PE-STOP: A versatile tool for installing nonsense substitutions amenable for precise reversion.

The rapid advances in genome editing technologies have revolutionized the study of gene functions in cell or animal models. The recent generation of double-stranded DNA cleavage-independent base editors has been suitably adapted for interrogation of protein-coding genes on the basis of introducing premature stop codons or disabling the start codons. However, such versions of stop/start codon-oriented genetic tools still present limitations on their versatility, base-level precision, and target specificity.

IL-6/ERK signaling pathway participates in type I IFN-programmed, unconventional M2-like macrophage polarization.

Type I interferons (IFN-Is) have been harnessed for cancer therapies due to their immunostimulatory functions. However, certain tumor-tolerating activities by IFN-Is also exist, and may potentially thwart their therapeutic effects. In this respect, our previous studies have demonstrated a monocyte-orchestrated, IFN-I-to-IL-4 cytokine axis, which can subsequently drive M2-skewed pro-tumoral polarization of macrophages.

Development of a versatile nuclease prime editor with upgraded precision.

The applicability of nuclease-based form of prime editor (PEn) has been hindered by its complexed editing outcomes. A chemical inhibitor against DNA-PK, which mediates the nonhomologous end joining (NHEJ) pathway, was recently shown to promote precise insertions by PEn. Nevertheless, the intrinsic issues of specificity and toxicity for such a chemical approach necessitate development of alternative strategies.

A New Mechanism for Early-Time Plasmaspheric Refilling: The Role of Charge Exchange Reactions in the Transport of Energy and Mass Throughout the Ring Current-Plasmasphere System.

Cold H produced via charge exchange reactions between ring current ions and exospheric neutral hydrogen constitutes an additional source of cold plasma that further contributes to the plasmasphere and affects the plasma dynamics in the Earth's magnetosphere system; however, its production and associated effects on the plasmasphere dynamics have not been fully assessed and quantified. In this study, we perform numerical simulations mimicking an idealized three-phase geomagnetic storm to investigate the role of heavy ion composition in the ring current (O vs. N) and exospheric neutral hydrogen density in the production of cold H via charge exchange reactions.

Precise somatic genome editing for treatment of inborn errors of immunity.

Rapid advances in high throughput sequencing have substantially expedited the identification and diagnosis of inborn errors of immunity (IEI). Correction of faulty genes in the hematopoietic stem cells can potentially provide cures for the majority of these monogenic immune disorders. Given the clinical efficacies of vector-based gene therapies already established for certain groups of IEI, the recently emerged genome editing technologies promise to bring safer and more versatile treatment options.

Enhancement of prime editing via xrRNA motif-joined pegRNA.

The prime editors (PEs) have shown great promise for precise genome modification. However, their suboptimal efficiencies present a significant technical challenge. Here, by appending a viral exoribonuclease-resistant RNA motif (xrRNA) to the 3'-extended portion of pegRNAs for their increased resistance against degradation, we develop an upgraded PE platform (xrPE) with substantially enhanced editing efficiencies in multiple cell lines.

Precise tumor immune rewiring via synthetic CRISPRa circuits gated by concurrent gain/loss of transcription factors.

Reinvigoration of antitumor immunity has recently become the central theme for the development of cancer therapies. Nevertheless, the precise delivery of immunotherapeutic activities to the tumors remains challenging. Here, we explore a synthetic gene circuit-based strategy for specific tumor identification, and for subsequently engaging immune activation.

A Monocyte-Orchestrated IFN-I-to-IL-4 Cytokine Axis Instigates Protumoral Macrophages and Thwarts Poly(I:C) Therapy.

Type I IFNs (IFN-I) are important for tumor immune surveillance and contribute to the therapeutic responses for numerous treatment regimens. Nevertheless, certain protumoral activities by IFN-I have been increasingly recognized. Indeed, our recent work showed that systemic poly(I:C)/IFN treatment can undesirably trigger high arginase (ARG1) expression within the tumor-associated monocyte/macrophage compartment.

Internalization Characterization of Si Nanorod with Camouflaged Cell Membrane Proteins Reveals as a Negative Regulator.

The fabrication of shape-controlled nanocarriers is critical for efficient delivery of biomolecules across the cell membrane. Surface coating of the nanocarrier can improve internalization efficiency. Here, we developed a facile method of silicon nanorod fabrication leading to a controlled size and shape.

Golgi-resident TRIO regulates membrane trafficking during neurite outgrowth.

Neurite outgrowth requires coordinated cytoskeletal rearrangements in the growth cone and directional membrane delivery from the neuronal soma. As an essential Rho guanine nucleotide exchange factor (GEF), TRIO is necessary for cytoskeletal dynamics during neurite outgrowth, but its participation in the membrane delivery is unclear. Using co-localization studies, live-cell imaging, and fluorescence recovery after photobleaching analysis, along with neurite outgrowth assay and various biochemical approaches, we here report that in mouse cerebellar granule neurons, TRIO protein pools at the Golgi and regulates membrane trafficking by controlling the directional maintenance of both RAB8 (member RAS oncogene family 8)- and RAB10-positive membrane vesicles.

Concomitant type I IFN and M-CSF signaling reprograms monocyte differentiation and drives pro-tumoral arginase production.

Background: Type I IFN-based therapies against solid malignancies have yielded only limited success. How IFN affects tumor-associated macrophage (TAM) compartment to impact the therapeutic outcomes are not well understood.

Methods: The effect of an IFN-inducer poly(I:C) on tumor-infiltrating monocytes and TAMs were analyzed using a transplantable mouse tumor model (LLC).

PFKFB3-Driven Macrophage Glycolytic Metabolism Is a Crucial Component of Innate Antiviral Defense.

Signaling by viral nucleic acids and subsequently by type I IFN is central to antiviral innate immunity. These signaling events are also likely to engage metabolic changes in immune and nonimmune cells to support antiviral defense. In this study, we show that cytosolic viral recognition, by way of secondary IFN signaling, leads to upregulation of glycolysis preferentially in macrophages.

Targeted Gene Delivery to Macrophages by Biodegradable Star-Shaped Polymers.

In this report, two biodegradable star-shaped polyasparamide derivatives and four analogues modified with either mannose or folic acid moiety for preferential targeting of a difficult-to-transfect immune cell type, i.e., macrophage, have been synthesized.

Rapamycin-sensitive mTORC1 signaling is involved in physiological primordial follicle activation in mouse ovary.

In mammals, resting female oocytes reside in primordial ovarian follicles. An individual primordial follicle may stay quiescent for a protracted period of time before initiating follicular growth, which is also termed “activation.” Female reproductive capacity is sustained by the gradual, streamlined activation of the entire population of primordial follicles, but this process also results in reproductive senescence in older animals.

Regulation of interleukin-10 receptor ubiquitination and stability by beta-TrCP-containing ubiquitin E3 ligase.

Interleukin-10 (IL-10) initiates potent anti-inflammatory effects via activating its cell surface receptor, composed of IL-10R1 and IL-10R2 subunits. The level of IL-10R1 is a major determinant of the cells' responsiveness to IL-10. Here, via a series of biochemical analyses using 293T cells reconstituted with IL-10R1, we identify the latter as a novel substrate of βTrCP-containing ubiquitin E3 ligase.

Pathogen recognition receptor signaling accelerates phosphorylation-dependent degradation of IFNAR1.

An ability to sense pathogens by a number of specialized cell types including the dendritic cells plays a central role in host's defenses. Activation of these cells through the stimulation of the pathogen-recognition receptors induces the production of a number of cytokines including Type I interferons (IFNs) that mediate the diverse mechanisms of innate immunity. Type I IFNs interact with the Type I IFN receptor, composed of IFNAR1 and IFNAR2 chains, to mount the host defense responses.

Inducible priming phosphorylation promotes ligand-independent degradation of the IFNAR1 chain of type I interferon receptor.

Phosphorylation-dependent ubiquitination and ensuing down-regulation and lysosomal degradation of the interferon alpha/beta receptor chain 1 (IFNAR1) of the receptor for Type I interferons play important roles in limiting the cellular responses to these cytokines. These events could be stimulated either by the ligands (in a Janus kinase-dependent manner) or by unfolded protein response (UPR) inducers including viral infection (in a manner dependent on the activity of pancreatic endoplasmic reticulum kinase). Both ligand-dependent and -independent pathways converge on phosphorylation of Ser(535) within the IFNAR1 degron leading to recruitment of beta-Trcp E3 ubiquitin ligase and concomitant ubiquitination and degradation.

Mammalian casein kinase 1alpha and its leishmanial ortholog regulate stability of IFNAR1 and type I interferon signaling.

Phosphorylation of the degron of the IFNAR1 chain of the type I interferon (IFN) receptor triggers ubiquitination and degradation of this receptor and, therefore, plays a crucial role in negative regulation of IFN-alpha/beta signaling. Besides the IFN-stimulated and Jak activity-dependent pathways, a basal ligand-independent phosphorylation of IFNAR1 has been described and implicated in downregulating IFNAR1 in response to virus-induced endoplasmic reticulum (ER) stress. Here we report purification and characterization of casein kinase 1alpha (CK1alpha) as a bona fide major IFNAR1 kinase that confers basal turnover of IFNAR1 and cooperates with ER stress stimuli to mediate phosphorylation-dependent degradation of IFNAR1.

Virus-induced unfolded protein response attenuates antiviral defenses via phosphorylation-dependent degradation of the type I interferon receptor.

Phosphorylation-dependent ubiquitination and degradation of the IFNAR1 chain of the type I interferon (IFN) receptor is regulated by two different pathways, one of which is ligand independent. We report that this ligand-independent pathway is activated by inducers of unfolded protein responses (UPR), including viral infection, and that such activation requires the endoplasmic reticulum-resident protein kinase PERK. Upon viral infection, activation of this pathway promotes phosphorylation-dependent ubiquitination and degradation of IFNAR1, specifically inhibiting type I IFN signaling and antiviral defenses.

Cigarette smoking products suppress anti-viral effects of Type I interferon via phosphorylation-dependent downregulation of its receptor.

While negative effect of smoking on the resistance to viral infections was known, the underlying mechanisms remained unclear. Here we report that products of cigarette smoking compromise the cellular anti-viral defenses by inhibiting the signaling induced by Type I interferon (IFN). Cigarette smoking condensate (but not pure nicotine) stimulated specific serine phosphorylation-dependent ubiquitination and degradation of the IFNAR1 subunit of the Type I IFN receptor leading to attenuation of IFN signaling and decreased resistance to viral infection.

Ligand-independent pathway that controls stability of interferon alpha receptor.

Ligand-specific negative regulation of cytokine-induced signaling relies on down regulation of the cytokine receptors. Down regulation of the IFNAR1 sub-unit of the Type I interferon (IFN) receptor proceeds via lysosomal receptor proteolysis, which is triggered by ubiquitination that depends on IFNAR1 serine phosphorylation. While IFN-inducible phosphorylation, ubiquitination, and degradation requires the catalytic activity of the Tyk2 Janus kinase, here we found the ligand- and Tyk2-independent pathway that promotes IFNAR1 phosphorylation, ubiquitination, and degradation when IFNAR1 is expressed at high levels.

Site-specific ubiquitination exposes a linear motif to promote interferon-alpha receptor endocytosis.

Ligand-induced endocytosis and lysosomal degradation of cognate receptors regulate the extent of cell signaling. Along with linear endocytic motifs that recruit the adaptin protein complex 2 (AP2)-clathrin molecules, monoubiquitination of receptors has emerged as a major endocytic signal. By investigating ubiquitin-dependent lysosomal degradation of the interferon (IFN)-alpha/beta receptor 1 (IFNAR1) subunit of the type I IFN receptor, we reveal that IFNAR1 is polyubiquitinated via both Lys48- and Lys63-linked chains.