How Do Cells Know Up From Down?

Erin Cline
Department of Molecular & Cellular Physiology
Stanford University School of Medicine
December 2001

The research in my biology laboratory concentrates on understanding how cells' interactions with each other give them directional information. This directional information allows cells to set up tissues that create a barrier between the inside and outside of the animal. Essentially for the cells this means separating "up" from "down". We call this cell polarity. I work on a set of proteins called "Pars" that has been found to be important in this process. Research like mine that deals with cell polarity is important because numerous diseases, like cancer, involve cells that have lost the ability to tell up from down.

The interactions of cells with each other and their environment cause multiple changes to occur within them. Proteins move to different locations and the cytoskeleton - literally the skeleton of the cell that gives it structure just like our skeletons give us structure - changes its shape and orientation. These changes start a cycle where changes in the cell shape cause changes in the location of proteins within the cell, and then these changes in protein location cause changes in the cell shape. Eventually all of these changes that were initiated by the interaction of the cells with things outside of themselves, cause the cell to assume its proper orientation which is needed for its designated role in the animal. For example, cells in our kidneys have their "upper side" facing our blood flow in order to filter out wastes. This important polarity is set up when cells that will eventually become our kidneys interact while we develop before we are born.

I use imaging techniques that allow me to see proteins within cells and biochemistry techniques that analyze them directly to investigate the Pars. These proteins are necessary for correct cell polarity in cells from all types of organisms. I hope to understand what these proteins are doing to generate cell polarity. I also hope to understand the pathway that leads from cells' interactions with other cells to the location and activity of these proteins. My approach to understanding these questions is to look at where these proteins are in cells, when and how they get there, and with what other proteins they are associated.

The imaging techniques I use rely on antibodies. These are the same kind of antibodies that are made in our bodies to help our immune systems fight off disease. The antibodies I use are from animals like mice or rabbits. We send a sample of the protein we want to study to a company that injects the animals with the protein. They then send us blood samples from the animals. These blood samples contain antibodies to our proteins because when it was injected into the animals, the animals' immune systems saw the protein as an invader that it wanted to fight off. We then use these antibodies to find our protein of interest in actual cells. We kill the cells we are studying and add the antibody, which clamps on to our protein wherever it is in the cell. We then use other antibodies that are hooked to fluorescent dyes to clamp onto the first antibodies we added to the cells. Finally, we look at the cells under a microscope that allows us to see fluorescent dyes. This whole process allows us to see where in the cell our protein of interest is located, and therefore what the protein might be doing inside of the cell

The biochemistry techniques I use investigate the Pars once I have taken all of the proteins out of the cells. I use gels, which are polymers that form a lattice-work with little holes in it, to separate the PAR proteins from each other and from other proteins in cells on the basis of size. Doing this in combination with some other techniques allows me to see what proteins interact with the PAR proteins. This is important because this helps me to understand the role of the PAR proteins in the context of all the other proteins in the cell. Depending on how I set up the experiment, I can also investigate where the PAR proteins are in the cell.

With our imaging and biochemical studies of the Pars we hope to contribute to the field of biology by showing how these proteins are involved in cell polarity. This knowledge combined with that of other researchers will give all biologists a better understanding of how cell organize themselves.