Patterning the Cytoplasm to Generate Protein Gradients

Last Thursday, Erik Griffin, a post-doctoral fellow at Johns Hopkins University, spoke at a Faculty Candidate Seminar hosted by the Dartmouth biological sciences department. Griffin focused on his research regarding the formation of protein gradients in embryological cells.

While the ability of mother cells to asymmetrically localize certain traits between daughter cells has been well documented, Griffin hoped to identify the mechanism that leads to asymmetrical protein distribution. Griffin’s presentation dealt exclusively with two specific proteins in the cell: MEX-5 and PIE-1.

Griffin’s use of embryological cells from C. elegans, a species of nematode, proved effective because the cells are large enough to be easily imaged, allowing him to watch the process of cell polarization as it occurred. Polarization in the embryological cells of C. elegans is important because it allows each mother cell to give rise to two daughter cells with distinct adult functions. MEX-5 and PIE-1 gradients play a significant role in determining cell fate and establishing cell diversity.

In the context of Griffin’s experiment, cell polarization means that one daughter cell forms with a high concentration of MEX-5 and a low concentration of PIE-1. The opposite is true of the second daughter cell. Cells with high concentrations of MEX-5 develop into soma (body cells), while those with high concentrations of PIE-1 are destined to be germoid (sperm and egg) cells designated for reproduction in adult form.

Griffin used “pulse-charge” imaging techniques to show that new synthesis of these proteins in different cell regions does not affect relative concentrations after cell division has begun. Rather, MEX-5 and PIE-1 proteins are redistributed to the anterior and posterior of the mother cell, respectively. Griffin’s primary challenge was to identify the means of redistribution within the mother cell.

Griffin observed that the rate of diffusion of MEX-5 through the cytoplasm was much faster in the posterior than the anterior of the cell. He hypothesized that PAR-1 proteins in the posterior of the cell phosphorylate MEX-5 molecules to speed up their diffusion. Phosphorylated MEX-5 molecules diffuse more rapidly because they exhibit a decreased affinity for RNA molecules in the cytoplasm. MEX-5 diffusion is relatively slow in the anterior of the cell because the molecules get “caught” on RNA molecules in the cytoplasm.

Just as important as the PAR-1 proteins in establishing MEX-5 gradients in the cell, however, are PP2A phosphatases that perform the reverse functions and slow down MEX-5 diffusion. Griffin found that these phosphatases are distributed relatively equally throughout the cell. On the other hand, he found that PAR-1 molecules formed a gradient opposite that of the MEX-5 gradient in the cell to help speed the process of polarization.

Ultimately, the established MEX-5 gradient across the mother cell leads to the formation of the PIE-1 gradient. The PIE-1 gradient that is formed is “stronger” than the MEX-5 gradient, meaning the relative concentration difference between posterior and anterior in the mother cell is higher for PIE-1 than for MEX-5. In the near future, Griffin hopes to better understand why this is true, as well as how RNA binding proteins like MEX-5 can gather and transmit information to create other protein gradients in the cell.