Senior Honors Projects, 2020-current

Creative Commons License

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 License.

Date of Graduation

5-9-2020

Document Type

Thesis

Degree Name

Bachelor of Science (BS)

Department

Department of Engineering

Advisor(s)

Callie Miller

Abstract

F-actin networks have different structures throughout the cell depending on their location or mechanical role. For example, at the leading edge of a migrating cell, F-actin is organized in a region called the lamellipodia as a branched network responsible for pushing the membrane outwards. Behind the lamellipodia is a lamellar actin network where focal adhesions and stress fibers originate, and then within the cell cortex, actin is arranged in a gel-like network. Stress fibers are an important organization of F-actin and how they arise from either the branched lamellipodia network or the gel-like cortex network is poorly understood. Our approach is to create a computational simulation to model a mechanism of bundling by crosslinking proteins specifically modeling the interaction of two cylindrical rods (individual F-actin filaments). We created a 2D simulation of two cylindrical rods in MATLAB that contact one another. We specified the initial speeds of the rods, and plotted their location after impact based on the principles of linear momentum. Although this model demonstrated how parallel filaments might collide, we also considered how actin-binding proteins work together to form tightly bundled F-actin stress fibers, arising from the branched network of F-actin at the leading edge. Our approach is two-fold: first, we conducted biochemical studies focused on the interactions of actin filaments and the actin binding proteins ARP2/3, alpha-actinin, dynamin2, and cortactin and second, we developed a computational model to validate experimental hypotheses for actin filament crosslinking to promote filament bundling. Our biochemical studies used quantitative binding analysis and imaged reconstituted networks in real time to show that dynamin2 and cortactin create bundled actin filaments in vitro. Additionally, cortactin decreases the association of alpha-actinin with bundled actin filaments. We hypothesize that the transition from a branched to bundled F-actin network architecture involves balancing the activities of two competing filament crosslinkers: dynamin2 and cortactin vs. alpha-actinin. We will test this hypothesis by creating an emergent Monte Carlo model in MATLAB that simulates filament crosslinking in the presence of alpha-actinin, cortactin and dynamin2. Our model lays the foundation for testing hypotheses of the mechanisms by which cortactin and dynamin2 organize F-actin networks.

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