Tryptophan C-mannosylation is critical for Plasmodium falciparum transmission.

Tryptophan C-mannosylation stabilizes proteins bearing a thrombospondin repeat (TSR) domain in metazoans. Here we show that Plasmodium falciparum expresses a DPY19 tryptophan C-mannosyltransferase in the endoplasmic reticulum and that DPY19-deficiency abolishes C-glycosylation, destabilizes members of the TRAP adhesin family and inhibits transmission to mosquitoes. Imaging P. falciparum gametogenesis in its entirety in four dimensions using lattice light-sheet microscopy reveals defects in ΔDPY19 gametocyte egress and exflagellation. While egress is diminished, ΔDPY19 microgametes still fertilize macrogametes, forming ookinetes, but these are abrogated for mosquito infection. The gametogenesis defects correspond with destabilization of MTRAP, which we show is C-mannosylated in P. falciparum, and the ookinete defect is concordant with defective CTRP secretion on the ΔDPY19 background. Genetic complementation of DPY19 restores ookinete infectivity, sporozoite production and C-mannosylation activity. Therefore, tryptophan C-mannosylation by DPY19 ensures TSR protein quality control at two lifecycle stages for successful transmission of the human malaria parasite.

Epitope-coated polymer particles elicit neutralising antibodies against Plasmodium falciparum sporozoites.

The current Malaria RTS,S vaccine is based on virus-like particles (VLPs) comprising the NANP repetitive epitopes from the cicumsporozoite protein (CSP) of Plasmodium falciparum. This vaccine has limited efficacy, only preventing severe disease in about 30% of vaccinated individuals. A more efficacious vaccine is urgently needed to combat malaria. Here we developed a particulate malaria vaccine based on the same CSP epitopes but using biopolymer particles (BPs) as an antigen carrier system. Specific B- and T-cell epitope-coated BPs were assembled in vivo inside an engineered endotoxin-free mutant of Escherichia coli. A high-yield production process leading to ~27% BP vaccine weight over biomass was established. The epitope-coated BPs were purified and their composition, i.e., the polymer core and epitope identity, was confirmed. Epitope-coated BPs were used alongside soluble peptide epitopes and empty BPs to vaccinate sheep. Epitope-coated BPs showed enhanced immunogenicity by inducing anti-NANP antibody titre of EC50 > 150,000 that were at least 20 times higher than induced by the soluble peptides. We concluded that the additional T-cell epitope was not required as it did not enhance immunogenicity when compared with the B-cell epitope-coated BPs. Antibodies specifically bound to the surface of Plasmodium falciparum sporozoites and efficiently inhibited sporozoite motility and traversal of human hepatocytes. This study demonstrated the utility of biologically self-assembled epitope-coated BPs as an epitope carrier for inclusion in next-generation malaria vaccines.

Ptpn2 and KLRG1 regulate the generation and function of tissue-resident memory CD8+ T cells in skin.

Tissue-resident memory T cells (TRM cells) are key elements of tissue immunity. Here, we investigated the role of the regulator of T cell receptor and cytokine signaling, Ptpn2, in the formation and function of TRM cells in skin. Ptpn2-deficient CD8+ T cells displayed a marked defect in generating CD69+ CD103+ TRM cells in response to herpes simplex virus type 1 (HSV-1) skin infection. This was accompanied by a reduction in the proportion of KLRG1- memory precursor cells and a transcriptional bias toward terminal differentiation. Of note, forced expression of KLRG1 was sufficient to impede TRM cell formation. Normalizing memory precursor frequencies by transferring equal numbers of KLRG1- cells restored TRM generation, demonstrating that Ptpn2 impacted skin seeding with precursors rather than downstream TRM cell differentiation. Importantly, Ptpn2-deficient TRM cells augmented skin autoimmunity but also afforded superior protection from HSV-1 infection. Our results emphasize that KLRG1 repression is required for optimal TRM cell formation in skin and reveal an important role of Ptpn2 in regulating TRM cell functionality.

Inhibition of Plasmepsin V Activity Blocks Plasmodium falciparum Gametocytogenesis and Transmission to Mosquitoes.

Plasmodium falciparum gametocytes infect mosquitoes and are responsible for malaria transmission. New interventions that block transmission could accelerate malaria elimination. Gametocytes develop within erythrocytes and activate protein export pathways that remodel the host cell. Plasmepsin V (PMV) is an aspartyl protease that is required for protein export in asexual parasites, but its function and essentiality in gametocytes has not been definitively proven, nor has PMV been assessed as a transmission-blocking drug target. Here, we show that PMV is expressed and can be inhibited specifically in P. falciparum stage I-II gametocytes. PMV inhibitors block processing and export of gametocyte effector proteins and inhibit development of stage II-V gametocytes. Gametocytogenesis in the presence of sublethal inhibitor concentrations results in stage V gametocytes that fail to infect mosquitoes. Therefore, PMV primes gametocyte effectors for export, which is essential for the development and fitness of gametocytes for transmission to mosquitoes.

Author Correction: Tissue-resident memory CD8+ T cells promote melanoma-immune equilibrium in skin.

Panel j was inadvertently labelled as panel k in the caption to Fig. 4. Similarly, 'Fig. 4k' should have been 'Fig. 4j' in the sentence beginning 'TNF-α-deficient gBT-I cells were…'. In addition, the surname of author Umaimainthan Palendira was misspelled 'Palendria'. These errors have been corrected online.

Tissue-resident memory CD8+ T cells promote melanoma-immune equilibrium in skin.

The immune system can suppress tumour development both by eliminating malignant cells and by preventing the outgrowth and spread of cancer cells that resist eradication1. Clinical and experimental data suggest that the latter mode of control-termed cancer-immune equilibrium1-can be maintained for prolonged periods of time, possibly up to several decades2-4. Although cancers most frequently originate in epithelial layers, the nature and spatiotemporal dynamics of immune responses that maintain cancer-immune equilibrium in these tissue compartments remain unclear. Here, using a mouse model of transplantable cutaneous melanoma5, we show that tissue-resident memory CD8+ T cells (TRM cells) promote a durable melanoma-immune equilibrium that is confined to the epidermal layer of the skin. A proportion of mice (~40%) transplanted with melanoma cells remained free of macroscopic skin lesions long after epicutaneous inoculation, and generation of tumour-specific epidermal CD69+ CD103+ TRM cells correlated with this spontaneous disease control. By contrast, mice deficient in TRM formation were more susceptible to tumour development. Despite being tumour-free at the macroscopic level, mice frequently harboured melanoma cells in the epidermal layer of the skin long after inoculation, and intravital imaging revealed that these cells were dynamically surveyed by TRM cells. Consistent with their role in melanoma surveillance, tumour-specific TRM cells that were generated before melanoma inoculation conferred profound protection from tumour development independently of recirculating T cells. Finally, depletion of TRM cells triggered tumour outgrowth in a proportion (~20%) of mice with occult melanomas, demonstrating that TRM cells can actively suppress cancer progression. Our results show that TRM cells have a fundamental role in the surveillance of subclinical melanomas in the skin by maintaining cancer-immune equilibrium. As such, they provide strong impetus for exploring these cells as targets of future anticancer immunotherapies.