Cells on the Catwalk

BIOLOGY & MEDICINE_Biophysics

Life is motion and interaction with the environment. This is equally true of cells within an organism, but for cells to get from one place to another, they not only have to be able to move, they also have to interact with their environment. Joachim Spatz and his team at the Max Planck Institute for Medical Research in Heidelberg are studying how cells manage this. In his search for answers, the winner of the 2017 Leibniz Prize puts cells through their paces on catwalks and obstacle courses to test their adhesive properties.

TEXT CATARINA PIETSCHMANN

he adult human body com- Many cells have to cover distances of distinguish whether the surface is hard prises 100 trillion cells – an various lengths within the body: or soft by essentially tugging on it. “It’s almost unimaginably large during embryonic development, for like us probing the ground,” Spatz says. number, a one followed by example, but also during continuous “If it’s soft enough, I can safely fall to 14 zeroes. The cells of our remodeling processes in the adult the ground. If it’s made of stone, I’d bodiesT form organs such as the heart and body, cells must migrate from their better not. “Cells adapt their behavior kidneys, and tissues such as the skin and place of origin to the sites where they in a similar manner. “If a stem cell per- nerves. Some drift through the highly are needed. To do this, they have to ceives a hard environment, it will ma- branched vascular system in the form of know where they are and where they ture into bone or tissue cells. If, in con- blood cells, while others patrol the body need to go. trast, the substrate is soft, it matures for the immune system. But regardless into a nerve cell.” of the task they perform for the body as RECEPTORS SENSE THE Cells are even able to estimate the a whole, every cell is an individual. ENVIRONMENT number of small molecules near them. “Each must be able to perceive its envi- This gives them a sense of how import- ronment and respond to it,” says Joa- But how does a cell interact with its en- ant the latter are as chemical messengers chim Spatz. The biophysicist headed the vironment? “For one thing, it detects and whether a response is appropriate. New Material and Biosystems Depart- chemical signals via receptors on the “It is also important for migrating cells ment of the Max Planck Institute for In- cell membrane,” Spatz explains. Small to know how densely a surface is cov- telligent Systems until the end of 2015. molecules attach themselves to mem- ered by molecules,” Spatz says. “If the In 2016, he and his team moved to the brane-spanning proteins and activate distance between the molecules is 60 Max Planck Institute for Medical Re- signaling pathways in the cell’s interi- nanometers or more, the cells are unable search in Heidelberg. Together, they are or. Depending on the information to read the information.” devising biophysical experiments, mea- transmitted, genes are revved up, throt- In order to observe the behavior of suring techniques and model systems to tled or switched on or off. individual cells, Spatz’s coworkers have investigate the motility and adhesion of In addition, the cell also perceives developed a sort of cellular catwalk,

individual cells and cell collectives. its underlying surface tactically. It can which they call cellwalks. Each cellwalk Photo: MPI for Intelligent Systems

48 MaxPlanckResearch 1 | 17 A cell holds fast: Adhesive proteins concentrate at binding sites (yellow) to form a textured surface (blue). A meshwork of actin proteins (red) links adhesion points across the cell. Photo: MPI for Intelligent Systems

1 | 17 MaxPlanckResearch 49 Jacopo Di Russo studies the migration behavior of human skin cells and measures the force the cells exert to cling to a surface. To do this, he binds various proteins to gels to alter the properties of the surface.

consists of a polymer surface coated cell type. Even within a single cell line from Heidelberg University Hospital with ultrafine gold particles. The gold there are “tortoises” and “hares” be- approached Spatz: couldn’t they put a acts as anchoring points for biomole- cause the speed is also determined by “real” protozoon, namely the malaria cules – peptides, for example, or anti- the cell’s stage of development. organism Plasmodium, on the cellwalk bodies, to which, in turn, cell receptors Cells, of course, have no feet, but for a change? can bind. This provides support for the they do have something similar. They The tropical disease, which is caused cell, because without these “nubs,” the crawl forward on sheet-like exten- by the bite of an infected Anopheles surface would be far too slippery. By sions, called lamellipodia. The cell mosquito, still claims half a million varying the base polymer, the spacing membrane bulges out and spreads for- lives every year. The mobility of the between the anchoring points can be ward. “It’s comparable to us taking a pathogens, which are injected into the adjusted to 30, 50, 70, 100 or 150 nano- step forward,” Spatz explains. The human skin from the mosquito’s sali- meters. For the cellwalk, a small piece lamellipodium adheres to the sub- vary glands, is crucial for the “success” of the synthetic surface is placed in a strate by rearranging and aligning pro- of the infection. “The sickle-shaped culture dish with nutrient solution. In- tein molecules in the cell’s interior – sporozoites move at a top speed of 10 dividual cells are then placed on top of integrins as well as actin and myosin micrometers per second, nearly 100 it, the microscope is focused and the filaments. The rest of the cell body is times faster than human cells,” camera is activated. then pulled forward, and the anchor- Frischknecht explains. “We’re interest- ing site is released again. This process ed in finding out how they do it.” MIGRATION AT A SNAIL’S PACE is repeated for each “step” – a form of Sporozoites drill through the skin locomotion also used by amoebas. until they reach a blood capillary. They How fast do cells actually move? “On In recent years some members of are then carried by the bloodstream to average, 30 micrometers an hour, but Spatz’s team have conducted research the liver, where they multiply for the they can reach a brisk 50 micrometers not only in Stuttgart but also on the first time. Until now it was a puzzle ex- an hour,” says Spatz with a chuckle. campus of the University of Heidelberg, actly how sporozoites move forward. They can travel about one millimeter a where word about their work on cell The only thing that was clear was that day, assuming they are fit enough. How- migration soon got around. One day they neither crawl nor paddle like bac-

ever, the speed of travel depends on the parasitologist Friedrich Frischknecht teria and single-cell algae, because they Photo: Wolfram Scheible

50 MaxPlanckResearch 1 | 17 BIOLOGY & MEDICINE_Biophysics

» Malaria pathogens move nearly 100 times faster than human cells.

form neither lamellipodia nor flagella. A comparison of various mutants, each they are connected by a meshwork of “Sporozoites glide gracefully without of which lacks a different protein, actin molecules. “The parasite causes changing their shape thanks to special yielded fresh insights. “We can now di- the membrane proteins to move to- proteins on their surface,” Kai Matu rectly observe what actually happens. ward the posterior end and then repels schewski explains. The scientist at the If protein X is missing, the parasite is them,” Matuschuweski explains. The Max Planck Institute for Infection Biol- unable to hold on; if Y is missing, it motion is similar to that of a millipede ogy in Berlin and professor at Hum- sticks fast to the surface,” Matuschew – except that these so-called feet con- boldt University is the third member of ski explains. sist only of single molecules and are so the project. small that they can’t even be seen un- The Plasmodium experiments didn’t TINY PROTEIN FEET der an electron microscope. The force require a non-slip cellwalk, but an ob- the parasite has to exert to hold and stacle course to imitate the parasite’s re- In its cell membrane, a sporozoite has then release itself can be measured lentless progress through the skin. a huge range of proteins that are re- with the help of optical tweezers. This Spatz constructed a miniature bed of quired for locomotion, some of which provided further insights into how the nails, of sorts, from polymers through perform identical functions. Hundreds single-cell organism moves. which the parasites can glide as if of them are situated at the anterior The malaria organism has a com- through a forest of ultrafine needles. end of the tiny cell body, to which plex life cycle, in the course of which

Below Malaria parasites (yellow) crawl over a surface studded with tiny columns. If the single-cell organisms have the same curvature as the columns, the pathogens, which measure just one hundredth of a millimeter in length, begin to circle around the obstacles. Right A migrating cell. It forms contact points with the surface at specially struc- tured sites located at the ends of thread-like processes (small image). At these points, it 250 mm senses the properties of the surface and uses

Photos and graphic: MPI for Intelligent Systems (2) the anchoring points to pull itself forward.

2 µm Photo: Wolfram Scheible

1 | 17 MaxPlanckResearch 51 Above Jacopo Di Russo, Katharina Quadt and Medhavi Vishwakarma (from left) preparing a new experiment in which they will modify and measure the movement of cell collectives. Below The protein merlin (green dots) controls the direction of travel of follower cells: 1 Without physical or chemical stimulation, merlin blocks the formation of lamellipodia. Without these protuberances, the cell is unable to move. 2 + 3 If, in contrast, the leader cell pulls on the cell behind it, merlin in both cells leaves its site on the cell membrane, allowing a lamellipodium to form. The follower cells are then able to follow the leader cells. The signal molecule Rac1 (red dots) must also be activated for a lamellipodium to form. The cells always migrate toward areas of higher concentrations of activated Rac1. 4 If Rac1 is inactivated (gray dots), merlin again blocks the formation of lamellipodia at the cell membrane.

1 Follower cell Leader cell it takes on various forms. However, ex- Whether it’s a small cut to a finger, a perts agree that the sporozoite is a scraped knee or deep cuts following an promising – if not the most promising operation, epithelial cells have to – target for a vaccine, for the sole rea- “rush” into the wound to close it and son that only around 100 pathogens begin rebuilding the tissue. This sounds Merlin Actin Inactive Rac1 2 are present immediately after an infec- simple, but it is a highly complex pro- Force tion. In the later stages of infection, cess. It has been compared to marching Lamellipodium there are billions. “Ideal vaccine can- in lockstep. Joachim Spatz calls it col- didates are antibodies that block two lective cell migration. It is one of the or three of the pathogen’s proteins processes that no longer function prop- Active Rac1 involved in movement,” Matuschew erly in chronic wounds. 3 ski says. If the sporozoite is unable to glide, it literally gets stuck in the MIGRATION UNDER skin, and the infection is stopped in A MICROSCOPE its tracks. Having studied the movements of The scientists needed a suitable wound 4 individual cells in detail, Spatz’s team model for their experiments. They cov- turned its attention to the migration ered the bottom of a petri dish with a behavior of entire groups of cells. They nutrient medium and grew epithelial posed the question: how exactly does a cells on it. They blocked a region of the

wound heal? dish to prevent the cells from migrat- Photo: Wolfram Scheible; graphic: MPI for Intelligent Systems

52 MaxPlanckResearch 1 | 17 BIOLOGY & MEDICINE_Biophysics

» Mechanical tension at tissue edges promotes wound healing.

ing into it. As soon as they removed the like snaps. At the same time, the actin blockade, the cells started to colonize skeleton in the interior of the cell the free space. Their movements were reaches out to neighboring cells for recorded by a microscope camera, stability. “The edge of a cell collective which took snapshots of the process ev- is sort of like a herd of sheep that has ery ten minutes. been penned behind a fence,” Spatz After just a few hours, bulges ap- says. The actin fence is under mechan- peared in the originally straight wound ical tension. If the curvature is the edges. “That’s because some cells move same everywhere, as in the case of a faster than others,” Spatz says. In oth- circle or a straight line, it’s difficult to er words, the repair crews don’t ad- break away from the herd. “Neighbor- vance in ordered lines. Rather, there are ing cells take off together and jockey leaders, “pioneering” cells that the oth- for the lead. Eventually, one prevails 100 µm ers trail along behind. “They form the and the others fall behind.” However, vanguard of the collective and can eas- a cell that marches out of the corner ily be identified because they’re bigger of a square, for example, has no neigh- and move ahead of the others.” bors and automatically becomes a Wound healing in a petri dish: Epithelial What makes a cell a leader? And leader. Surgeons unknowingly exploit skin cells migrate together into a region how do they communicate with the this effect when they make zigzag scal- with no cells. The colored paths show the cells’ movements over a period of more other cells in their tow? To answer pel cuts rather than straight ones, thus than five hours. these questions, the researchers varied stimulating subsequent cell migration their experiments. They covered the into the wound. bottom of the petri dishes with a hy- The researchers first measured the drogel, which cells find slippery, and ONE LEADS, THE COLLECTIVE speed and direction of motion of each sowed the epithelial cells within only FOLLOWS cell in a collective. “That showed us a small geometrical shape in the cen- that domains of 20 to 30 cells form that ter. As soon as the colonies filled these The researchers suspect that, at least in then quickly march together in one di- circular, triangular or square areas, the principle, every cell has leadership po- rection,” Spatz explains. These cells researchers briefly exposed the gel to tential. Whether they can exploit that form what is termed the persistence UV light. This transformed the slip- potential depends largely on their po- length, that is, the distance over which pery surface into a non-slip surface, sition at the edge of the wound. If the cells march in a coordinated manner in and the epithelial cells quickly set off. position is not crucial, for example if one direction. “We found that leader cells tend to the edge of the wound is straight, then Then the real work began. “In mo- form at places with a pronounced cur- the cell rows in the rear determine lecular cell biology, we know hundreds vature, for example in the corners,” which cell becomes the leader. Ulti- of proteins that serve as signal mole- Spatz says. This is easily explained: the mately, the decision regarding the lead- cules for cell migration,” Spatz says. contacts between the cells are me- ership of a collective is not made inde- “We switched off the gene for each mi- chanically stabilized by what are pendently by the leader cell, but by the gration-associated protein so that the known as tight junctions – short struts collective as a whole. This decision is cell was unable to synthesize that pro- of membrane proteins such as cadher- physically regulated in the collective by tein. Then we measured the persistence ins. If the cadherins of cell A match purely mechanical means. The ques- length of the mutants by biophysical those of cell B, the extracellular do- tion that remains is how the leader means.” The scientists were amazed to Photo: Wolfram Scheible; graphic: MPI for Intelligent Systems Photo: MPI for Intelligent Systems mains of those proteins click together communicates with its followers. discover that very few of the proteins

1 | 17 MaxPlanckResearch 53 Medhavi Vishwakarma, Freddy Frischknecht, Joachim Spatz, Jacopo di Russo and Katharina Quadt (from left) analyzing data. The scientists have established that the protein merlin plays a key role in the collective migration of cells.

have anything to do with collective found that the protein leaves its origi- substances are released that convey di- movement. Only if the membrane pro- nal location to reappear in the interior rectional information to the cells along tein merlin is missing does the collec- of the cell. But why? They hypothesize the wound’s edge.” Spatz’s team, togeth- Profiles of Leading Women Scientists tive break apart. that the leader cell rushing ahead cre- er with the University of Heidelberg, has It is astonishing that the cell relies ates tension between the cells. “It’s like since repeated and confirmed the exper- on just a single type of molecule and a runner pulling another runner be- iments on human skin models. The project doesn’t have at least one backup pro- hind him by the hand,” Spatz explains. tein at the ready. Merlin is not a new Merlin is like a force sensor that re- UNCOORDINATED MOTION AcademiaNet is a database containing the discovery – it was already known from sponds to this tension. It disappears WITHOUT MERLIN profiles of over 2,200 outstanding women cancer biology. Unlike in the case of into the interior of the cell, leaving wound healing, a metastatic growth room on the membrane for the follow- The researchers also discovered that, researchers from all disciplines. wants to prevent the cellular “herd in- er cell to extend its lamellipodium and without merlin, more leader cells form, stinct” because single cancer cells are follow the leader. “If merlin didn’t do but the wound healing proceeds more Our objective more efficient and can progress into tis- that, the cell would be unable to follow, slowly, because the mechanism by sue more quickly than a group. They let because membrane-bound merlin acts which the cell collective moves is no The aim of our search portal is to make it the bloodstream carry them to distant essentially as a lamellipodium brake.” longer coordinated. easier to find female academics to fill leading regions of the body, where they estab- And because the protein makes room Spatz is currently also looking more lish themselves at suitable locations only at places where the leader cell closely at the movement of cancer cells. positions and to sit on executive committees. and begin to divide uncontrollably. pulls, the direction is automatically es- “Unlike healthy cells, they have a ten- “Merlin is a metastasis inhibitor,” Spatz tablished. This process continues down dency to ignore their environment. explains. “It’s a good sign when the the rows of the cell collective – literal- That’s bad for patients, but good for the protein is very active in cancer cells. ly pull by pull. So mechanical tension cancer!” Metastatic cancer cells reduce The partners The cells then tend to stay together and promotes the formation of cell collec- not only their merlin production, but are thus less likely to form metastases.” tives and thus wound healing. also that of cadherin contact proteins. It was already known that merlin is And how does the lead cell know They are therefore able to make them- positioned between the cell membrane which direction it must go in? “In our selves extremely long and narrow and and the cellular skeleton. The research- model, it can move in only one direc- slip between tissue cells to invade new ers observed collective cell migration tion, namely into the free space. In the regions of the body without being de-

after labeling merlin with a dye, and case of a real wound, however, signaling tected or stopped by other cells. Photo: Wolfram Scheible Would you like to find out more? www.academia-net.de 54 MaxPlanckResearch 1 | 17 BIOLOGY & MEDICINE_Biophysics

However, cell migration is not specific to wounds and cancer cells. It occurs TO THE POINT continuously everywhere in the body. l Membrane-bound proteins convey information about a cell’s environment, “If cells did not cluster together, com- such as the nature of the substrate. By remodeling its internal skeleton, the cell municate with each other and migrate can form small protuberances in the membrane, which it uses to move about. together, we wouldn’t exist. The migra- l When epithelial cells migrate into an uncolonized area, some of them become leader cells that the others follow. Such leader cells tend to develop at sites tion and interaction behavior of cells where the tissue edge has a pronounced curvature, for example at corners was the prerequisite for the evolution and edges.

of multicellular organisms – and thus l Merlin, a membrane protein, coordinates groups of migrating cells. It senses for life as we know it.” the physical tension that arises when a leader cell pulls on the follower cells, Communication with the environ- enabling the formation of small protuberances at the pulling points. The protu- ment is the lifeblood of cells, some- berances are then used for locomotion. thing they have in common with us, their one-hundred-trillion-cell collectives. What happens to people when GLOSSARY communication is cut off for extend- ed periods? They become lonely. Some Lamellipodia: Broad protuberances of the membrane at the anterior end of motile cells. They are also known as pseudopodia. A two-dimensional meshwork of thread-like actin may become despondent, while others proteins provides stability and tensile force. Lamellipodia are used for locomotion and might find that they are sufficient navigation, and for engulfing nutrient particles (phagocytosis). unto themselves. Yet others would Optical tweezers: A method that allows scientists to measure the forces that individual seek out old acquaintances or establish molecules exert on each other. Each of the molecules to be measured is placed on a new ones. Cells, in contrast, have no plastic bead that is held in position by two laser beams. As soon as the molecules start choice: they activate a built-in suicide to interact and attract each other, sensors measure the force that the lasers must exert to hold the beads apart. program and die.

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Our objective The aim of our search portal is to make it easier to ﬁnd female academics to ﬁll leading positions and to sit on executive committees.

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