Understanding protamine-induced DNA folding in sperm

We examine how protamine causes DNA folding within the sperm nucleus. Specifically, we are interested in how protamine loops the DNA into toroids that allow for tight packaging of the paternal genome. The formation of toroids have implications in epigenetics and fertility. Toroids may be formed in a single step (collapse model) or multistep (sequential model) manner. In the collapse model, all loops form along the DNA at once and then collapse into the toroid. In the sequential model, loops stack sequentially into a toroid. To decipher between these two models, we perform an in vitro Tethered Particle Motion (TPM) assay. In a TPM assay, an individual DNA molecule is attached to a bead and tethered to a cover-slip. By observing the position of the bead over time, we can measure the DNA length to a precision of about 10 nm and infer the folding trajectory of our DNA. We observe that as the length of the DNA increases, the number of folding events increases, suggesting that toroid formation is multistep (sequential model). If DNA folding is multistep, the location of nucleation loops is important as they may indicate where histones, and thus epigenetic tags, may be present. Moreover, the ratio of different protamine proteins may affect how toroids form and play a role in fertility.