@article {47648, title = {Multiscale diffusion in the mitotic Drosophila melanogaster syncytial blastoderm.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {109}, year = {2012}, month = {2012 May 29}, pages = {8588-93}, abstract = {

Despite the fundamental importance of diffusion for embryonic morphogen gradient formation in the early Drosophila melanogaster embryo, there remains controversy regarding both the extent and the rate of diffusion of well-characterized morphogens. Furthermore, the recent observation of diffusional "compartmentalization" has suggested that diffusion may in fact be nonideal and mediated by an as-yet-unidentified mechanism. Here, we characterize the effects of the geometry of the early syncytial Drosophila embryo on the effective diffusivity of cytoplasmic proteins. Our results demonstrate that the presence of transient mitotic membrane furrows results in a multiscale diffusion effect that has a significant impact on effective diffusion rates across the embryo. Using a combination of live-cell experiments and computational modeling, we characterize these effects and relate effective bulk diffusion rates to instantaneous diffusion coefficients throughout the syncytial blastoderm nuclear cycle phase of the early embryo. This multiscale effect may be related to the effect of interphase nuclei on effective diffusion, and thus we propose that an as-yet-unidentified role of syncytial membrane furrows is to temporally regulate bulk embryonic diffusion rates to balance the multiscale effect of interphase nuclei, which ultimately stabilizes the shapes of various morphogen gradients.

}, keywords = {Algorithms, Animals, Animals, Genetically Modified, Blastoderm, Cell Compartmentation, Cell Membrane, Computer Simulation, Cytoplasm, Diffusion, Drosophila melanogaster, Embryo, Nonmammalian, Fluorescence Recovery After Photobleaching, Giant Cells, Green Fluorescent Proteins, Kinetics, Microscopy, Fluorescence, Mitosis, Models, Biological}, issn = {1091-6490}, doi = {10.1073/pnas.1204270109}, author = {Daniels, Brian R and Rikhy, Richa and Renz, Malte and Dobrowsky, Terrence M and Lippincott-Schwartz, Jennifer} } @article {47646, title = {Plasticity of the asialoglycoprotein receptor deciphered by ensemble FRET imaging and single-molecule counting PALM imaging.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {109}, year = {2012}, month = {2012 Oct 30}, pages = {E2989-97}, abstract = {

The stoichiometry and composition of membrane protein receptors are critical to their function. However, the inability to assess receptor subunit stoichiometry in situ has hampered efforts to relate receptor structures to functional states. Here, we address this problem for the asialoglycoprotein receptor using ensemble FRET imaging, analytical modeling, and single-molecule counting with photoactivated localization microscopy (PALM). We show that the two subunits of asialoglycoprotein receptor [rat hepatic lectin 1 (RHL1) and RHL2] can assemble into both homo- and hetero-oligomeric complexes, displaying three forms with distinct ligand specificities that coexist on the plasma membrane: higher-order homo-oligomers of RHL1, higher-order hetero-oligomers of RHL1 and RHL2 with two-to-one stoichiometry, and the homo-dimer RHL2 with little tendency to further homo-oligomerize. Levels of these complexes can be modulated in the plasma membrane by exogenous ligands. Thus, even a simple two-subunit receptor can exhibit remarkable plasticity in structure, and consequently function, underscoring the importance of deciphering oligomerization in single cells at the single-molecule level.

}, keywords = {Asialoglycoprotein Receptor, Fluorescence Resonance Energy Transfer, Ligands, Microscopy, Confocal}, issn = {1091-6490}, doi = {10.1073/pnas.1211753109}, author = {Renz, Malte and Daniels, Brian R and V{\'a}mosi, Gy{\"o}rgy and Arias, Irwin M and Lippincott-Schwartz, Jennifer} } @article {47656, title = {Plasticity of the asialoglycoprotein receptor deciphered by ensemble FRET imaging and single-molecule counting PALM imaging.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {109}, year = {2012}, month = {2012 Oct 30}, pages = {E2989-97}, abstract = {

The stoichiometry and composition of membrane protein receptors are critical to their function. However, the inability to assess receptor subunit stoichiometry in situ has hampered efforts to relate receptor structures to functional states. Here, we address this problem for the asialoglycoprotein receptor using ensemble FRET imaging, analytical modeling, and single-molecule counting with photoactivated localization microscopy (PALM). We show that the two subunits of asialoglycoprotein receptor [rat hepatic lectin 1 (RHL1) and RHL2] can assemble into both homo- and hetero-oligomeric complexes, displaying three forms with distinct ligand specificities that coexist on the plasma membrane: higher-order homo-oligomers of RHL1, higher-order hetero-oligomers of RHL1 and RHL2 with two-to-one stoichiometry, and the homo-dimer RHL2 with little tendency to further homo-oligomerize. Levels of these complexes can be modulated in the plasma membrane by exogenous ligands. Thus, even a simple two-subunit receptor can exhibit remarkable plasticity in structure, and consequently function, underscoring the importance of deciphering oligomerization in single cells at the single-molecule level.

}, keywords = {Asialoglycoprotein Receptor, Fluorescence Resonance Energy Transfer, Ligands, Microscopy, Confocal}, issn = {1091-6490}, doi = {10.1073/pnas.1211753109}, author = {Renz, Malte and Daniels, Brian R and V{\'a}mosi, Gy{\"o}rgy and Arias, Irwin M and Lippincott-Schwartz, Jennifer} }