scholarly journals LC3B phosphorylation regulates FYCO1 binding and directional transport of autophagosomes

2020 ◽  
Author(s):  
Jose L. Nieto-Torres ◽  
Sean-Luc Shanahan ◽  
Romain Chassefeyre ◽  
Sara Landeras-Bueno ◽  
Sandra E. Encalada ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Translational Regulation ◽  
Membrane Vesicles ◽  
Adaptor Proteins ◽  
Cell Types ◽  
Cell Periphery ◽  
Anterograde Transport ◽  
Disease Etiology ◽  
Multiple Cell ◽  
Directional Transport

SUMMARYMacroautophagy (hereafter referred to as autophagy) is a conserved process that promotes cellular homeostasis through the degradation of cytosolic components, also known as cargo. During autophagy, cargo is sequestered into double-membrane vesicles called autophagosomes, which are predominantly transported in the retrograde direction to the perinuclear region to fuse with lysosomes, thus ensuring cargo degradation [1]. The mechanisms regulating directional autophagosomal transport remain unclear. The ATG8 family of proteins associate with autophagosome membranes [2] and play key roles in autophagy, such as the movement of autophagosomes. This is achieved via the interaction of ATG8 with adaptor proteins, including FYCO1, a protein involved in the anterograde transport of autophagosomes toward the cell periphery [1,3–5]. We previously reported that phosphorylation of LC3B/ATG8 on threonine 50 (LC3B-T50) by the Hippo kinase STK4 is required for autophagy through unknown mechanisms [6]. Here, we show that LC3B-T50 phosphorylation decreases the interaction between LC3B and FYCO1, which in turn regulates the starvation-induced perinuclear positioning of autophagosomes. Moreover, non-phosphorylatable LC3B-T50A aberrantly switches the predominant retrograde movement of autophagosomes to anterograde movement towards the cell periphery in multiple cell types, including in mouse primary hippocampal neurons. Our data support a role of a nutrient-sensitive STK4–LC3B–FYCO1 axis in the regulation of the directional transport of autophagosomes via the post-translational regulation of LC3B. Given that autophagy is impaired in many human conditions, including neurodegenerative diseases, our findings may highlight new principles of vesicle transport regulation critical for disease etiology.Graphical abstract

scholarly journals Phospholipase D2 Localizes to the Plasma Membrane and Regulates Angiotensin II Receptor Endocytosis

2004 ◽  
Vol 15 (3) ◽  
pp. 1024-1030 ◽  
Author(s):  
Guangwei Du ◽  
Ping Huang ◽  
Bruce T. Liang ◽  
Michael A. Frohman
Keyword(s):  
Plasma Membrane ◽  
Angiotensin Ii ◽  
Membrane Vesicle ◽  
Membrane Vesicles ◽  
Cell Types ◽  
Dominant Negative ◽  
Secretory Cells ◽  
Resting Cells ◽  
Angiotensin Ii Receptor ◽  
Multiple Cell

Phospholipase D (PLD) is a key facilitator of multiple types of membrane vesicle trafficking events. Two PLD isoforms, PLD1 and PLD2, exist in mammals. Initial studies based on overexpression studies suggested that in resting cells, human PLD1 localized primarily to the Golgi and perinuclear vesicles in multiple cell types. In contrast, overexpressed mouse PLD2 was observed to localize primarily to the plasma membrane, although internalization on membrane vesicles was observed subsequent to serum stimulation. A recent report has suggested that the assignment of PLD2 to the plasma membrane is in error, because the endogenous isoform in rat secretory cells was imaged and found to be present primarily in the Golgi apparatus. We have reexamined this issue by using a monoclonal antibody specific for mouse PLD2, and find, as reported initially using overexpression studies, that endogenous mouse PLD2 is detected most readily at the plasma membrane in multiple cell types. In addition, we report that mouse, rat, and human PLD2 when overexpressed all similarly localize to the plasma membrane in cell lines from all three species. Finally, studies conducted using overexpression of wild-type active or dominant-negative isoforms of PLD2 and RNA interference-mediated targeting of PLD2 suggest that PLD2 functions at the plasma membrane to facilitate endocytosis of the angiotensin II type 1 receptor.

scholarly journals Nitric Oxide Transiently Converts Synaptic Inhibition to Excitation in Retinal Amacrine Cells

2006 ◽  
Vol 95 (5) ◽  
pp. 2866-2877 ◽  
Author(s):  
Brian Hoffpauir ◽  
Emily McMains ◽  
Evanna Gleason
Keyword(s):  
Nitric Oxide ◽  
Hippocampal Neurons ◽  
Reversal Potential ◽  
Amacrine Cells ◽  
Cell Types ◽  
Synaptic Function ◽  
Positive Shift ◽  
Gabaergic Synapses ◽  
Multiple Cell

Nitric oxide (NO) is generated by multiple cell types in the vertebrate retina, including amacrine cells. We investigate the role of NO in the modulation of synaptic function using a culture system containing identified retinal amacrine cells. We find that moderate concentrations of NO alter GABAA receptor function to produce an enhancement of the GABA-gated current. Higher concentrations of NO also enhance GABA-gated currents, but this enhancement is primarily due to a substantial positive shift in the reversal potential of the current. Several pieces of evidence, including a similar effect on glycine-gated currents, indicate that the positive shift is due to an increase in cytosolic Cl−. This change in the chloride distribution is especially significant because it can invert the sign of GABA- and glycine-gated voltage responses. Furthermore, current- and voltage-clamp recordings from synaptic pairs of GABAergic amacrine cells demonstrate that NO transiently converts signaling at GABAergic synapses from inhibition to excitation. Persistence of the NO-induced shift in ECl− in the absence of extracellular Cl− indicates that the increase in cytosolic Cl− is due to release of Cl− from an internal store. An NO-dependent release of Cl− from an internal store is also demonstrated for rat hippocampal neurons indicating that this mechanism is not restricted to the avian retina. Thus signaling in the CNS can be fundamentally altered by an NO-dependent mobilization of an internal Cl− store.

scholarly journals Kinesin-mediated organelle translocation revealed by specific cellular manipulations.

1994 ◽  
Vol 127 (4) ◽  
pp. 1021-1039 ◽  
Author(s):  
F Feiguin ◽  
A Ferreira ◽  
K S Kosik ◽  
A Caceres
Keyword(s):  
Hippocampal Neurons ◽  
Brefeldin A ◽  
Growth Cones ◽  
Cell Periphery ◽  
Fluorescent Staining ◽  
Anterograde Transport ◽  
Membrane Bound ◽  
Living Neurons ◽  
Vacuolar System ◽  
Cultured Hippocampal Neurons

The distribution of membrane-bound organelles was studied in cultured hippocampal neurons after antisense oligonucleotide suppression of the kinesin-heavy chain (KHC). We observed reduced 3,3'-dihexyloxacarbocyanine iodide (DiOC6(3)) fluorescent staining in neurites and growth cones. In astrocytes, KHC suppression results in the disappearance of the DiOC6(3)-positive reticular network from the cell periphery, and a parallel accumulation of label within the cell center. On the other hand, mitochondria microtubules and microfilaments display a distribution that closely resembles that observed in control cells. KHC suppression of neurons and astrocytes completely inhibited the Brefeldin A-induced spreading and tubulation of the Golgi-associated structure enriched in mannose-6-phosphate receptors. In addition, KHC suppression prevents the low pH-induced anterograde redistribution of late endocytic structures. Taken collectively, these observations suggest that in living neurons, kinesin mediates the anterograde transport of tubulovesicular structures originated in the central vacuolar system (e.g., the endoplasmic reticulum) and that the regulation of kinesin-membrane interactions may be of key importance for determining the intracellular distribution of selected organelles.

scholarly journals Multiple cell types contribute to the atherosclerotic lesion fibrous cap by PDGFRβ and bioenergetic mechanisms

2021 ◽  
Vol 3 (2) ◽  
pp. 166-181 ◽  
Author(s):  
Alexandra A. C. Newman ◽  
Vlad Serbulea ◽  
Richard A. Baylis ◽  
Laura S. Shankman ◽  
Xenia Bradley ◽  
...  
Keyword(s):  
Atherosclerotic Lesion ◽  
Cell Types ◽  
Fibrous Cap ◽  
Multiple Cell

scholarly journals MyD88 Is Not Required for Muscle Injury-Induced Endochondral Heterotopic Ossification in a Mouse Model of Fibrodysplasia Ossificans Progressiva

Biomedicines ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 630
Author(s):  
Huili Lyu ◽  
Cody M. Elkins ◽  
Jessica L. Pierce ◽  
C. Henrique Serezani ◽  
Daniel S. Perrien
Keyword(s):  
Heterotopic Ossification ◽  
Muscle Injury ◽  
Cell Types ◽  
Fibrodysplasia Ossificans Progressiva ◽  
Inflammatory Signaling ◽  
Conditional Deletion ◽  
Pathway Crosstalk ◽  
Multiple Cell

Excess inflammation and canonical BMP receptor (BMPR) signaling are coinciding hallmarks of the early stages of injury-induced endochondral heterotopic ossification (EHO), especially in the rare genetic disease fibrodysplasia ossificans progressiva (FOP). Multiple inflammatory signaling pathways can synergistically enhance BMP-induced Smad1/5/8 activity in multiple cell types, suggesting the importance of pathway crosstalk in EHO and FOP. Toll-like receptors (TLRs) and IL-1 receptors mediate many of the earliest injury-induced inflammatory signals largely via MyD88-dependent pathways. Thus, the hypothesis that MyD88-dependent signaling is required for EHO was tested in vitro and in vivo using global or Pdgfrα-conditional deletion of MyD88 in FOP mice. As expected, IL-1β or LPS synergistically increased Activin A (ActA)-induced phosphorylation of Smad 1/5 in fibroadipoprogenitors (FAPs) expressing Alk2R206H. However, conditional deletion of MyD88 in Pdgfrα-positive cells of FOP mice did not significantly alter the amount of muscle injury-induced EHO. Even more surprisingly, injury-induced EHO was not significantly affected by global deletion of MyD88. These studies demonstrate that MyD88-dependent signaling is dispensable for injury-induced EHO in FOP mice.

scholarly journals Microtubule–microtubule sliding by kinesin-1 is essential for normal cytoplasmic streaming in Drosophila oocytes

2016 ◽  
Vol 113 (34) ◽  
pp. E4995-E5004 ◽  
Author(s):  
Wen Lu ◽  
Michael Winding ◽  
Margot Lakonishok ◽  
Jill Wildonger ◽  
Vladimir I. Gelfand
Keyword(s):  
Cytoplasmic Streaming ◽  
Cell Types ◽  
Nurse Cells ◽  
Wild Type ◽  
Actin Cortex ◽  
Microtubule Motor ◽  
Multiple Cell ◽  
Cytoplasmic Microtubules ◽  
Two Populations

Cytoplasmic streaming in Drosophila oocytes is a microtubule-based bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, a major microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule–microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants.

scholarly journals Single-cell mapping of focused ultrasound-transfected brain

Gene Therapy ◽  
2021 ◽  
Author(s):  
A. S. Mathew ◽  
C. M. Gorick ◽  
R. J. Price
Keyword(s):  
Single Cell ◽  
Focused Ultrasound ◽  
Cell Types ◽  
Brain Cell ◽  
Therapeutic Modality ◽  
Full Potential ◽  
Stress Genes ◽  
Multiple Cell ◽  
Differential Gene

AbstractGene delivery via focused ultrasound (FUS) mediated blood-brain barrier (BBB) opening is a disruptive therapeutic modality. Unlocking its full potential will require an understanding of how FUS parameters (e.g., peak-negative pressure (PNP)) affect transfected cell populations. Following plasmid (mRuby) delivery across the BBB with 1 MHz FUS, we used single-cell RNA-sequencing to ascertain that distributions of transfected cell types were highly dependent on PNP. Cells of the BBB (i.e., endothelial cells, pericytes, and astrocytes) were enriched at 0.2 MPa PNP, while transfection of cells distal to the BBB (i.e., neurons, oligodendrocytes, and microglia) was augmented at 0.4 MPa PNP. PNP-dependent differential gene expression was observed for multiple cell types. Cell stress genes were upregulated proportional to PNP, independent of cell type. Our results underscore how FUS may be tuned to bias transfection toward specific brain cell types in vivo and predict how those cells will respond to transfection.

scholarly journals Outflow Tract Formation—Embryonic Origins of Conotruncal Congenital Heart Disease

2021 ◽  
Vol 8 (4) ◽  
pp. 42
Author(s):  
Sonia Stefanovic ◽  
Heather C. Etchevers ◽  
Stéphane Zaffran
Keyword(s):  
Congenital Heart ◽  
Cardiac Development ◽  
Heart Defects ◽  
Dynamic Structure ◽  
Cell Types ◽  
Outflow Tract ◽  
Heart Tube ◽  
Heart Field ◽  
Multiple Cell ◽  
The Many

Anomalies in the cardiac outflow tract (OFT) are among the most frequent congenital heart defects (CHDs). During embryogenesis, the cardiac OFT is a dynamic structure at the arterial pole of the heart. Heart tube elongation occurs by addition of cells from pharyngeal, splanchnic mesoderm to both ends. These progenitor cells, termed the second heart field (SHF), were first identified twenty years ago as essential to the growth of the forming heart tube and major contributors to the OFT. Perturbation of SHF development results in common forms of CHDs, including anomalies of the great arteries. OFT development also depends on paracrine interactions between multiple cell types, including myocardial, endocardial and neural crest lineages. In this publication, dedicated to Professor Andriana Gittenberger-De Groot and her contributions to the field of cardiac development and CHDs, we review some of her pioneering studies of OFT development with particular interest in the diverse origins of the many cell types that contribute to the OFT. We also discuss the clinical implications of selected key findings for our understanding of the etiology of CHDs and particularly OFT malformations.

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