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94 Janelia Publications

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    12/22/11 | Observations on clustered synaptic plasticity and highly structured input patterns.
    Magee JC
    Neuron. 2011 Dec 22;72(6):887-8. doi: 10.1016/j.neuron.2011.12.009

    In this issue of Neuron, Makino and Malinow and Kleindienst et al. present evidence of a behaviorally induced form of synaptic plasticity that would encourage the development of fine-scale structured input patterns and the binding of features within single neurons.

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    12/16/11 | Synthesis of rhodamines from fluoresceins using Pd-catalyzed C-N cross-coupling.
    Grimm JB, Lavis LD
    Organic Letters. 2011 Dec 16;13(24):6354-7. doi: 10.1021/ol202618t

    A unified, convenient, and efficient strategy for the preparation of rhodamines and N,N’-diacylated rhodamines has been developed. Fluorescein ditriflates were found to undergo palladium-catalyzed C-N cross-coupling with amines, amides, carbamates, and other nitrogen nucleophiles to provide direct access to known and novel rhodamine derivatives, including fluorescent dyes, quenchers, and latent fluorophores.

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    12/12/11 | The BTB/POZ zinc finger protein Broad-Z3 promotes dendritic outgrowth during metamorphic remodeling of the peripheral stretch receptor dbd.
    Scott JA, Williams DW, Truman JW
    Neural Development. 2011 Dec 12;6:39. doi: 10.1186/1749-8104-6-39

    Various members of the family of BTB/POZ zinc-finger transcription factors influence patterns of dendritic branching. One such member, Broad, is notable because its BrZ3 isoform is widely expressed in Drosophila in immature neurons around the time of arbor outgrowth. We used the metamorphic remodeling of an identified sensory neuron, the dorsal bipolar dendrite sensory neuron (dbd), to examine the effects of BrZ3 expression on the extent and pattern of dendrite growth during metamorphosis.

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    Eddy/Rivas Lab
    12/07/11 | Phosphorylation at the interface.
    Davis FP
    Structure . 2011 Dec 7;19:1726-7. doi: 10.1016/j.str.2011.11.006

    Proteomic studies have identified thousands of eukaryotic phosphorylation sites (phosphosites), but few are functionally characterized. Nishi et al., in this issue of Structure, characterize phosphosites at protein-protein interfaces and estimate the effect of their phosphorylation on interaction affinity, by combining proteomics data with protein structures.

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    12/02/11 | Structure of the escherichia coli phosphonate binding protein PhnD and rationally optimized phosphonate biosensors.
    Alicea I, Marvin JS, Miklos AE, Ellington AD, Looger LL, Schreiter ER
    Journal of Molecular Biology. 2011 Dec 2;414(3):356-69. doi: 10.1016/j.jmb.2011.09.047

    The phnD gene of Escherichia coli encodes the periplasmic binding protein of the phosphonate (Pn) uptake and utilization pathway. We have crystallized and determined structures of E. coli PhnD (EcPhnD) in the absence of ligand and in complex with the environmentally abundant 2-aminoethylphosphonate (2AEP). Similar to other bacterial periplasmic binding proteins, 2AEP binds near the center of mass of EcPhnD in a cleft formed between two lobes. Comparison of the open, unliganded structure with the closed 2AEP-bound structure shows that the two lobes pivot around a hinge by \~{}70° between the two states. Extensive hydrogen bonding and electrostatic interactions stabilize 2AEP, which binds to EcPhnD with low nanomolar affinity. These structures provide insight into Pn uptake by bacteria and facilitated the rational design of high signal-to-noise Pn biosensors based on both coupled small-molecule dyes and autocatalytic fluorescent proteins.

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    12/01/11 | A critical role of mitochondrial phosphatase Ptpmt1 in embryogenesis reveals a mitochondrial metabolic stress-induced differentiation checkpoint in embryonic stem cells.
    Shen J, Liu X, Yu W, Liu J, Nibbelink MG, Guo C, Finkel T, Qu C
    Molecular and Cellular Biology. 2011 Dec;31:4902-16. doi: 10.1128/MCB.05629-11

    Mitochondria are highly dynamic organelles that play multiple roles in cells. How mitochondria cooperatively modulate embryonic stem (ES) cell function during development is not fully understood. Global disruption of Ptpmt1, a mitochondrial Pten-like phosphatidylinositol phosphate (PIP) phosphatase, resulted in developmental arrest and postimplantation lethality. Ptpmt1(-/-) blastocysts failed to outgrow, and inner-cell-mass cells failed to thrive. Depletion of Ptpmt1 in conditional knockout ES cells decreased proliferation without affecting energy homeostasis or cell survival. Differentiation of Ptpmt1-depleted ES cells was essentially blocked. This was accompanied by upregulation of cyclin-dependent kinase inhibitors and a significant cell cycle delay. Reintroduction of wild-type but not of catalytically deficient Ptpmt1 C132S or truncated Ptpmt1 lacking the mitochondrial localization signal restored the differentiation capabilities of Ptpmt1 knockout ES cells. Intriguingly, Ptpmt1 is specifically important for stem cells, as ablation of Ptpmt1 in differentiated embryonic fibroblasts did not disturb cellular function. Further analyses demonstrated that oxygen consumption of Ptpmt1-depleted cells was decreased, while glycolysis was concomitantly enhanced. In addition, mitochondrial fusion/dynamics were compromised in Ptpmt1 knockout cells due to accumulation of PIPs. These studies, while establishing a crucial role for Ptpmt1 phosphatase in embryogenesis, reveal a mitochondrial metabolic stress-activated checkpoint in the control of ES cell differentiation.

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    12/01/11 | Advances in the chemistry of small molecule fluorescent probes.
    Wysocki LM, Lavis LD
    Current Opinion in Chemical Biology. 2011 Dec;15(6):752-9. doi: 10.1016/j.cbpa.2011.10.013

    Small molecule fluorophores are essential tools for chemical biology. A benefit of synthetic dyes is the ability to employ chemical approaches to control the properties and direct the position of the fluorophore. Applying modern synthetic organic chemistry strategies enables efficient tailoring of the chemical structure to obtain probes for specific biological experiments. Chemistry can also be used to activate fluorophores; new fluorogenic enzyme substrates and photoactivatable compounds with improved properties have been prepared that facilitate advanced imaging experiments with low background fluorescence. Finally, chemical reactions in live cells can be used to direct the spatial distribution of the fluorophore, allowing labeling of defined cellular regions with synthetic dyes.

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    12/01/11 | Super-resolution 3D microscopy of live whole cells using structured illumination.
    Shao L, Kner P, Rego EH, Gustafsson MG
    Nature Methods. 2011 Dec;8:1044-6. doi: 10.1038/nmeth.1734

    Three-dimensional (3D) structured-illumination microscopy (SIM) can double the lateral and axial resolution of a wide-field fluorescence microscope but has been too slow for live imaging. Here we apply 3D SIM to living samples and record whole cells at up to 5 s per volume for >50 time points with 120-nm lateral and 360-nm axial resolution. We demonstrate the technique by imaging microtubules in S2 cells and mitochondria in HeLa cells.

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    12/01/11 | Superresolution fluorescence imaging of mitochondrial nucleoids reveals their spatial range, limits, and membrane interaction.
    Brown TA, Tkachuk AN, Shtengel G, Kopek BG, Bogenhagen DF, Hess HF, Clayton DA
    Molecular and Cellular Biology. 2011 Dec;31:4994-5010. doi: 10.1128/MCB.05694-11

    A fundamental objective in molecular biology is to understand how DNA is organized in concert with various proteins, RNA, and biological membranes. Mitochondria maintain and express their own DNA (mtDNA), which is arranged within structures called nucleoids. Their functions, dimensions, composition, and precise locations relative to other mitochondrial structures are poorly defined. Superresolution fluorescence microscopy techniques that exceed the previous limits of imaging within the small and highly compartmentalized mitochondria have been recently developed. We have improved and employed both two- and three-dimensional applications of photoactivated localization microscopy (PALM and iPALM, respectively) to visualize the core dimensions and relative locations of mitochondrial nucleoids at an unprecedented resolution. PALM reveals that nucleoids differ greatly in size and shape. Three-dimensional volumetric analysis indicates that, on average, the mtDNA within ellipsoidal nucleoids is extraordinarily condensed. Two-color PALM shows that the freely diffusible mitochondrial matrix protein is largely excluded from the nucleoid. In contrast, nucleoids are closely associated with the inner membrane and often appear to be wrapped around cristae or crista-like inner membrane invaginations. Determinations revealing high packing density, separation from the matrix, and tight association with the inner membrane underscore the role of mechanisms that regulate access to mtDNA and that remain largely unknown.

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    12/01/11 | Wiring economy and volume exclusion determine neuronal placement in the Drosophila brain.
    Rivera-Alba M, Vitaladevuni SN, Mischenko Y, Lu Z, Takemura S, Scheffer L, Meinertzhagen I, Chklovskii D, Polavieja G
    Current Biology. 2011 Dec;21(23):2000-5. doi: 10.1016/j.cub.2011.10.022

    Wiring economy has successfully explained the individual placement of neurons in simple nervous systems like that of Caenorhabditis elegans [1-3] and the locations of coarser structures like cortical areas in complex vertebrate brains [4]. However, it remains unclear whether wiring economy can explain the placement of individual neurons in brains larger than that of C. elegans. Indeed, given the greater number of neuronal interconnections in larger brains, simply minimizing the length of connections results in unrealistic configurations, with multiple neurons occupying the same position in space. Avoiding such configurations, or volume exclusion, repels neurons from each other, thus counteracting wiring economy. Here we test whether wiring economy together with volume exclusion can explain the placement of neurons in a module of the Drosophila melanogaster brain known as lamina cartridge [5-13]. We used newly developed techniques for semiautomated reconstruction from serial electron microscopy (EM) [14] to obtain the shapes of neurons, the location of synapses, and the resultant synaptic connectivity. We show that wiring length minimization and volume exclusion together can explain the structure of the lamina microcircuit. Therefore, even in brains larger than that of C. elegans, at least for some circuits, optimization can play an important role in individual neuron placement.

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