Generating Force at the Leading Edge

Last week I saw an awesome lecture by Gaudenz Danuser who has a lab at the Scripps institute in San Diego. It has taken me a week to fully digest what was said, plus I haven't had the time to jot this down.

Over the past few years the Danuser lab along with Claire Waterman-Storer's group (see this post and then this post) have used speckle microscopy in order to figure out how the actin cytoskeleton is reorganized during migration. In the process they've really made quite a bit of headway in figuring out how cells crawl.

Actin is just cool. It is the main polymer responsible for shaping the cell's morphology. It forms polymers that are constantly assembling and disassembling in the cell. This perpetual remodeling of the actin network combined with the action of motors, which pull on the actin filaments, is responsible for generating much of the force that helps cells contract, stretch and change shape.

Another important item that you should know are focal adhesions. These are large aggregates of proteins that help connect the cell to extra-cellular fibers, such as collagen. Focal adhesions contain receptors for the extra-cellular fibers, signaling molecules that help coordinate the cell's shape and behavior, and actin binding proteins. Thus there is a direct protein connection between fibers on the outside of the cell and the actin meshwork that shapes the inside of the cell.

Now I'll refrain from giving any details about Danuser's unpublished data, but I will say a couple of general things regarding his most recent work, which dissects the coordination between actin, cell-substratum attachments and G-proteins in migrating cells.

(A warning, for those not fully versed in the cytoskeleton & cell signalling, the following paragraphs will have lots of jargon that may be hard to follow .)

1) At the leading edge, different sub-domains of the periphery will transiently spread and then retract like waves hitting a shoreline. Thus each small region takes two steps forward, then one step back. Over time this pulse like expansion, interspersed by small elastic-collapses of the overstretched membrane, results in forward movement.

By imaging this whole cycle of membrane expansion and collapse in small regions of the cell periphery, and correlating the exact timing of these events with changes in the actin cytoskeleton, focal adhesion dynamics and G-protein activation state, the Gaudenz lab in collaboration with Waterman-Storer (and Klause Han Hahn's?) lab(s) were able to build a model of how changes in actin produce forward movement.

The model: During the initial membrane expansion, certain actin structures are assemble in response to one G-protein. Then as the overstretched membrane starts to collapse, the first G-protein is turned off and a second G protein becomes activated. This new signaling molecule stimulates the assembly of a second type of actin network that stabilizes the fragile cell protrusion and helps to stop total membrane collapse. As the membrane applies pressure to the actin meshwork, actin filaments are driven rearwards (as seen above, in the speckle microscopy movie of actin at the front of a migrating cell.) This backward force is opposed by the focal adhesions that are anchored both to the actin meshwork and to the extra-cellular environment (or "matrix" as it is commonly called). The activation and inhibition of the G-proteins is probably mediated by the formation of new focal adhesion within the new protrusions. Remember that these adhesions are full of signaling molecules many of which can affect the activity of G-proteins (sounds like stuff that I've personally published!)

2) This talk was one of the best "System biology" talks that I've seen. In fact it was so good that I was simultaneously happy and depressed ... and happy. Happy because the model proposed by Gaudenz, is the best explanation that I've seen for force generation at the leading edge. In addition, the data presented validate a prediction that came out of my thesis advisor's lab. (To be honest others have also measured that RhoA is active at the leading edge and not the rear of crawling cells.) But I was simultaneously depressed. Gaudenz's work has really raised the bar. If you want to make some serious progress in the cytoskeletal field, you'll need to know a lot of biology that you can only get from hands-on experience AND you'll need a good foundation in mathematics, statistics and modeling. Thus if systems biology is going to contribute to our knowledge of cell biology it will be through individuals such as Gaudenz who both understand the intricacies of the biological process (in this case the cytoskeleton and G-proteins) while being adept at developing new statistical analysis methods in order to figure out how the system is built. But I was also happy because I'm no longer in the cytoskeletal field. It would seem to me that our knowledge of how cells handle RNA is much more primitive then how G-proteins, actin, focal adhesion and membrane states are coordinated to regulate cellular morphology.