OZITX, a pertussis toxin-like protein for occluding inhibitory G protein signalling including Gαz

Keen, Alastair C.; Pedersen, Maria; Lemel, Laura; Scott, Daniel J.; Canals, Meritxell; Littler, Dene R.; Beddoe, Travis; Ono, Yukiko; Shi, Lei; Inoue, Asuka; Javitch, Jonathan A.; Lane, J. Robert

doi:10.1038/s42003-022-03191-5

Cited by 8 publications

(9 citation statements)

References 51 publications

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“…Furthermore, PTX is incapable of inhibiting Gαz. An Escherichia coli pertussis toxin-like AB 5 toxin called OZITX was recently found to inhibit all Gαi subfamily proteins, including Gαz 30 , 31 . Even with this toxin, a Gαi C-terminal mutation, which may affect coupling properties, is required to make Gαi subfamily proteins insensitive to OZITX.…”

Section: Discussionmentioning

confidence: 99%

Generation of Gαi knock-out HEK293 cells illuminates Gαi-coupling diversity of GPCRs

et al. 2023

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G-protein-coupled receptors (GPCRs) are pivotal cell membrane proteins that sense extracellular molecules and activate cellular responses. The G-protein α subunit i (Gαi) family represents the most common GPCR-coupling partner and consists of eight subunits with distinct signaling properties. However, analyzing the coupling pattern has been challenging owing to endogenous expression of the Gαi subunits in virtually all cell lines. Here, we generate a HEK293 cell line lacking all Gαi subunits, which enables the measurement of GPCR-Gαi coupling upon transient re-expression of a specific Gαi subunit. We profile Gαi-coupling selectivity across 11 GPCRs by measuring ligand-induced inhibitory activity for cAMP accumulation. The coupling profiles are then classified into three clusters, representing those preferentially coupled to Gαz, those to Gαo, and those with unapparent selectivity. These results indicate that individual Gαi-coupled GPCRs fine-tune Gαi signaling by exerting coupling preference at the Gαi-subunit level.

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Section: Discussionmentioning

confidence: 99%

Generation of Gαi knock-out HEK293 cells illuminates Gαi-coupling diversity of GPCRs

et al. 2023

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“…To eliminate Gi/o signaling influence, this experiment was conducted in the presence of 0.05 μg/mL pertussis toxin. 74 Upon melanopsin activation, Lyn-mRFP expressing RAW264.7 cells showed a minor migration away from the blue light (Figure S7D, top row). The cells expressing Lyn-mRFP-dHTH showed significantly higher migration than the migration observed with Lyn-mRFP (Figure S7E) (one-way ANOVA: F 1, 10 = 14.0369, p = 0.00381; Table S15A,B).…”

Section: Opto-dhth Does Not Disrupt Gβγ Signaling or Gαqgtp-grk2 Inte...mentioning

confidence: 97%

Spatiotemporal Optical Control of Gαq-PLCβ Interactions

Ubeysinghe,

Kankanamge,

Thotamune

et al. 2023

ACS Synth. Biol.

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Cells experience time-varying and spatially heterogeneous chemokine signals in vivo, activating cell surface proteins including G protein-coupled receptors (GPCRs). The Gαq pathway activation by GPCRs is a major signaling axis with broad physiological and pathological significance. Compared with other Gα members, GαqGTP activates many crucial effectors, including PLCβ (Phospholipase Cβ) and Rho GEFs (Rho guanine nucleotide exchange factors). PLCβ regulates many key processes, such as hematopoiesis, synaptogenesis, and cell cycle, and is therefore implicated in terminal-debilitating diseases, including cancer, epilepsy, Huntington's Disease, and Alzheimer's Disease. However, due to a lack of genetic and pharmacological tools, examining how the dynamic regulation of PLCβ signaling controls cellular physiology has been difficult. Since activated PLCβ induces several abrupt cellular changes, including cell morphology, examining how the other pathways downstream of Gq-GPCRs contribute to the overall signaling has also been difficult. Here we show the engineering, validation, and application of a highly selective and efficient optogenetic inhibitor (Opto-dHTH) to completely disrupt GαqGTP-PLCβ interactions reversibly in user-defined cellular-subcellular regions on optical command. Using this newly gained PLCβ signaling control, our data indicate that the molecular competition between RhoGEFs and PLCβ for GαqGTP determines the potency of Gq-GPCR-governed directional cell migration.

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“…In contrast to pathogenicity factors, which require the presence of living bacteria, bacterial toxins can exert their function once released from the pathogen and often the toxin is sufficient to cause disease. Many bacterial toxins target molecules of the host that are central for the cell to survive, replicate or exert its function, for example actin, Rho GTPases or heterotrimeric G proteins (74)(75)(76). Typical modifications are ADP ribosylation (74), glycosylation (77), but other functional modifications that can change the activation status of the target molecule, like deamidation (78) are also described.…”

Section: Bacterial Pathogenicity Factorsa Short Introductionmentioning

confidence: 99%

Pasteurella multocida toxin – lessons learned from a mitogenic toxin

Kubatzky

2022

Front. Immunol.

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The gram-negative, zoonotic bacterium Pasteurella multocida was discovered in 1880 and found to be the causative pathogen of fowl cholera. Pasteurella-related diseases can be found in domestic and wild life animals such as buffalo, sheep, goat, deer and antelope, cats, dogs and tigers and cause hemorrhagic septicemia in cattle, rhinitis or pneumonia in rabbits or fowl cholera in poultry and birds. Pasteurella multocida does not play a major role in the immune-competent human host, but can be found after animal bites or in people with close contact to animals. Toxigenic strains are most commonly found in pigs and express a phage-encoded 146 kDa protein, the Pasteurella multocida toxin (PMT). Toxin-expressing strains cause atrophic rhinitis where nasal turbinate bones are destroyed through the inhibition of bone building osteoblasts and the activation of bone resorbing osteoclasts. After its uptake through receptor-mediated endocytosis, PMT specifically targets the alpha subunit of several heterotrimeric G proteins and constitutively activates them through deamidation of a glutamine residue to glutamate in the alpha subunit. This results in cytoskeletal rearrangement, proliferation, differentiation and survival of cells. Because of the toxin’s mitogenic effects, it was suggested that it might have carcinogenic properties, however, no link between Pasteurella infections and cell transformation could be established, neither in tissue culture models nor through epidemiological data. In the recent years it was shown that the toxin not only affects bone, but also the heart as well as basically all cells of innate and adaptive immunity. During the last decade the focus of research shifted from signal transduction processes to understanding how the bacteria might benefit from a bone-destroying toxin. The primary function of PMT seems to be the modulation of immune cell activation which at the same time creates an environment permissive for osteoclast formation. While the disease is restricted to pigs, the implications of the findings from PMT research can be used to explore human diseases and have a high translational potential. In this review our current knowledge will be summarized and it will be discussed what can be learned from using PMT as a tool to understand human pathologies.

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OZITX, a pertussis toxin-like protein for occluding inhibitory G protein signalling including Gαz

Cited by 8 publications

References 51 publications

Generation of Gαi knock-out HEK293 cells illuminates Gαi-coupling diversity of GPCRs

Generation of Gαi knock-out HEK293 cells illuminates Gαi-coupling diversity of GPCRs

Spatiotemporal Optical Control of Gαq-PLCβ Interactions

Pasteurella multocida toxin – lessons learned from a mitogenic toxin

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