Cook Lab website
Timely ubiquitin-mediated protein degradation is fundamental to cell cycle control, but the precise degradation order at each cell cycle phase transition is still unclear. We investigated the degradation order among substrates of a single human ubiquitylating enzyme, and discovered a consistent order of destruction during both S-phase entry and DNA repair. This order is attributable to the enzyme targeting motif known as a degron. Early or late degradation kinetics are transferably via the degron and are associated with differences in affinity for both the enzyme and an essential targeting cofactor. Importantly, cells with an artificially manipulated degradation order display evidence of stalled replication in mid-S phase and sensitivity to replication arrest. We propose that sequential degradation ensures orderly S-phase progression to avoid replication stress and genome instability.
Ohio State University
772 Biosciences Building
Columbus, OH 43210
Living organisms rely on macroscopic and microscopic structures that produce and transform force to survive: from cell volume regulation to sound transduction, handling of forces is essential to life. I will present our studies on three protein systems involved in force transduction. First, I will describe simulations of MscS, a mechanosensitive ion channel gated by membrane tension that acts as a safety valve to protect bacterial cells from osmotic shocks. These simulations established the conduction state of the first MscS crystal structure, triggered the search for wider, truly open states, and proposed a general mechanism for lipid-mediated channel gating. I will also show our work on the elasticity of ankyrin repeats present in TRP channels thought to be involved in mechanosensation. Steered molecular dynamics simulations of long ankyrin repeat domains showed that they extend and straighten reversibly and without secondary structure modifications in response to low force. We called this "tertiary structure elasticity". Stepwise unfolding of repeats was predicted to occur at larger forces and subsequently confirmed by experiments suggesting that ankyrin repeats can act as gating springs with a built-in safety mechanism. Finally, I will present our simulations of classical cadherin repeats involved in calcium-dependent cell-cell adhesion and mechanotransduction. These simulations showed how calcium induces straightening and rigidification of tandem repeats to favor binding to other cadherins, how it prevents mechanical unfolding to protect cadherins from large forces, and how it regulates the availability of key tryptophan residues involved in bond formation. Overall, these simulations have shed light on the molecular mechanisms of mechanosensation.