Characterization of a novel NKG2D and NKp46 double-mutant mouse reveals subtle variations in the NK cell repertoire

Sheppard, Sam; Triulzi, Chiara; Ardolino, Michele; Serna, Daniel; Zhang, Lily; Raulet, David H.; Guerra, Nadia

doi:10.1182/blood-2012-12-471607

Cited by 31 publications

(61 citation statements)

References 51 publications

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“…However, elevated levels of the Helios transcript were not observed in the Ncr1 gfp/gfp mouse (Ref. 31 and data not shown). We speculated that the reported elevation in the Helios transcript might result from the presence of the mutated Ncr1-Noé protein.…”

Section: Helios Transcript Is Elevated In Ncr1-noe´-expressing Cellsmentioning

confidence: 89%

“…In these mice, similarly to the Ncr1 gfp/gfp mice, NK cell hyperactivation was not observed and Helios was not overexpressed, either in the single or in the double knockout mice (31). The major difference between the Noé and Ncr1 gfp/gfp mice is that the Ncr1 protein was mutated in the Noé mice.…”

mentioning

confidence: 81%

“…Recently, another mouse was generated by crossing Nkg2d knockout mice with the Ncr1 gfp/gfp mice (31). In these mice, similarly to the Ncr1 gfp/gfp mice, NK cell hyperactivation was not observed and Helios was not overexpressed, either in the single or in the double knockout mice (31).…”

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confidence: 99%

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Expression, Function, and Molecular Properties of the Killer Receptor Ncr1-Noé

Glasner¹,

Šimić

Miklić

et al. 2015

The Journal of Immunology

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NK cells kill various cells using activating receptors, such as the natural cytotoxicity receptors (NCRs). NKp46 is a major NCR and is the only NCR expressed in mice (denoted Ncr1). Using Ncr1-deficient mice (Ncr1gfp/pfp) we demonstrated that Ncr1 controls various pathologies, and that in its absence Ncr1-related functions are impaired. In 2012, another Ncr1-related mouse was generated, named Noé, in which a random mutation, W32R, in position 32, impaired the Ncr1-Noé cell surface expression. Interestingly, in the Noé mice, Ncr1-dependent deficiencies were not observed. Additionally, the Noé-NK cells were hyperactivated, probably due to increased Helios expression, and the Noé mice demonstrate increased clearance of influenza and murine CMV. In contrast, in the Ncr1gfp/pfp mice infection with influenza was lethal and we show in the present study no difference in murine CMV infection between Ncr1gfp/pfp and wild-type (WT) mice. Because the foremost difference between the Noé and Ncr1gfp/gfp mice is the presence of a mutated Ncr1-Noé protein, we studied its properties. We show that Ncr1-Noé and various other Ncr1 mutants in position 32 can be expressed on the surface, albeit slowly and unstably, and that ligand recognition and function of the various Ncr1-Noé is similar to the WT Ncr1. We further show that the glycosylation pattern of Ncr1-Noé is aberrant, that the Ncr1-Noé proteins accumulate in the endoplasmic reticulum, and that the expression of Ncr1-Noé proteins, but not WT Ncr1, leads to increased Helios expression. Thus, we suggest that the NK hyperactivated phenotype observed in the Noé mice might result from the presence of the Ncr1-Noé protein.

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Section: Helios Transcript Is Elevated In Ncr1-noe´-expressing Cellsmentioning

confidence: 89%

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confidence: 81%

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Expression, Function, and Molecular Properties of the Killer Receptor Ncr1-Noé

Glasner¹,

Šimić

Miklić

et al. 2015

The Journal of Immunology

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“…Ncr1 gfp/gfp mice appear to exhibit slightly reduced IFN-γ responses to NK1.1 stimulation in vitro and murine cytomegalovirus infection. 19,37 Stimulation through the NKp46 receptor has been shown to drive IFN-γ production. This NK cell-driven IFN-γ production has recently been shown to induce the extracellular matrix protein fibronectin 1 that could modulate tumor formation.…”

Section: Discussionmentioning

confidence: 99%

“…12,17,18 These lines ( Ncr gfp/gfp , Ncr1 Noe/Noe and Ncr1 iCre/iCre ) exhibit different phenotypes, possibly as a consequence of their difference in NKp46 protein expression. For example, Ncr1 Noe/Noe and Ncr1 iCre/iCre strains harbor modest gene changes but preserve the protein while Ncr1 gfp/gfp mice, 17 and Ncr1 gfp/gfp Klrk1 −/- double knockout mice which have defects in both NKp46 and NKG2D expression, 19 lack exons 5–7 resulting in a complete disruption of the protein.…”

Section: Introductionmentioning

confidence: 99%

A point mutation in the Ncr1 signal peptide impairs the development of innate lymphoid cell subsets

et al. 2018

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NKp46 (CD335) is a surface receptor shared by both human and mouse natural killer (NK) cells and innate lymphoid cells (ILCs) that transduces activating signals necessary to eliminate virus-infected cells and tumors. Here, we describe a spontaneous point mutation of cysteine to arginine (C14R) in the signal peptide of the NKp46 protein in congenic Ly5.1 mice and the newly generated NCRB6C14R strain. Ly5.1C14R NK cells expressed similar levels of Ncr1 mRNA as C57BL/6, but showed impaired surface NKp46 and reduced ability to control melanoma tumors in vivo. Expression of the mutant NKp46C14R in 293T cells showed that NKp46 protein trafficking to the cell surface was compromised. Although Ly5.1C14R mice had normal number of NK cells, they showed an increased number of early maturation stage NK cells. CD49a+ILC1s were also increased but these cells lacked the expression of TRAIL. ILC3s that expressed NKp46 were not detectable and were not apparent when examined by T-bet expression. Thus, the C14R mutation reveals that NKp46 is important for NK cell and ILC differentiation, maturation and function.SignificanceInnate lymphoid cells (ILCs) play important roles in immune protection. Various subsets of ILCs express the activating receptor NKp46 which is capable of recognizing pathogen derived and tumor ligands and is necessary for immune protection. Here, we describe a spontaneous point mutation in the signal peptide of the NKp46 protein in congenic Ly5.1 mice which are widely used for tracking cells in vivo. This Ncr1 C14R mutation impairs NKp46 surface expression resulting in destabilization of Ncr1 and accumulation of NKp46 in the endoplasmic reticulum. Loss of stable NKp46 expression impaired the maturation of NKp46+ ILCs and altered the expression of TRAIL and T-bet in ILC1 and ILC3, respectively.

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The Power of Three: Nanomaterials for Natural Killer (NK) Cell Immunoengineering Maximize Their Potency if They Exploit Multireceptor Stimulation

Dodd,

Guerra,

Dunlop

2024

Adv Healthcare Materials

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Many emerging cancer treatments are immunotherapies that modulate Natural Killer‐ (NK) or T cell activation, posing a challenge to develop immunoengineering nanomaterials that improve on the performance of molecular reagents. In physiological activation, multiple immunoreceptors signal in consort, however current biomaterials do not replicate this. Here, we create for the first time NK cell activating bionanomaterials that stimulate >2 immunoreceptors. We exploit nanoclusters of monoclonal antibodies (mAb), templated by nanoscale graphene oxide sheets (NGO) (∼75 nm size). To inform nanoreagent design, we first investigated a model system of planar substrates with anchored mAb. Combining mAb that stimulate 3 NK cell activating receptors (αNKP46 + αNKG2D + αDNAM‐1), activated NK cells more potently than any single receptor or pair. Applying this insight, we created an NGO‐mAb nanocluster that combined three distinct mAb: NGO‐mAb(αNKP46 + αNKG2D + αDNAM‐1). This construct was potent and outperformed single‐receptor‐simulating nanoclusters, activating nearly twice as many NK cells as NGO‐mAb(αNKP46) at a similar mAb dose, or delivering similar activation at 10x lower dosage. Furthermore, NGO‐mAb were more potent than planar substrates for both single‐ and triple‐mAb stimulation. Our results imply a new concept for immunoengineering biomaterials: both nanoclustering and multi‐receptor stimulation should be incorporated for maximum effect.This article is protected by copyright. All rights reserved

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Characterization of a novel NKG2D and NKp46 double-mutant mouse reveals subtle variations in the NK cell repertoire

Cited by 31 publications

References 51 publications

Expression, Function, and Molecular Properties of the Killer Receptor Ncr1-Noé

Expression, Function, and Molecular Properties of the Killer Receptor Ncr1-Noé

A point mutation in the Ncr1 signal peptide impairs the development of innate lymphoid cell subsets

The Power of Three: Nanomaterials for Natural Killer (NK) Cell Immunoengineering Maximize Their Potency if They Exploit Multireceptor Stimulation

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