Biology subjects

Tuzel, E.

Publications and source records attributed to Tuzel, E..

5 recordsLinked to original sources

bioRxiv2024

DACIT: Device for Axon - Cancer cell Interaction Testing in 2D and 3D

There is increasing interest in studying the role of peripheral innervation in tumor growth and metastasis. However, in vitro studies of interactions between cancer cells and axonal projections are technically challenging. To address this, we have developed a microfluidic Device for Axon-Cancer cell Interaction Testing in 2D and 3D (DACIT). We show that DACIT successfully separates neuronal soma from the axons and cancer cells into two compartments, which can be exposed to similar, or different growth conditions, depending on the experimental needs. We compare neoaxonogenesis using either the PC-12 cell line or primary embryonic or adult sensory neurons, demonstrating superior neurite growth in primary cells. Additionally, we show that DACIT can accommodate assessing growth and 3D invasion of tumor spheroids, due to its unique height profile. Hence, DACIT can be used to analyze cancer cell interactions with axons in most typical cell biology assays such as proliferation, invasion, and calcium activity which we demonstrate on examples of imaging transients in GCaMP6-labeled neurons, invadopodia assay, and 3D cancer spheroid invasion.

cancer biology↗

bioRxiv2021

Invadopodia enable cooperative invasion and metastasis of breast cancer cells

Invasive and non-invasive cancer cells can invade together during the cooperative invasion. However, the events leading to it, role of EMT and the consequences this may have on metastasis are unknown. In this study, we demonstrate that the isogenic 4T1 and 67NR breast cancer cells sort from each other in 3D spheroids, followed by cooperative invasion. By time-lapse microscopy, we show that the invasive 4T1 cells move more persistently compared to non-invasive 67NR, sorting and accumulating at the spheroid-ECM interface, a process dependent on cell-ECM adhesions and independent from E-cadherin cell-cell adhesions. Elimination of invadopodia in 4T1 cells blocks invasion, demonstrating that invadopodia requirement is limited to leader cells. Importantly, we demonstrate that cells with and without invadopodia can also engage in cooperative metastasis in preclinical mouse models. Altogether, our results suggest that a small number of cells with invadopodia can drive the metastasis of heterogeneous cell clusters.

cancer biology↗

bioRxiv2020

Myosin XI drives polarized growth by vesicle clustering and local enrichment of F-actin in Physcomitrium (Physcomitrella) patens

In tip-growing plant cells, growth results from myosin XI and F-actin mediated deposition of cell wall polysaccharides contained in secretory vesicles. Previous evidence showed that myosin XI anticipates F-actin accumulation at the cells tip, suggesting a mechanism where vesicle clustering via myosin XI promotes F-actin polymerization. To evaluate this model, we used a conditional loss-of-function strategy by generating Physcomitrium (Physcomitrella) patens plants harboring a myosin XI temperature-sensitive allele. We found that loss of myosin XI function alters tip cell morphology, vacuolar homeostasis, and cell viability, but not following F-actin depolymerization. Importantly, our conditional loss-of-function analysis shows that myosin XI clusters and directs vesicles at the tip of the cell, which induces F-actin polymerization, increasing F-actins local concentration. Our findings support the role of myosin XI in vesicle clustering and F-actin organization, necessary for tip growth, and deepen our understanding of additional myosin XI functions.

cell biology↗

bioRxiv2019

In vivo Interactions between Myosin XI, Vesicles, and Filamentous Actin Are Fast and Transient

The apical actin cytoskeleton and active membrane trafficking machinery are essential in driving polarized cell growth. To better understand the interactions between myosin XI, vesicles, and actin filament in vivo, we performed Fluorescence Recovery After Photobleaching (FRAP) and showed that the dynamics of myosin XIa at the tip are actin-dependent and that approximately half of myosin XI is bound to vesicles in the cell. To obtain single particle information, we used Variable Angle Epifluorescence Microscopy (VAEM) in Physcomitrella patens protoplasts to demonstrate that myosin XIa and VAMP72-labeled vesicles localize in time and space for periods lasting only a few seconds. Using tracking data with Hidden Markov Modeling (HMM), we showed that myosin XIa and VAMP72-labeled vesicles exhibit short runs of actin-dependent directed transport. We also found that the interaction of myosin XI with vesicles is short lived. Together, this bound fraction, fast off-rate, and short run lengths are expected to be critical for the dynamic oscillations observed at the cell apex, and may be vital for the regulation and recycling of the exocytosis machinery; while simultaneously promoting the vesicle focusing and secretion at the tip, necessary for cell wall expansion.

cell biology↗

bioRxiv2018

Measurement of the Persistence Length of Cytoskeletal Filaments using Curvature Distributions

Cytoskeletal filaments such as microtubules and actin filaments play important roles in the mechanical integrity of cells and the ability of cells to respond to their environment. Measuring the mechanical properties of cytoskeletal structures is crucial for gaining insight into intracellular mechanical stresses and their role in regulating cellular processes. One of the ways to characterize these mechanical properties is by measuring their persistence length, the average length over which filaments stay straight. There are several approaches in the literature for measuring filament deformations, including Fourier analysis of images obtained using fluorescence microscopy. Here, we show how curvature distributions can be used as an alternative tool to quantify bio-filament deformations, and investigate how the apparent stiffness of filaments depends on the resolution and noise of the imaging system. We present analytical calculations of the scaling curvature distributions as a function of filament discretization, and test our predictions by comparing Monte Carlo simulations to results from existing techniques. We also apply our approach to microtubules and actin filaments obtained from in vitro gliding assay experiments with high densities of non-functional motors, and calculate the persistence length of these filaments. The presented curvature analysis is significantly more accurate compared to existing approaches for small data sets, and can be readily applied to both in vitro or in vivo filament data through the use of an ImageJ plugin we provide.

biophysics↗