SIDT2 Transports Extracellular dsRNA into the Cytoplasm for Innate Immune Recognition

Nguyen, Tracy; Smith, Blake R. C.; Tate, Michelle D.; Belz, Gabrielle T.; Barrios, Marilou H.; Elgass, Kirstin; Weisman, Alexandra S.; Baker, Paul J.; Preston, Simon; Whitehead, Lachlan; Garnham, Alexandra L.; Lundie, Rachel J.; Smyth, Gordon K.; Pellegrini, Marc; O’Keeffe, Meredith; Wicks, Ian P.; Masters, Seth L.; Hunter, Craig P.; Pang, Ken C

doi:10.1016/j.immuni.2017.08.007

Cited by 124 publications

(134 citation statements)

References 78 publications

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“…SIDT2, a mammalian ortholog of the Caenorhabditis elegans dsRNA transporter SID-1, localizes to endolysosomes and functions as a transporter of dsRNA, promoting escape from intracellular compartments [1]. Sixty minutes after its addition, poly(I:C) accumulates in the endosomal compartments of Sidt2 knockout bone marrow-derived DC, whereas poly(I:C) is diffuse in wild-type (WT) cells.…”

Section: Internalization Of Extracellular Rna Into the Endosome And Cmentioning

confidence: 99%

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Extracellular RNA Sensing by Pattern Recognition Receptors

et al. 2018

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RNA works as a genome and messenger in RNA viruses, and it sends messages in most of the creatures of the Earth, including viruses, bacteria, fungi, plants, and animals. The human innate immune system has evolved to detect single- and double-stranded RNA molecules from microbes by pattern recognition receptors and induce defense reactions against infections such as the production of type I interferons and inflammatory cytokines. To avoid cytokine toxicity causing chronic inflammation or autoimmunity by sensing self-RNA, the activation of RNA sensors is strictly regulated. All of the Toll-like receptors that recognize RNA are localized to endosomes/lysosomes, which require internalization of RNA for sensing through an endocytic pathway. RIG-I-like receptors sense RNA in cytosol. These receptors are expressed in a cell type-specific fashion, enabling sensing of RNA for a wide range of microbial invasions. At the same time, both endosomal and cytoplasmic receptors have strategies to respond only to RNA of pathogenic microorganisms or dying cells. RNA are potential vaccine adjuvants for immune enhancement against cancer and provide a benefit for vaccinations. Understanding the detailed molecular mechanisms of the RNA-sensing system will help us to broaden the clinical utility of RNA adjuvants for patients with incurable diseases.

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Section: Internalization Of Extracellular Rna Into the Endosome And Cmentioning

confidence: 99%

“…However, although extracellular RNA activate RLR-mediated signaling, the mechanisms via which RNA reach cytosolic sensors is unclear. One component that may be involved is SIDT2, which was recently reported to play a role in trafficking of dsRNA from the endosome to the cytosol after TLR3 activation [1] (Fig. 1).…”

Section: Introductionmentioning

confidence: 99%

Extracellular RNA Sensing by Pattern Recognition Receptors

et al. 2018

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“…5e). These findings suggest a connection between synaptic plasticity and cellular transfer of RNA as Sidt1 and Sidt2 bind and transport double-stranded RNA 58,59 .…”

Section: Homeostatic Plasticity Induces Reorganization Of the Neuronamentioning

confidence: 83%

Surfaceome dynamics during neuronal development and synaptic plasticity reveal system-wide surfaceome reorganization independent of global proteostasis

Oostrum

Campbell

Müller

et al. 2019

Preprint

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Neurons are highly compartmentalized cells with tightly controlled subcellular protein organization. While broad brain transcriptome, connectome and global proteome maps are being generated, system-wide analysis of temporal protein dynamics at the subcellular level are currently lacking for neuronal development and synapse formation. We performed a temporally-resolved surfaceome analysis of developing primary neuron cultures to a depth of 1000 bona fide surface proteins and reveal dynamic surface protein clusters that reflect the functional requirements during distinct stages of neuronal development. Moreover, our data shows that synaptic proteins are globally trafficked to the surface prior to synapse formation. Direct comparison of surface and total protein pools demonstrates that, depending on the time scale, surface abundance changes can correlate or differ from total protein abundance. The uncoupling of surface and total abundance changes has direct functional implications as shown in the context of synaptic vesicle transport. To demonstrate the utility of our approach we analyzed the surfaceome modulation in response to homeostatic synaptic scaling and found dynamic remodeling of the neuronal surface, which was largely independent of global proteostasis, indicative of wide-spread regulation on the level of surface trafficking. Finally, we present a quantitative analysis of the neuronal surface during early-phase long-term potentiation (LTP) and reveal fast externalization of diverse classes of surface proteins beyond the AMPA receptor, providing new insights into the requirement of exocytosis for LTP. Our resource and finding of organizational principles highlight the importance of subcellular resolution for systems-level understanding of cellular processes, which are typically masked by broad omics-style approaches.Recently, we reported a miniaturization and automation of the Cell Surface Capture (autoCSC) method, enabling sensitive and multiplexed interrogation of the surfaceome landscape of primary cells by direct identification of extracellular N-glycopeptides 30,31 . On living cells, cell-surface carbohydrates are tagged with cell-impermeable biocytin-hydrazide. After cell lysis and tryptic digestion, glycopeptides are captured using streptavidin-filled tips using a process which is automated by using a liquid handling robot. After peptide:N-glycosidase F (PNGase F) treatment, which catalyzes the cleavage of asparagine-linked glycans, de-glycosylated peptides are eluted. This leaves a deamidation within the N-X-S/T consensus sequence of formerly N-glycosylated peptides, enabling specific identification of extracellular N-glycosylation sites and quantification of protein abundance with subcellular resolution by data-independent acquisition (DIA) mass spectrometry (MS) 32 . The main advantage of this chemoproteomic strategy focusing on

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“…Furthermore, it is possible that other nucleotide species such as RNA:DNA hybrids [11][12][13] and long dsRNAs [17] are stained by the PicoGreen variants.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of PicoGreen variants for use in microscopy and flow cytometry

Khatoo

Yang

Gee

et al. 2019

Preprint

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PicoGreen is a fluorescent probe that binds dsDNA and forms a highly luminescent complex when compared to the free dye in solution. This unique probe is widely used in DNA quantitation assays but has limited application in flow cytometry and microscopy. Here we have investigated various PicoGreen variants for the ability to stain low amounts of cytosolic DNA present in many tumor cells. Analysis of stained cells by flow cytometry and fluorescent microscopy showed that certain variants improved the ability to stain low levels of cytosolic DNA when compared to the commercially available PicoGreen molecule.

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SIDT2 Transports Extracellular dsRNA into the Cytoplasm for Innate Immune Recognition

Cited by 124 publications

References 78 publications

Extracellular RNA Sensing by Pattern Recognition Receptors

Extracellular RNA Sensing by Pattern Recognition Receptors

Surfaceome dynamics during neuronal development and synaptic plasticity reveal system-wide surfaceome reorganization independent of global proteostasis

Evaluation of PicoGreen variants for use in microscopy and flow cytometry

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