scholarly journals Autoinhibition of kinesin-1 is essential to the dendrite-specific localization of Golgi outposts

2018 ◽  
Author(s):  
Michael T. Kelliher ◽  
Yang Yue ◽  
Ashley Ng ◽  
Daichi Kamiyama ◽  
Bo Huang ◽  
...  
Keyword(s):  
Structure Function ◽  
Molecular Motors ◽  
Genetic Interaction ◽  
Function Analysis ◽  
Molecular Motor ◽  
Neuronal Polarity ◽  
Growth Defects ◽  
Specific Localization ◽  
Selective Localization

AbstractNeuronal polarity relies on the selective localization of cargo to axons or dendrites. The molecular motor kinesin-1 moves cargo into axons but is also active in dendrites. This raises the question of how kinesin-1 activity is regulated to maintain the compartment-specific localization of cargo. Our in vivo structure-function analysis of endogenous Drosophila kinesin-1 reveals a novel role for autoinhibition in enabling the dendrite-specific localization of Golgi outposts. Mutations that disrupt kinesin-1 autoinhibition result in the axonal mislocalization of Golgi outposts. Autoinhibition also regulates kinesin-1 localization. Uninhibited kinesin-1 accumulates in axons and is depleted from dendrites, correlating with the change in outpost distribution and dendrite growth defects. Genetic interaction tests show that a balance of kinesin-1 inhibition and dynein activity is necessary to localize Golgi outposts to dendrites and keep them from entering axons. Our data indicate that kinesin-1 activity is precisely regulated by autoinhibition to achieve the selective localization of dendritic cargo.SummaryNeuronal polarity relies on the axon-or dendrite-specific localization of cargo by molecular motors such as kinesin-1. These studies show autoinhibition regulates both kinesin-1 activity and localization to keep dendritic cargo from entering axons.

scholarly journals Autoinhibition of kinesin-1 is essential to the dendrite-specific localization of Golgi outposts

2018 ◽  
Vol 217 (7) ◽  
pp. 2531-2547 ◽  
Author(s):  
Michael T. Kelliher ◽  
Yang Yue ◽  
Ashley Ng ◽  
Daichi Kamiyama ◽  
Bo Huang ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Structure Function ◽  
Genetic Interaction ◽  
Function Analysis ◽  
Molecular Motor ◽  
Neuronal Polarity ◽  
Growth Defects ◽  
Specific Localization ◽  
Selective Localization

Neuronal polarity relies on the selective localization of cargo to axons or dendrites. The molecular motor kinesin-1 moves cargo into axons but is also active in dendrites. This raises the question of how kinesin-1 activity is regulated to maintain the compartment-specific localization of cargo. Our in vivo structure–function analysis of endogenous Drosophila melanogaster kinesin-1 reveals a novel role for autoinhibition in enabling the dendrite-specific localization of Golgi outposts. Mutations that disrupt kinesin-1 autoinhibition result in the axonal mislocalization of Golgi outposts. Autoinhibition also regulates kinesin-1 localization. Uninhibited kinesin-1 accumulates in axons and is depleted from dendrites, correlating with the change in outpost distribution and dendrite growth defects. Genetic interaction tests show that a balance of kinesin-1 inhibition and dynein activity is necessary to localize Golgi outposts to dendrites and keep them from entering axons. Our data indicate that kinesin-1 activity is precisely regulated by autoinhibition to achieve the selective localization of dendritic cargo.

scholarly journals Regulated nuclear import of the Drosophila rel protein dorsal: structure-function analysis.

1996 ◽  
Vol 16 (3) ◽  
pp. 1103-1114 ◽  
Author(s):  
S Govind ◽  
E Drier ◽  
L H Huang ◽  
R Steward
Keyword(s):  
Amino Acids ◽  
Structure Function ◽  
Nuclear Localization ◽  
Nuclear Import ◽  
Function Analysis ◽  
Drosophila Embryo ◽  
Nuclear Targeting ◽  
Homology Region ◽  
Early Drosophila Embryo

The formation of a gradient of nuclear Dorsal protein in the early Drosophila embryo is the last step in a maternally encoded dorsal-ventral signal transduction pathway. This gradient is formed in response to a ventral signal, which leads to the dissociation of cytoplasmic Dorsal from the I kappa B homolog Cactus. Free Dorsal is then targeted to the nucleus. Dorsal is a Rel-family transcription factor. Signal-dependent nuclear localization characterizes the regulation of Rel proteins. In order to identify regions of Dorsal that are essential for its homodimerization, nuclear targeting, and interaction with Cactus, we have performed an in vivo structure-function analysis. Our results show that all these functions are carried out by regions within the conserved Rel-homology region of Dorsal. The C-terminal divergent half of Dorsal is dispensable for its selective nuclear import. A basic stretch of 6 amino acids at the C terminus of the Rel-homology region is necessary for nuclear localization. This nuclear localization signal is not required for Cactus binding. Removal of the N-terminal 40 amino acids abolished the nuclear import of Dorsal, uncovering a potentially novel function for this highly conserved region.

scholarly journals Dense granule trafficking in Toxoplasma gondii requires a unique class 27 myosin and actin filaments

2016 ◽  
Vol 27 (13) ◽  
pp. 2080-2089 ◽  
Author(s):  
Aoife T. Heaslip ◽  
Shane R. Nelson ◽  
David M. Warshaw
Keyword(s):  
Toxoplasma Gondii ◽  
Molecular Motors ◽  
Fluorescent Protein ◽  
Molecular Motor ◽  
Structural Similarity ◽  
Dense Granule ◽  
Lytic Cycle ◽  
Secretory Vesicles ◽  
Filamentous Actin

The survival of Toxoplasma gondii within its host cell requires protein release from secretory vesicles, called dense granules, to maintain the parasite’s intracellular replicative niche. Despite the importance of DGs, nothing is known about the mechanisms underlying their transport. In higher eukaryotes, secretory vesicles are transported to the plasma membrane by molecular motors moving on their respective cytoskeletal tracks (i.e., microtubules and actin). Because the organization of these cytoskeletal structures differs substantially in T. gondii, the molecular motor dependence of DG trafficking is far from certain. By imaging the motions of green fluorescent protein–tagged DGs in intracellular parasites with high temporal and spatial resolution, we show through a combination of molecular genetics and chemical perturbations that directed DG transport is independent of microtubules and presumably their kinesin/dynein motors. However, directed DG transport is dependent on filamentous actin and a unique class 27 myosin, TgMyoF, which has structural similarity to myosin V, the prototypical cargo transporter. Actomyosin DG transport was unexpected, since filamentous parasite actin has yet to be visualized in vivo due in part to the prevailing model that parasite actin forms short, unstable filaments. Thus our data uncover new critical roles for these essential proteins in the lytic cycle of this devastating pathogen.

scholarly journals Structure/Function Analysis of the Vaccinia Virus F18 Phosphoprotein, an Abundant Core Component Required for Virion Maturation and Infectivity

2010 ◽  
Vol 84 (13) ◽  
pp. 6846-6860 ◽  
Author(s):  
Nadi T. Wickramasekera ◽  
Paula Traktman
Keyword(s):  
Amino Acids ◽  
Structure Function ◽  
Protein Interactions ◽  
Function Analysis ◽  
Aromatic Amino Acids ◽  
Protein Protein Interactions ◽  
Core Component ◽  
Virion Morphogenesis

ABSTRACT Poxvirus virions, whose outer membrane surrounds two lateral bodies and a core, contain at least 70 different proteins. The F18 phosphoprotein is one of the most abundant core components and is essential for the assembly of mature virions. We report here the results of a structure/function analysis in which the role of conserved cysteine residues, clusters of charged amino acids and clusters of hydrophobic/aromatic amino acids have been assessed. Taking advantage of a recombinant virus in which F18 expression is IPTG (isopropyl-β-d-thiogalactopyranoside) dependent, we developed a transient complementation assay to evaluate the ability of mutant alleles of F18 to support virion morphogenesis and/or to restore the production of infectious virus. We have also examined protein-protein interactions, comparing the ability of mutant and WT F18 proteins to interact with WT F18 and to interact with the viral A30 protein, another essential core component. We show that F18 associates with an A30-containing multiprotein complex in vivo in a manner that depends upon clusters of hydrophobic/aromatic residues in the N′ terminus of the F18 protein but that it is not required for the assembly of this complex. Finally, we confirmed that two PSSP motifs within F18 are the sites of phosphorylation by cellular proline-directed kinases in vitro and in vivo. Mutation of both of these phosphorylation sites has no apparent impact on virion morphogenesis but leads to the assembly of virions with significantly reduced infectivity.

scholarly journals In vivo structure-function analysis of Drosophila HAIRLESS

1997 ◽  
Vol 67 (1) ◽  
pp. 97-106 ◽  
Author(s):  
Dieter Maier ◽  
Jörg Marquart ◽  
Annick Thompson-Fontaine ◽  
Irmtraud Beck ◽  
Elisa Wurmbach ◽  
...  
Keyword(s):  
Structure Function ◽  
Function Analysis

scholarly journals In vivo structure-function analysis of human Dicer reveals directional processing of precursor miRNAs

RNA ◽  
2012 ◽  
Vol 18 (6) ◽  
pp. 1116-1122 ◽  
Author(s):  
A. M. Gurtan ◽  
V. Lu ◽  
A. Bhutkar ◽  
P. A. Sharp
Keyword(s):  
Structure Function ◽  
Function Analysis ◽  
Precursor Mirnas

scholarly journals Structure-function analysis of β-arrestin Kurtz reveals a critical role of receptor interactions in downregulation of GPCR signaling in vivo

2019 ◽  
Vol 455 (2) ◽  
pp. 409-419 ◽  
Author(s):  
Fei Chai ◽  
Wenjian Xu ◽  
Timothy Musoke ◽  
George Tarabelsi ◽  
Steven Assaad ◽  
...  
Keyword(s):  
Structure Function ◽  
Function Analysis ◽  
Critical Role ◽  
Gpcr Signaling ◽  
Receptor Interactions

scholarly journals Non-Canonical G-quadruplexes cause the hCEB1 minisatellite instability in Saccharomyces cerevisiae

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Aurèle Piazza ◽  
Xiaojie Cui ◽  
Michael Adrian ◽  
Frédéric Samazan ◽  
Brahim Heddi ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Structure Function ◽  
Genomic Instability ◽  
Function Analysis ◽  
Structural Features ◽  
Detectable Effect ◽  
Definition Of ◽  
And Function

G-quadruplexes (G4) are polymorphic four-stranded structures formed by certain G-rich nucleic acids in vitro, but the sequence and structural features dictating their formation and function in vivo remains uncertain. Here we report a structure-function analysis of the complex hCEB1 G4-forming sequence. We isolated four G4 conformations in vitro, all of which bear unusual structural features: Form 1 bears a V-shaped loop and a snapback guanine; Form 2 contains a terminal G-triad; Form 3 bears a zero-nucleotide loop; and Form 4 is a zero-nucleotide loop monomer or an interlocked dimer. In vivo, Form 1 and Form 2 differently account for 2/3rd of the genomic instability of hCEB1 in two G4-stabilizing conditions. Form 3 and an unidentified form contribute to the remaining instability, while Form 4 has no detectable effect. This work underscores the structural polymorphisms originated from a single highly G-rich sequence and demonstrates the existence of non-canonical G4s in cells, thus broadening the definition of G4-forming sequences.

Impact for molecular biology in cardiology

1991 ◽  
Vol 261 (4) ◽  
pp. L8-L14
Author(s):  
Robert Roberts
Keyword(s):  
Molecular Biology ◽  
Structure Function ◽  
Recombinant Dna ◽  
Function Analysis ◽  
Coronary Thrombosis ◽  
Genetic Regulation ◽  
Cardiac Growth ◽  
Genetically Engineer ◽  
Hereditary Disorders

The recent development and application of the techniques of recombinant DNA and molecular biology ignited an explosion in biomedical research, which has been embraced by medicine. However, cardiology as a subspecialty has been slower in adopting these techniques, in part because the heart is a nonproliferating organ and in part because it was not easily accessible until recently. The techniques of recombinant DNA were not possible until the 1970s. In that decade four major discoveries occurred that launched molecular biology into the 21st century. These seminal contributions were 1) the discovery and application of specific restriction endonucleases, 2) the discovery of reverse transcriptase, 3) the development of the cloning technique, and 4) the ability to rapidly sequence nucleic acids. The techniques of recombinant DNA offer several unique advantages over existing scientific disciplines, such as the abilities: 1) to perform in vivo structure-function analysis, 2) to genetically engineer drugs, 3) to perform diagnostic in situ hybridization, 4) to isolate genes responsible for hereditary disorders, and 5) to understand the genetic regulation of cardiac growth. These techniques are discussed in their application to cardiac disorders, including the development of new recombinant molecules for the treatment of coronary thrombosis and the potential to modulate the cardiac growth response to various forms of injury such as myocardial infarction and hypertension. cardiac growth; genetic engineering; molecular genetics; structure function analysis

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