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192 Publications

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    05/13/22 | Recovery mechanisms in the dragonfly righting reflex.
    Wang ZJ, Melfi J, Leonardo A
    Science. 2022 May 13;376(6594):754-758. doi: 10.1126/science.abg0946

    Insects have evolved sophisticated reflexes to right themselves in mid-air. Their recovery mechanisms involve complex interactions among the physical senses, muscles, body, and wings, and they must obey the laws of flight. We sought to understand the key mechanisms involved in dragonfly righting reflexes and to develop physics-based models for understanding the control strategies of flight maneuvers. Using kinematic analyses, physical modeling, and three-dimensional flight simulations, we found that a dragonfly uses left-right wing pitch asymmetry to roll its body 180 degrees to recover from falling upside down in ~200 milliseconds. Experiments of dragonflies with blocked vision further revealed that this rolling maneuver is initiated by their ocelli and compound eyes. These results suggest a pathway from the dragonfly's visual system to the muscles regulating wing pitch that underly the recovery. The methods developed here offer quantitative tools for inferring insects' internal actions from their acrobatics, and are applicable to a broad class of natural and robotic flying systems.

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    05/11/22 | Single molecule microscopy to profile the effect of zinc status of transcription factor dynamics
    Leah J. Damon , Jesse Aaron , Amy E. Palmer
    bioRxiv. 2022 May 11:. doi: 10.1101/2022.05.10.491421

    Transcription factors (TFs) are DNA binding proteins that control the expression of genes. The regulation of transcription is a complex process that involves binding of TFs to specific sequences, recruitment of cofactors and chromatin remodelers, assembly of the pre-initiation complex and ultimately the recruitment of RNA polymerase II. Increasing evidence suggests that TFs are highly dynamic and interact only transiently with DNA. Single molecule microscopy techniques are powerful approaches for visualizing and tracking individual TF molecules as they diffuse in the nucleus and interact with DNA. In this work, we employ multifocus microscopy and highly inclined and laminated optical sheet microscopy to track TF dynamics in response to perturbations in labile zinc inside cells. We sought to define whether zinc-dependent TFs sense changes in the labile zinc pool by determining whether their dynamics and DNA binding can be modulated by zinc. While it is widely appreciated that TFs need zinc to bind DNA, whether zinc occupancy and hence TF function are sensitive to changes in cellular zinc remain open questions. We utilized fluorescently tagged versions of the glucocorticoid receptor (GR), with two C4 zinc finger domains, and CCCTC-binding factor (CTCF), with eleven C2H2 zinc finger domains. We found that the biophysical dynamics of both TFs are susceptible to changes in zinc, but in subtly different ways. These results indicate that at least some transcription factors are sensitive to zinc dynamics, revealing a potential new layer of transcriptional regulation.

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    05/11/22 | Super-resolution microscopy reveals actomyosin dynamics in medioapical arrays.
    Moore RP, Fogerson SM, Tulu US, Yu JW, Cox AH, Sican MA, Li D, Legant WR, Weigel AV, Crawford JM, Betzig E, Kiehart DP
    Molecular Biology of the Cell. 2022 May 11:mbcE21110537. doi: 10.1091/mbc.E21-11-0537

    Arrays of actin filaments (F-actin) near the apical surface of epithelial cells (medioapical arrays) contribute to apical constriction and morphogenesis throughout phylogeny. Here, super-resolution approaches (grazing incidence structured illumination, GI-SIM and lattice light sheet, LLSM) microscopy resolve individual, fluorescently labeled F-actin and bipolar myosin filaments that drive amnioserosa cell shape changes during dorsal closure in . In expanded cells, F-actin and myosin form loose, apically domed meshworks at the plasma membrane. The arrays condense as cells contract, drawing the domes into the plane of the junctional belts. As condensation continues, individual filaments are no longer uniformly apparent. As cells expand, arrays of actomyosin are again resolved - some F-actin turnover likely occurs, but a large fraction of existing filaments rearrange. In morphologically isotropic cells, actin filaments are randomly oriented and during contraction, are drawn together but remain essentially randomly oriented. In anisotropic cells, largely parallel actin filaments are drawn closer to one another. Our images offer unparalleled resolution of F-actin in embryonic tissue show that medioapical arrays are tightly apposed to the plasma membrane, are continuous with meshworks of lamellar F-actin and thereby constitute modified cell cortex. In concert with other tagged array components, super-resolution imaging of live specimens will offer new understanding of cortical architecture and function. [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text].

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    05/10/22 | Rescue of behavioral and electrophysiological phenotypes in a Pitt-Hopkins syndrome mouse model by genetic restoration of expression.
    Kim H, Gao EB, Draper A, Berens NC, Vihma H, Zhang X, Higashi-Howard A, Ritola KD, Simon JM, Kennedy AJ, Philpot BD
    eLife. 2022 May 10;11:. doi: 10.7554/eLife.72290

    Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder caused by monoallelic mutation or deletion in the () gene. Individuals with PTHS typically present in the first year of life with developmental delay and exhibit intellectual disability, lack of speech, and motor incoordination. There are no effective treatments available for PTHS, but the root cause of the disorder, haploinsufficiency, suggests that it could be treated by normalizing gene expression. Here, we performed proof-of-concept viral gene therapy experiments using a conditional mouse model of PTHS and found that postnatally reinstating expression in neurons improved anxiety-like behavior, activity levels, innate behaviors, and memory. Postnatal reinstatement also partially corrected EEG abnormalities, which we characterized here for the first time, and the expression of key TCF4-regulated genes. Our results support a genetic normalization approach as a treatment strategy for PTHS, and possibly other TCF4-linked disorders.

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    05/09/22 | Gene structure-based homology search identifies highly divergent putative effector gene family.
    Stern DL, Han C
    Genome Biology and Evolution. 2022 May 09:. doi: 10.1093/gbe/evac069

    Homology of highly divergent genes often cannot be determined from sequence similarity alone. For example, we recently identified in the aphid Hormaphis cornu a family of rapidly evolving bicycle genes, which encode novel proteins implicated as plant gall effectors, and sequence similarity search methods yielded few putative bicycle homologs in other species. Coding sequence-independent features of genes, such as intron-exon boundaries, often evolve more slowly than coding sequences, however, and can provide complementary evidence for homology. We found that a linear logistic regression classifier using only structural features of bicycle genes identified many putative bicycle homologs in other species. Independent evidence from sequence features and intron locations supported homology assignments. To test the potential roles of bicycle genes in other aphids, we sequenced the genome of a second gall-forming aphid, Tetraneura nigriabdominalis, and found that many bicycle genes are strongly expressed in the salivary glands of the gall forming foundress. In addition, bicycle genes are strongly overexpressed in the salivary glands of a non-gall forming aphid, Acyrthosiphon pisum, and in the non-gall forming generations of Hormaphis cornu. These observations suggest that Bicycle proteins may be used by multiple aphid species to manipulate plants in diverse ways. Incorporation of gene structural features into sequence search algorithms may aid identification of deeply divergent homologs, especially of rapidly evolving genes involved in host-parasite interactions.

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    05/07/22 | Microbial models of development: Inspiration for engineering self-assembled synthetic multicellularity.
    Ricci-Tam C, Kuipa S, Kostman MP, Aronson MS, Sgro AE
    Semin Cell Dev Biol. 05/2022:. doi: 10.1016/j.semcdb.2022.04.014

    While the field of synthetic developmental biology has traditionally focused on the study of the rich developmental processes seen in metazoan systems, an attractive alternate source of inspiration comes from microbial developmental models. Microbes face unique lifestyle challenges when forming emergent multicellular collectives. As a result, the solutions they employ can inspire the design of novel multicellular systems. In this review, we dissect the strategies employed in multicellular development by two model microbial systems: the cellular slime mold Dictyostelium discoideum and the biofilm-forming bacterium Bacillus subtilis. Both microbes face similar challenges but often have different solutions, both from metazoan systems and from each other, to create emergent multicellularity. These challenges include assembling and sustaining a critical mass of participating individuals to support development, regulating entry into development, and assigning cell fates. The mechanisms these microbial systems exploit to robustly coordinate development under a wide range of conditions offer inspiration for a new toolbox of solutions to the synthetic development community. Additionally, recreating these phenomena synthetically offers a pathway to understanding the key principles underlying how these behaviors are be coordinated naturally.

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    05/03/22 | Bromodomains regulate dynamic targeting of the PBAF chromatin remodeling complex to chromatin hubs.
    Kenworthy CA, Haque N, Liou S, Chandris P, Wong V, Dziuba P, Lavis LD, Liu W, Singer RH, Coleman RA
    Biophysical Journal. 2022 May 3;121(9):1738-1752. doi: 10.1016/j.bpj.2022.03.027

    Chromatin remodelers actively target arrays of acetylated nucleosomes at select enhancers and promoters to facilitate or shut down the repeated recruitment of RNA Pol II during transcriptional bursting. It is poorly understood how chromatin remodelers such as PBAF dynamically target different chromatin states inside a live cell. Our live-cell single molecule fluorescence microscopy study reveals chromatin hubs throughout the nucleus where PBAF rapidly cycles on and off the genome. Deletion of PBAF's bromodomains impairs targeting and stable engagement of chromatin in hubs. Dual color imaging reveals that PBAF targets both euchromatic and heterochromatic hubs with distinct genome binding kinetic profiles that mimic chromatin stability. Removal of PBAF's bromodomains stabilizes H3.3 binding within chromatin indicating that bromodomains may play a direct role in remodeling of the nucleosome. Our data suggests that PBAF's dynamic bromodomain mediated engagement of a nucleosome may reflect the chromatin remodeling potential of differentially bound chromatin states.

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    05/01/22 | Cryo-EM structures of the human GATOR1-Rag-Ragulator complex reveal a spatial-constraint regulated GAP mechanism.
    Egri SB, Ouch C, Chou H, Yu Z, Song K, Xu C, Shen K
    Molecular Cell. 2022 May 01;36(1):. doi: 10.1016/j.molcel.2022.03.002

    mTORC1 controls cellular metabolic processes in response to nutrient availability. Amino acid signals are transmitted to mTORC1 through the Rag GTPases, which are localized on the lysosomal surface by the Ragulator complex. The Rag GTPases receive amino acid signals from multiple upstream regulators. One negative regulator, GATOR1, is a GTPase activating protein (GAP) for RagA. GATOR1 binds to the Rag GTPases via two modes: an inhibitory mode and a GAP mode. How these two binding interactions coordinate to process amino acid signals is unknown. Here, we resolved three cryo-EM structural models of the GATOR1-Rag-Ragulator complex, with the Rag-Ragulator subcomplex occupying the inhibitory site, the GAP site, and both binding sites simultaneously. When the Rag GTPases bind to GATOR1 at the GAP site, both Rag subunits contact GATOR1 to coordinate their nucleotide loading states. These results reveal a potential GAP mechanism of GATOR1 during the mTORC1 inactivation process.

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    04/29/22 | Cryo-EM structure of the EBV ribonucleotide reductase BORF2 and mechanism of APOBEC3B inhibition.
    Shaban NM, Yan R, Shi K, Moraes SN, Cheng AZ, Carpenter MA, McLellan JS, Yu Z, Harris RS
    Science Advances. 2022 Apr 29;8(17):eabm2827. doi: 10.1126/sciadv.abm2827

    Viruses use a plethora of mechanisms to evade immune responses. A recent example is neutralization of the nuclear DNA cytosine deaminase APOBEC3B by the Epstein-Barr virus (EBV) ribonucleotide reductase subunit BORF2. Cryo-EM studies of APOBEC3B-BORF2 complexes reveal a large >1000-Å binding surface composed of multiple structural elements from each protein, which effectively blocks the APOBEC3B active site from accessing single-stranded DNA substrates. Evolutionary optimization is suggested by unique insertions in BORF2 absent from other ribonucleotide reductases and preferential binding to APOBEC3B relative to the highly related APOBEC3A and APOBEC3G enzymes. A molecular understanding of this pathogen-host interaction has potential to inform the development of drugs that block the interaction and liberate the natural antiviral activity of APOBEC3B. In addition, given a role for APOBEC3B in cancer mutagenesis, it may also be possible for information from the interaction to be used to develop DNA deaminase inhibitors.

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    04/25/22 | Regulation of Drosophila courtship behavior by the Tlx/tailless-like nuclear receptor, dissatisfaction.
    Duckhorn JC, Cande J, Metkus MC, Song H, Altamirano S, Stern DL, Shirangi TR
    Current Biology. 2022 Apr 25;32(8):1703-1714. doi: 10.1016/j.cub.2022.02.031

    Sexually dimorphic courtship behaviors in Drosophila melanogaster develop from the activity of the sexual differentiation genes, doublesex (dsx) and fruitless (fru), functioning with other regulatory factors that have received little attention. The dissatisfaction (dsf) gene encodes an orphan nuclear receptor homologous to vertebrate Tlx and Drosophila tailless that is critical for the development of several aspects of female- and male-specific sexual behaviors. Here, we report the pattern of dsf expression in the central nervous system and show that the activity of sexually dimorphic abdominal interneurons that co-express dsf and dsx is necessary and sufficient for vaginal plate opening in virgin females, ovipositor extrusion in mated females, and abdominal curling in males during courtship. We find that dsf activity results in different neuroanatomical outcomes in females and males, promoting and suppressing, respectively, female development and function of these neurons depending upon the sexual state of dsx expression. We posit that dsf and dsx interact to specify sex differences in the neural circuitry for dimorphic abdominal behaviors.

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