CORE 1: TECHNOLOGY Core 1 Project 1 – MEASURE a. Network and Pathway Data Integration. b. Virtual Experiment. c. Optical Probe Development. d. Fluorescence.

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CORE 1: TECHNOLOGY Core 1 Project 1 – MEASURE a. Network and Pathway Data Integration. b. Virtual Experiment. c. Optical Probe Development. d. Fluorescence Correlation Spectroscopy. Core 1 Project 2 - MODEL a. Modularity and Multistate Complexes. b. Molecular Flux in Crowded Spaces. c. Stochastic Modeling and Discrete Particles. d. Moving Boundaries. Core 1 Project 3 – MANIPULATE a. Signaling Platforms. b. In Vivo Nanofabrication. c. Holographic Optical Tweezers. CORE 2: DRIVING BIOLOGICAL PROJECTS DBP 1 “Study of Neutrophil Polarity” Dianqing Wu DBP 2 “Cell Polarity and Cell Fusion” William Mohler DBP 3 “Polarity in A2 RNA Trafficking Pathway in Neurons John H. Carson DBP 4 “Regulation of Cell Locomotion by Microtubules” Vladimir I. Rodionov DBP 5 “Spatial Asymmetry, Nuclear Transport, and Signaling” Ian G. Macara DBP 6 “Simulation of Cytoskeletal Structure and Mobility” Thomas D. Pollard CORE 3: FACILITIES CORE 4: TRAINING CORE 5: DISSEMINATION CORE 6: ADMINISTRATION Measure ModelManipulate

Quantitative Cell Biology Predictions Dynamics of Cellular Structures and Molecules Simulation Hypothesis (Model) What are the initial concentrations, diffusion coefficients and locations of all the implicated molecules? What are the rate laws and rate constants for all the biochemical transformations? What are the membrane fluxes and how are they regulated? How are the forces controlling cytoskeletal mechanics regulated? Experiment Trends in Cell Biology 13: (2003)

Today’s Talks Bill Mohler – Contrast Mechanisms in Non- linear Optics (SHG of muscle) Paul Campagnola - New Photoactivators for Nano/Microfabrication Les Loew – Novel Environmentally- Sensitive Chromophores for Fluorescence and SHG Imaging

Measure - Measure - Fluorescent Probes Les Loew Membrane Domains Membrane Association and Fusion Environmentally-Sensitive Labels TPEFSHGOverlay

Characterization and Application of a New Optical Probe for Membrane Lipid Domains Lei Jin et al., Biophys. J., 2006

mergeSHG TPF 540/50 675/50 SHG and 2PF images of GUVs stained by di-4-ANEPPDHQ. The dye shows stronger SHG in the liquid disordered phase, corresponding to the red-shifted 2PF.

Di-4-ANEPPDHQ Reveals Lipid Polarity in Polarized Migrating Neutrophils

Newly synthesized dyes and their emission maxima in liquid ordered and liquid disordered lipid membranes Ping Yan and Joe Wuskell

DPPC:Cholesterol DOPC

DPPC:Cholesterol DOPC

Fertilization of Di-8-ANEPPS Stained Sea Urchin Egg Andrew Millard and Mark Terasaki SHG TPEF

Unique Contrast Patterns from Resonance Enhanced Chiral SHG of Cell Membranes Ping Yan et al., J. Am. Chem. Soc., 2006 Monomer Racemic Chiral SHG2PF SHG

Functionalization of 10KD Dextran for Covalent Linkage to Rose Bengal (Ping Yan)

Short Term Goals Develop covalent labeling reagents from the most environmentally sensitive of our new chromophores (Ping Yan, Joe Wuskell) Support Nanofabrication with improved intracellular photosensitizers (Ping Yan) Determine the mechanism of 2PF increase and SHG decrease upon fusion of exocytic vesicles (Aifang Xie)