hers. This is how science should work, and it’s a great feeling to be a part of it.

She spoke about the importance of distributing plasmids evenly during cell division. If one of the daughter cells doesn’t inherit an essential plasmid for some reason, it will not survive. That’s why it is crucial to have a method in place to guarantee the survival of both daughter cells. The ParA and ParB proteins make up such a system. This was the focus of her research.

Untitled
Plasmids should be distributed like these green dots

The ParA proteins bind to the nucleoid of a bacterium, and the ParB associates with plasmid DNA. When they interact, ParB induces a reaction which breaks the ParA-DNA bond. This is all well understood, but how would this have the desired effect of distributing all plasmids evenly? Well, it might be easier to observe what happens when there’s only one plasmid in the cell. It starts to oscillate along the long axis, but why?

 

 

Then she proceeded with an in silico model, a computer simulation. Unfortunately, the first model didn’t show the desired behaviour. The plasmid made a random walk, so the Par proteins didn’t do anything. At least, not until she adapted the model to make the DNA non-static. She could model ParA molecules as tiny springs, which oscillate around an equilibrium position. This was a very small change, but it had a great impact. It was all she needed to accurately model the oscillatory behaviour.

It was really cool to see such a small change make such a big difference. For further research and exploration of the ParA-B system, an analytical model would be useful, but that will also be a big challenge. I’m excited for the future!

Joachim Spatz: How wounds heal

Speaker: Joachim Spatz
Department:
Casimir Research School
Location: Delft
Date: September 8, 2017Transmission electron microscopy image of a biomimetic nuclear pore complex. Stefan Kowalczyk

Subject: Mechanotransduction in Collective Cell Migration and its Synthetic Mimic

Wound healing is a very important part of the medical sciences, so it is not very surprising that scientists have been studying it for a long time. Some 20 years ago, when looking at the way cells in the dermis – a specific layer of the human skin – closes a wound, they noticed a peculiar pattern. Although there is a lot of movement and fluctuations in this layer, some ‘leader cells’ will inevitably arise, evenly spaced and venturing into the void of the wound. These would be followed by many other cells and thus does healing begin.

 

In his talk, Joachim Spatz explained how he discovered the mechanism behind electing these leader cells by means of traction force microscopy. Essentially, he prepared a glass plate, covered with an elastic hydrogel containing a matrix of fluorescent beads to which cells adhere. Following their movement, one can measure the forces these cells exert on each other. The cells themselves were grown in a rectangular stencil on top of this gel. When he removed this stencil, he essentially created a wound, a void for the skin to grow into.

They made two important discoveries. Firstly, the cells are constantly moving around and fluctuating with respect to density and tension forces. They saw areas with high and low forces. This led them to measure the ‘force correlation length’. It’s a distance of about a 160 μm over which the beads are pulled in the same general direction and with about the same force. Secondly, just before a leader cell becomes visually apparent, there is an area of high forces just behind it. This means it is not a single cell’s autonomous decision to become a leader, but rather a consequence of the general population dynamics.

Along a long stretch of skin boundary, multiple leaders will emerge, and the spacing between them is always the same. This is explained by the fixed force correlation length. Next to the area of high tension behind the leader cell, there has got to be an area with low tension in which a leader cannot form. This means that the number of leader cells to form, important for the efficiency of the healing process, does not need to be actively regulated but is a consequence from the apparent chaos of the cells’ movements.

The second part of his seminar treated the creation of a synthetic cell. A subject that is being actively researched in Delft as well. It reminds me of Feynman’s famous quote: “What I cannot create, I do not understand”. The idea being, of course, that we’ll never completely understand how a cell works, until we can create it ourselves. Spatz’s approach was based on forming tiny water droplets in an oil solution. Since the two don’t mix such a water droplet can be the foundation for a synthetic cell. He then injected a mixture of hydrophilic and hydrophobic proteins into the droplet, which formed a membrane, further separating the cell from its environment.

Of course a normal cell has a membrane made of two layers of phospholipids, so he had to find a way to get these and other molecules into the cell. He ended up using an electrode to make the protein membrane porous allowing him to inject whatever he fancied. To make the lipid membrane, he injected a number of smaller vesicles, small membranes of lipid bilayers, and positive ions to destabilize them and fuse them together until they were big enough to form a membrane for the whole cell. FRAP experiments confirmed that these were similar to those of biological cells! Basically, they made the lipids fluorescent so they could see them under a microscope. Then they bleached an area by shining a very bright light on it. A property of biological membranes is that they are ‘liquid’, meaning that the individual phospholipid molecules are constantly moving around. This means that the extinguished phospholipids should diffuse into the greater, non-bleached, area and that other ones (still fluorescent) should be able to take their place. So the bleached area should disappear after some time. This was new to me and I find it very interesting.

But what is the link with the first part of his talk? Well, Spatz was also able to inject myosin and actin into the cell. These are essential to the movement of a cell and can be found in high concentrations in muscle cells, for example. When the cells floated up in the oil, they hit the glass plate and adhered to it. It meant that the cell suddenly had a fixed point of contact and that allowed the myosin to exert forces on the plate, which made the cell rotate. This can be seen through a microscope. If you also fix a little bead to the outside of this membrane, it can stop the rotation when it hits the glass. That is, until a sufficient force is generated to make the cell ‘jump’ over the bead, thereby actually moving the cell instead of rotating in place.

In conclusion, these artificial cells aren’t nearly as complex as their biological counterparts, but the technology is advancing and so does their complexity and capabilities. Joachim Spatz already succeeded in extracting them from the oil into an aqueous medium and he is working on an artificial mitochondrion. Lots of interesting research opportunities. I’m excited to hear what more will be discovered, or to work on it myself.

How to achieve cellular replication without fail: Lessons from bacterial cells

Speaker:     Christine Jacobs-Wagner

Department: Molecular, Cellular and Developmental Biology, Yale University

Subject:       How to achieve cellular replication without fail: Lessons from bacterial cells

Location:    TU Delft (BN)

Date:           13-10-2017

Author: Antoine Rolland

This talk was given by Christine Jacobs-Wagner. She started her talk by saying that most of the work in her lab is based on different phases in the self-replication of the cell. The research is mostly done on bacterial cells, since these are for example simpler than eukaryotic cells. In this talk, she introduced a mechanism that made sure that some low copy number plasmids in bacterial cells are evenly distributed among the two daughter cells after replication.

This mechanism actually consists of a pretty simple biochemical cycle, where two molecules are the most important: parA and parB. Both of these factors are encoded by the plasmids themselves. ParB can bind to the plasmid and cover it almost completely and parA can, after it has bound an ATP,  bind to the chromosomal DNA, which is spread almost all around the bacterial cell. ParB has a high affinity for parA that is bound to the DNA, and in this way the plasmid, which is covered with parB, can be connected to a given loci of the chromosomal DNA where parA is bound. After a small time step, the parB unbinds and the parA loses its ATP, after which the cycle can be repeated. Experiments have shown that this cycle creates a near-perfect distribution of the plasmids  along the long axis of bacterial cells. In case all the plasmids are merged, so only one large plasmid is present, this plasmid oscillates along the long axis over time, following the gradient of the parA concentration. However, in a first model in which only this cycle was introduced, this behavior wasn’t observed. This was caused by the fact that it was assumed that the parA molecules, once they were bound to the chromosomal DNA, would be static. This is not the case, since loci oscillate from its equilibrium position in the cell, because of a certain spring force.

When this part was introduced to the model, exactly the same behavior was observed as in the in vitro and in vivo experiments. Also, changing some parameters, like the amount of parA or the cell shape or size, still produced a nice distribution of the plasmids. What I would be interested in, is which parameters would disturb this distribution, and how this is done.

model parA parB

One of the models of a bacterium cell which was used, with in green the parB covered plasmid and in red parA

What I conclude from this talk is that a simple system, like the biochemical cycle of parA and parB, can still create a very nice distribution of the plasmids before replication. Before this talk, I would think that some really complex processes with a lot of components would be needed to create such a distribution. Also, I think it is nice to see how the model first didn’t work at all, but after one small change, namely having the parA molecules experience a spring force, it made the model work perfectly.

Agrobacterium-mediated transformation, pathogenesis by trans-kingdom conjugation

Speaker: Paul van Hooykaas
Department: Molecular Genetics, Leiden University
Subject: Agrobacterium-mediated transformation, pathogenesis by trans-kingdom conjugation
Location: Gorlaeus Laboratories, Leiden Bioscience Campus
Date: 28-09-2017

Author: Nemo Andrea

As is convention, Paul opened up his talk by outlining what the issues were that his research may be able to contribute to, and how these issues affect society. The genus of bacteria is the primary focus of his research, and thus he explained how and where these bacteria function. These bacteria are known to cause tumours in plants, which waste the plant’s resources. These tumours cell are plant cells with atypical morphology and will grow in growth factor deficient media. As plants are central to the food supply at all levels and as food supply for the increasing global population becomes an increasing challenge, research into such pathogens will be of importance. While fighting the reduction in crop yield due to this bacteria is a worthwhile exploration, the primary aim of this research was studying the means through which the bacterium induces this effects in plants – with the aim of generating molecular tools.

Agrobacterium will attach itself on plants and subsequently induce changes inside the plant cell. One of the effects of this is the induced production of opines, which are a type of chemical believed to aid the bacteria grow. This would explain the motivation of these bacteria to infect these plants. Relatively early on it was realized that a large plasmid was essential for the pathogenic effect. This plasmid, when present in non-pathogenic bacteria, was sufficient to make these bacteria cause tumors in plants. All the virulence ‘genes’ are therefore on the plasmid. A piece of this plasmid will end up in the plant cell, on which genes encoding opines and auxins and cytokinins are found. Not all genes on this section are fully understood yet, but many are believed to be related to cell growth – which would explain the tumour like behavior in the plant cells.

They found that this plasmid was quite resistant to random mutations, indicating that it is a very robust mechanism. This suggested it may a great method to inject specific genes into plants, by modifying the plasmid and inserting genes of interest. The method of infection used by agrobacteria works on many plant types. The monocots clade of plants do not form tumours like other clades, but it was found that they do produce opines, indicating that the plasmid section is successfully inserted in the plant genome. This means this method could be applied to a wide range of plant types. The speaker briefly highlighted the societal concern about GMO crops, but was of the opinion GMO crops are essential in solving food problems in the future, a notion I share. Another interesting discovery made was that certain plants (e.g. sweet potato) appear to still carry T-DNA (which is the DNA inserted by the bacterium) in their DNA outside of infection. While the speaker did not expand on this, it would be interesting to investigate why these genes have been retained (as they are detrimental to the plant in the context of bacterial infection).
They set out to identify the specific genes that mediate the transfer of the plasmid segment into the plant cells. They discovered that two genes, Vir D1 and D2, release the Vir genes from the plasmid. They induce nicks in the circular plasmid, which then releases the Vir strand as a linear piece of DNA. This single stranded DNA can then reform into a plasmid in the plant cell or be integrated in the genome. They noticed that this process resembles conjugation in bacteria, where bacteria can exchange genetic information by exchanging plasmids. They had also previously discovered that Agrobacterium can also transfer the T-DNA into fungi and even yeast, which is a remarkable fact. However, it has been known for a while that bacteria can conjugate DNA into yeast cells. These two things combined made them look at the conjugation machinery in greater detail.

Other discoveries made them believe that not only DNA is inserted in the plant cell, but also proteins. This would be unheard of and a significant discovery. They used a clever method of splitting up a GFP protein and leaving one part attached to the protein (in the virulent plasmid) and the other part expressed in the plant cell. If the protein from the bacterium would somehow be inserted in the plant cells, the GFP would recombine and become fluorescent. They had to use this method as GFP would not fit through the nanomachinery that is responsible for the transfer. By this approach they were able to confirm that even proteins were able to be transported into the plant cell.

The nanomachine in question is the Type 4 bacterial secretion system. Interesting to note is that this machinery is found in many bacteria and is used for conjugation, effector translation and more. Different types of bacteria have slightly altered versions of this nanomachine with different functions. The fact that this secretion system is responsible for the protein transport was surprising, but this does explain the parallels between conjugation and the mechanism of infection of Agrobacterium. The next question was how this machine could transport both DNA and proteins. As DNA and proteins are radically different structures, one would expect that only one of the two would be transported. They determined that VirD2 (nicking protein mentioned earlier) remains attached to the linear DNA and its C terminal domain is recognized by the secretion system, explaining DNA recognition and transport.


Structure of the type IV secretion system. Taken from: Waksman et al. 2010

When looking at the injected proteins in detail, they identified various proteins, of which they found one named virD5 to be of particular interest. Recent results have found that it inhibits growth in plants and yeast. Mechanistically, they found that it functions by binding to kinetochore associated proteins Spt4 and Ndc10. There, it destabilizes chromosomes and leads to chromosome missegregation. By overexpressing a gene that is known to counteract the pathway VirD5 deregulates, they were able to rescue the chromosome destabilization effect. The speaker postulated that the destabilization may buy time for the virulent T-DNA to be inserted in the DNA, as the cell cycle checks will be delayed. Additionally, the author described how this plasmid, provided the VirD5 protein is removed (as to not cause genetic instability), may prove to be very useful for industrial modification of plants.

When I was planning this seminar, I was worried it would have a heavy chemistry focus. Luckily, the chemistry aspects were kept to a minimum, making the talk quite accessible. I always enjoy hearing about plant biology, as this topic isn’t covered anywhere in the nanobiology course (understandably). The main attraction for me was the Type 4 Secretion system. Just like the bacterial flagella or ATP-synthase it is a wonderful (and versatile) nanomachine. There are many interesting directions to take in research regarding it, from the evolutionary reason for a particular adaption of the motor to the biophysics of unfolding and pulling a protein through it. I wonder what the reason is that the system is able to transport proteins into yeast but not animal cells. Perhaps modification of such a system may be an efficient way to inject proteins (for example medication) into cells.

Mechanotransduction in Collective Cell Migration and its Synthetic Mimic

Speaker:     Joachim P. Spatz

Department: Cellular Biophysics at Max Planck Institute for Medical Research,                                  Biophysical Chemistry at University of Heidelberg   

Subject:       Mechanotransduction in Collective Cell Migration and its Synthetic                                Mimic

Location:    TU Delft (BN)

Date:           8-9-2017

Author: Antoine Rolland

The lecture was divided into two parts. The first part was about the dynamic movement of skin cells, which is influenced by different forces acting on them. The second part was about how this behavior could potentially be mimicked in synthetic cells.

In the first part, Joachim Spatz began by saying that group movement is a very interesting topic in biology. From birds flying together to cells moving in groups, it is very exciting to study the way they move. Joachim Spatz has been looking at the way skin cells move when there is no boundary on one side of the cells. This is comparable to the way skin cells move in the process of healing a wound. What is well known is that there are so-called leader cells that begin to move into the open space, and that a group of cells go and follow that leader cell. What was found out is that these leader cells are already determined before they go and lead the other cells. This was discovered by measuring the force that was on the cells. What was observed was that there were strong forces behind the leader cells, when that cell cannot be distinguished yet. What was also very interesting, was that a leader cell always ‘recruited’ nearly the same amount of cells. This can be explained in a mechanical way, because this is the amount of cells that a leader cell can reach with the force that it exerts.  What I found the most interesting and surprising about this part is how mostly physical properties determine the way the cells move, rather than the biological properties of the cells. Joachim Spatz explained that there are sensors in the cells that can sense the force on them, and that in this way the leader cells can be determined. Moreover, the density had a big impact on the way the cells moved into the open space. The process went a lot faster when the density was lower. A higher density required to first eject some cells into the upper layer, before the process could start properly.

In the second part, Joachim Spatz explained how this movement of the cells could potentially be mimicked in synthetic cells. For this, a stable cell environment is needed. This can be achieved by creating nanodroplets with a polymer outside layer. To make an equivalent of the cell membrane, a lipid bilayer was added that bound to this polymer. It was proven that by adding components to this nanodroplet one after another, the result was better than when adding pre-assembled components. These nanodroplets also showed forms of adhesion, making it possible to apply and let them exert forces to potentially make the system from the first part in a synthetic manner. After the talk, we discussed some things with Joachim Spatz. What was interesting was the following question that arose: what exactly is life? To Joachim Spatz, for his cells to really become artificial life, they should be able to replicate, to move by their own and to create energy for themselves from the environment.

Leader cell with its follower cells

What really stood out to me from this talk is that physics and forces inside cells are way more important than I thought. Also, analyzing group movements of different kind of cells, could be very interesting for future research. Maybe we would discover that in group movement forces are often a very important factor. For me, this group movement is very interesting, and I wouldn’t mind learning more about this. Also, I think we are very close to creating synthetic life. Of course, ethical questions will rise if eventually real synthetic life could be created. I think this will be a big challenge for the future.

Mechanotransduction in collective cell migration and its synthetic mimic

Speaker: Joachim Spatz
Department Max Planck institute for medical research department of cellular biophysics
Subject: mechanotransduction in collective cell migration and its synthetic mimic
Location: TU Delft (BN Seminar)
Date: 08-09-2017
Author: Nemo Andrea

Joachim Spatz is a researcher from Germany who is working on a range of topics ranging from more traditional cell migration to microfluidics. We were informed that he was interested in the TU Delft’s BaSyC project. His talk today was a lecture in two parts with the first part focusing on cell migration and the latter on a new microfluidics-based approach to sequentially add components to a bilipid membrane.

 Part 1 – Cell migration

The talk started by introducing the motivation for his current work on migration: collective movement. Collective movement, as seen on the macro scale in flocks of birds and fish, is also seen in biology. Retinal cells are the textbook example of a collective form of cell migration during the formation of the eye. The two behaviors (on the macro and microscale) are very much the same behavior and are worth exploring from a physics point of view. Elucidating the principles that lead to collective motion is therefore of strong interest.

 To introduce his work on migration, we first had to be introduced to a few techniques, namely Traction Force Microscopy (TFM). This technique allows one to determine the forces exerted on the ECM by cells migrating on or through the material. From this, one can then also infer the stress between cells themselves (versus cells vs ECM). This can be done because the epithelial cells that they used exhibit a form of collective migration, where the cells are linked through various proteins. From these tractions and stresses, they were able to infer that each cell has a force correlation length that corresponds to about 10 cell lengths. This is then the maximum distance that cells can ‘’affect’’ other cells by exerting force/traction.

 The behavior they were interested in is related to wound healing. They spatially constrained the cells in a rectangular cutout, which could then be removed after which the cells would spread. Interestingly, they observed that cells do not spread out homogenously (equal dilation everywhere), but rather that certain cells would move outwards first and that these cells would drag others with them. These cells, dubbed ‘’leader cells’’, and their dynamics where studied. They wondered what the rules where for this system and how this leader cell first emerges. To do this, they observed the stress and force exerted by cells before the leader cell appears/can be identified. They found that the mean traction in the regions where a leader cell will appear is significantly higher than in cells that will not produce a leader cell, and similarly for stress. They then wished to find the rules for the spacing of leader cells, as they observed the spacing between leader cells (along the cell boundary) had a minimum value. This was not just an artifact, as when the aritificially patterned the cells in a way where the leader cells were spaced closer together, they would return to the natural spacing distance.

They found that the spacing of the leader cells depended on the ECM stiffness (which affects the traction exerted by each cell). This was verified by adding factors that increase or decrease internal actomyosin contraction, which similarly affects cell traction. An increase in traction was accompanied by an increase in leader cell spacing. They postulate is because with greater traction a leader cell can affect larger number of cells due to increased force correlation length.  

 Part 2 – Microfluidics

 In line with their work on migration, they wished to make bilipid membranes that had integrins in the membrane. Such vesicles could then adhere to the environment and could work as an artificial model for some aspects of cell migration. In their attempts to create these they ran into problems with the low mechanical strength and the low yield of the fabrication process. They then developed a new system: water droplets in oil with the water droplets being stabilized by a polymer shell. Through  this strong polymer shell (which can be moved using standard microfluidics platforms) they can inject many different proteins using a series of picoinjectors. As there is no limit to how many picoinjects can be used, and each picoinjector is placed after the other, one has the advantage of being able to sequentially add elements to the vesicle. This is a massive advantage, as various systems will not properly self-assemble if they are all ‘’’thrown in at once’’. Not only does this allow for precise control, the system also had very high throughput with the picoinjectors being able to handle roughly 500 cells per second.

microfluidics

image taken from a presentation by Marian Weiss titled ‘‘Droplet-Based Microfluidics for Sequential Bottom-Up Assembly of Functional Cell-Like Compartments

 Joachim showed us some beautiful demonstrations, with three differently labeled fluorescent actin types being injected into a single cell, and even adding actin and myosin into a vesicle. This then allowed the myosin to contract the actin, making the vesicles active in a real sense. As they did this they observed that the active actin vesicles were slowly rotating. They were able to turn this rotation into motion across a line using another bead attached to the outside of the vesicle, making essentially a very crude form of a migrating cell. (although mechanically completely different from natural cell migration).

 After this, they also added lipids to the vesicle, which, in the right concentration, allowed for the formation of a lipid bilayer in the polymer vesicle. This a remarkable feat, as making a membrane isn’t easy with traditional methods. Next to this, they also demonstrated ester formation and dynamics inside these vesicles in some truly breathtaking video fragments. Importantly, they also demonstrated that is was very easy to remove the polymer shell and just leave the bilipid membrane. To get back to their original goal, they managed to add integrins into this membrane using this technique. Depending on the concentration, they could modulate the extent to which cells adhered to fibrinogen. They are currently working on making artificial mitochondria, and were able to get ATPase and bacterial rhodopsin to work in this artificial context, which I consider to be a remarkable achievement.

 I found this to be the most excited seminar I have attended to date. The first part was interesting to me because I knew a fair amount about cell migration due to my Honours Programme Project, and the second part was extremely fascinating due to the wonderful things that they were able to do. I am very excited for work on the BaSyC project of the TU, so seeing a new promising method like this makes me very, very excited about what is to come.

Information, from the structure of the universe to life

Speaker: Robbert Dijkgraaf
Department: Kavli institute
Subject: Information, from the structure of the universe to life
Location: TU Delft
Date: 07-09-2017
Author: Renée van der Winden

For the Kavli Day on September 7th Robbert Dijkgraaf gave the opening lecture. In this seminar he did not talk about a particular research subject, but rather tried to give us an insight in the assumptions and dimensions we work with in science. In order to do this he discussed several ways in which we can distinguish different types of science and gave examples of many scientists and the different views they have on this matter.

The first thing Dijkgraaf discussed was the scale on which we operate. The smallest things in life we can explore are on the 10-35 m scale, while the biggest structures are about 1025 m. The things we are studying, life and cells, are right in the middle at around 10-5 m. When considering beauty and elegance in science, you can find it at the ends of this spectrum. Those are the places where the messy, everyday life structures crystallize into simple and clear-cut laws and phenomena. The fact that you see this at each end is a reflection of two ways to approach science: reductionism and emergence. Reductionism starts at bigger phenomena and tries to uncover the laws that underlie it. For example, going from a tree to photosynthesis. Emergence starts out at laws of nature and tries to see what they bring about. For instance, going from collisions of water molecules to fluid dynamics. Whichever approach you choose, the end product seems to be more elegant from that point of view, though not from the other, which is interesting. This realization is of the same order as one of the quotes Dijkgraaf used in his presentation, this particular one coming from John Wheeler: “Every law of physics is approximate, not precise”. Dijkgraaf used this quote to illustrate that no matter how elegant a theory might seem, it is still something we came up with to explain our data.Seminar 9

Figure 1: Exerpts from the movie ‘Powers of Ten’

Another thing Dijkgraaf illustrated is that what we consider to be truths, might not in fact be true completely and always. He pointed out the many theories Einstein has come up with to explain how relativity emerges from quantum theory, all while ignoring the possibility that things might be the other way around. Moreover, while discussing black holes, Dijkgraaf mentioned that something strange happens when you move within a certain zone around it, called the horizon. After the horizon, time flips and stops being endless, rather it moves to the center of the black hole, where time stops. What we consider as a given (the flow of time), turns out to be false under certain conditions.

Dijkgraaf ended his talk by saying that the scale of the powers of ten that he started his talk with, might not be the endpoint. He stated that it might be possible there is an even smaller layer below it, a so-called basement layer, which may be concerned with quantum information.

I had hoped to get a chance to see Robbert Dijkgraaf speak for a while, so I was very excited when that opportunity came around. His talk was easy to follow and interesting. I think Dijkgraaf is good at explaining even difficult topics concisely and clearly, so I would very much like to hear him speak about his own ongoing research as well at some point.