Protamine-induced toroid formation in sperm.

Abstract

In sperm cells, DNA is very tightly condensed into a toroid, which is a near crystalline state. If we can understand how DNA is condensed in sperm cells, this would give insights into the physical mechanism for DNA folding at this fundamental limit. Previous work in our lab shows how protamine binds DNA and bends it into a loop. Here we want to establish how these loops stack and eventually come to form toroids in vitro. To achieve this, we used atomic force microscopy (AFM) to image DNA at increasing protamine concentrations (0.1-3 micromolar) and used a Matlab program, Easyworm, to quantify the images. This allows us to measure changes in the DNA structure as the DNA-protamine complex folds. To start, we adhered DNA that was ∼3000 bp long onto a mica slide and then added protamine to induce DNA folding. We imaged structures with many folded loops, structures with stacks of loops, and long cylindrical structures that appear to be coils of loops. More work is needed to determine the mechanism for folding and how these structures fit into the toroid formation pathway.