Adriana Dawes, The Ohio State University
Hosted by Arpita Upadhyaya
Title: Pushing and pulling to properly position centrosomes in polarized cells
Time: 4:00PM - 5:00PM
Date: Monday January 30, 2017

Abstract

Asymmetric cell division, where daughter cells inherit unequal amounts of specific factors, is critical for development and cell fate specification. Asymmetric cell division occurs in polarized cells as a result of positioning the centrosomes along the polarity axis. Using an individual-based stochastic model of microtubule dynamics and experiments in the early C. elegans embryo, we explore potential sources of cortical force generation and demonstrate the need for both cortical and centrosomal asymmetries for recapitulating the in vivo dynamics and proper positioning of the centrosomes.

Ursula Perez-Salas, University of Illinois at Chicago
Hosted by Silvina Matysiak
Title: Lipids in Motion and the Energy to Get them "there"
Time: 4:00 - 5:00PM
Date: Monday February 20, 2017

Abstract

Lipids are amphiphilic molecules that love and hate water simultaneously and self-assemble into a lipid bilayer that is a universal structure of all cell membranes. The membrane’s hydrophobic interior is a 4nm-thick film that separates the interior of the cells from the surrounding environment. Structures inside the cell, such as the nucleus, evolved to be bounded by membranes in eukaryote cells; in fact, it is the hallmark that sets them apart from prokaryotic cells. Hence, biological membranes are not only life/death boundaries for cells but are also necessary for the existence of higher organisms like us. However, the mere function of “confining barrier” does not explain why eukaryotic cells invest substantial resources in generating thousands of different lipids (~5% of their genes!) to form membranes. Indeed, the lipid composition of individual membranes within a cell varies significantly. Consider the case of cholesterol: the endoplasmic reticulum, where it is synthesized, contains barely 1% of the total cell cholesterol, while the plasma membrane contains about 40%. So the question arises: how do cells create these unique compositions and multiple functionalities in their individual membranes? The answer to this question is explained in part by thermally-driven properties of lipid membranes. In this talk I will present results from our ongoing efforts to map the energy landscape of lipid motion between distinct membranes and within a single membrane. These recent efforts concentrate primarily on neutron scattering techniques that, I will argue, are highly desirable because other probing techniques potentially can be too invasive, resulting in skewed results.

Tae Yoon Kim, Purdue University
Hosted by Garegin Papoian
Title: Reconstructing the Mechanical Behaviors of Cells in Silico
Time: 4:00 - 5:00PM
Date: Monday March 6, 2017

Abstract

Actin cytoskeleton is a dynamic structural scaffold used by eukaryotic cells to provide mechanical integrity and resistance to deformation, while simultaneously remodeling itself and adapting to diverse extracellular stimuli. The actin cytoskeleton utilizes these properties to play crucial roles in essential cellular processes such as cell migration and division. However, despite its known mechanical role in cell behaviors, a clear understanding of the mechanical properties of actin cytoskeleton and the molecular origin of these properties still lacks, partly due to experimental limitations. Computer simulations can access time and length scales inaccessible by experiments, and thus aid in creating a descriptive model of the molecular interactions that evolve into the mechanical properties observed on cellular scales. To this end, we have developed a cutting-edge computational model which is designed to reproduce the mechanical and dynamic behaviors of actin cytoskeleton within cells. Guided by explicit experimental data, we systematically explored, via simulation, how the mechanics and dynamics of actins and actin-binding proteins determine the deformation, flow, and stiffness of the passive actin cytoskeleton. We also investigated how interactions between the passive cytoskeletal constituents and active molecular motors lead to force generation, contraction, and morphological changes in the active actin cytoskeleton.

Rodrigo Maillard, Georgetown University
Hosted by Christopher Jarzynski
Title: Investigating the Conformational Landscape of Protein Kinase A with Single Molecule Optical Tweezers
Time: 4:00 - 5:00PM
Date: Monday March 27, 2017

Abstract

Signaling proteins are dynamic macromolecular complexes that sample multiple conformational states. Such conformational plasticity allows these proteins to adapt and respond to different biological signals. Protein Kinase A (PKA) is a signaling protein that oscillates between inactive and active conformations depending on cAMP concentration. In this study, we use single molecule optical tweezers to dissect the pathways of communication between PKA domains that enable the progression from inactive to active conformations. We show that cAMP triggers networks of communication between the two cAMP binding domains of the PKA regulatory subunit that involve direct, interfacial domain contacts as well as long-range interactions between non-contiguous structural motifs. In contrast, without ligand the two cAMP binding domains behave as independent, non-interacting structures, illustrating how cAMP turns on and off domain communication networks. The selective mechanical manipulation of the regulatory subunit bound to the catalytic subunit reveal that the inactive PKA holoenzyme is in a dynamic equilibrium between two conformational states, wherein the two cAMP binding domains establish different sets of interactions with the catalytic subunit. By changing the pulling axis in the regulatory subunit, we identify an energetic hub in the regulatory subunit whose mechanical perturbation triggers the highly cooperative and coordinated dissociation of the PKA holoenzyme. Altogether, our results show how this signaling complex propagate ligand binding signals throughout the protein structure to regulate protein function. Our experimental approach based on optical tweezers should be readily applicable to dissect domain communication networks in other signaling proteins.

Stavroula Mili, NIH-NCI
Hosted by Helim Aranda-Espinoza
Title: RNAs at Cell Protrusions: Regulation by the Extracellular Environment and Functional Effects
Time: 4:00 - 5:00PM
Date: Monday April 3, 2017

Abstract

Dynamic formation and extension of protrusions is intimately associated with the process of cell migration. Cells exhibit a variety of protrusions that can be utilized during distinct modes of migration and in different extracellular environments. Local RNA translation is emerging as a mechanism required to stabilize protrusions and promote persistence and directionality during cell movement. Numerous RNAs are known to be enriched within protrusive regions. However, whether all protrusion-localized RNAs are coordinately regulated and how they respond to the properties of the extracellular matrix are significant, unanswered questions. I will discuss evidence showing that RNAs localized in protrusions of migrating fibroblasts can be distinguished in at least two groups, which are differentially enriched in high- or low-contractility protrusions, and are additionally differentially dependent on the Adenomatous Polyposis Coli (APC) tumor-suppressor. APC-dependent RNAs become enriched in high-contractility protrusions and, accordingly, their localization is promoted by increasing stiffness of the extracellular matrix. The underlying mechanism involves activation a RhoA-mDia1 signaling pathway that leads to formation of a detyrosinated-microtubule network, which in turn is required for localization of APC-dependent RNAs. Importantly, a competition-based approach to specifically and globally mislocalize APC-dependent RNAs, indicates that localization of the APC-dependent RNA subgroup is functionally important for cell migration both on 2D and 3D substrates. These studies provide new links between the extracellular environment, cellular behavior and regulation of RNA localization.

Imran Rizvi, Harvard Medical School
Hosted by Giuliano Scarcelli
Title: Biophysics in Cancer: Targeting Chemoresistance with Rationally-designed Combination Therapies in Bioengineered 3D Models
Time: 4:00 - 5:00PM
Date: Monday April 17, 2017

Abstract

Tumor heterogeneity and drug resistance to conventional therapies remain major causes of treatment failure, recurrence and dismal survival rates for patients with advanced stage cancers. A range of cellular, architectural, and physical cues in the tumor microenvironment influence the intrinsic and acquired resistance mechanisms that lead to treatment failure. These cues include physical forces such as hydrodynamic shear stress and communication with heterocellular stromal partners, which remain understudied as determinants of tumor heterogeneity and variability in treatment response. Strategies that leverage photodynamic therapy (PDT) a photochemistry-based biophysical treatment modality to regionally target and prime stubborn tumor populations may be essential to overcoming key barriers to durable cancer management while minimizing toxicity from traditional agents. A multi-faceted approach is needed to evaluate and optimize PDT-based combination therapies, including the development of bioengineered 3D models that integrate cues such as physical forces and heterotypic cellular communication. Here the impact of hydrodynamic stress and stromal biology is evaluated in the context of ovarian cancer (OvCa). Metastatic OvCa spreads predominantly via flushing of ascites along preferential fluidic pathways and communicates with local microenvironment, including the extracellular matrix (ECM) and TECs, to form peritoneal implants. A microfluidic model that supports 3D tumor growth was developed to investigate the role of fluidic stress on the heterogeneity of metastatic OvCa. The motivation for this study was based on clinical observations that the most stubborn tumors are often found in regions such as the peritoneal gutter, a common site of resistance and recurrence, and a region that is subjected to fluidic stress from ascites. Tumor nodules cultured under flow showed increased epithelialmesenchymal transition (EMT) compared to non-flow 3D cultures. Molecular and morphological changes consistent with EMT included a transcriptionally-regulated significant decrease in E-cadherin, a significant increase in vimentin, and significant decrease in fractal dimension, a metric adapted to quantify spindle-like morphology. A concomitant significant post-translational upregulation of epidermal growth factor receptor (EGFR) expression and activation was seen under flow. The impact of heterotypic communication between TECs and OvCa cells was investigated in a 3D model. Tumors grown in the presence of TECs were differentially susceptible to chemotherapy and benzoporphyrin derivative (BPD)-based PDT and showed increased heterogeneity in treatment response in the presence of endothelial cells. The potential value of using bioengineered models to guide customized, rationally-designed PDT-based combination regimens will be presented.

Michael Murrell, Yale University
Hosted by Wolfgang Losert
Title: Mechanical Force Production in a Biomimetic Cell Cytoskeleton
Time: 4:00 - 5:00PM
Date: Monday April 24, 2017

Abstract

While the molecular interactions between myosin motors and Factin are well known, the relationship between Factin organization and myosin-mediated force generation remains poorly understood. Here, we explore the accumulation of myosin-induced stresses within a 2D biomimetic model of the actomyosin cortex, where myosin activity is controlled spatially and temporally using light. By controlling the geometry and the duration of myosin activation, we show that contraction of disordered actomyosin is highly cooperative, telescopic with the activation area and generates a pattern of mechanical stresses consistent with those observed in contractile cells. We quantitatively reproduce these properties using an in vitro isotropic model of the actomyosin cytoskeleton, and explore the physical origins of telescopic contractility in disordered networks using agent-based simulations.

Eva Nogales (Marker Lecturer), University of California Berkeley, HHMI
Hosted by Dorothy Beckett
Title: Visualization of the Human Transcription Initiation Machinery
Time: 4:00 - 5:00PM
Date: Monday May 1, 2017
Location: 2212 Benjamin Banneker Room, Stamp Student Union

Abstract

Eukaryotic gene transcription requires the assembly at the promoter of a large pre-initiation complex (PIC) that includes RNA polymerase II (Pol II) and the general transcription factors TFIID, TFIIA, TFIIB, TFIIF, TFIIE, and TFIIH. The size and complexity of Pol II, TFIID, and TFIIH have precluded their reconstitution from heterologous systems, and purification relies on scarce endogenous sources. Together with their conformational flexibility and the transient nature of their interactions, these limitations had precluded structural characterization of the PIC. In the last few years, however,progress in cryo–electron microscopy (cryo-EM) has made possible the visualization, at increasingly better resolution, of large PIC assemblies in different functional states. These structures can now be interpreted in near-atomic detail and provide an exciting structural framework for past and future functional studies, giving us unique mechanistic insight into the complex process of transcription initiation.

Eva Nogales (Marker Lecturer), University of California Berkeley, HHMI
Hosted by Dorothy Beckett
Title: Structural Basis of Microtubule Dynamic Instability and its Regulation
Time: 11:00AM - 12:00PM
Date: Tuesday May 2, 2017
Location: 0112 Marker Seminar Room, Chemistry Building

Abstract

Microtubules (MTs) are crucial components of the cytoskeleton and play a central role in cell division. Essential to MT function is the property of dynamic instability, the stochastic switching between MT growing and shrinking linked to GTP binding and hydrolysis MT dynamics are tightly regulated in vivo by a number of MT-associated proteins (MAPs), while widely successful anti-mitotic chemotherapeutics, such as Taxol, bind to and stabilize MTs, inhibiting dynamic instability and preventing cells from dividing. Structural studies of microtubule in different nucleotide states, and bound to different MAPs or drugs are important to generate a mechanistic understanding of the regulated function of MTs. As non-crystallizable polymers, MT have been the target of cryo-electron microscopy (cryo-EM) studies since the technique was first established. Over the years, image processing strategies have been developed that take care of the unique, pseudo-helical symmetry of the microtubule. With recent progress in data quality and data processing, cryo-EM reconstructions are now reaching resolutions that allow the generation of atomic models of microtubules and the factors that bind them.