Metabolic Engineering of Salmonella Vaccine Bacteria To Boost Human Vγ2Vδ2 T Cell Immunity

Abstract

Human Vγ2Vδ2 T cells monitor isoprenoid metabolism by recognizing foreign (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP), a metabolite in the 2-C-methyl-D-erythritol-4-phosphate pathway used by most eubacteria and apicomplexan parasites, and self isopentenyl pyrophosphate, a metabolite in the mevalonate pathway used by humans. Whereas microbial infections elicit prolonged expansion of memory Vγ2Vδ2 T cells, immunization with prenyl pyrophosphates or aminobisphosphonates elicit short-term Vγ2Vδ2 expansion with rapid anergy and deletion upon subsequent immunizations. We hypothesized that a live, attenuated bacterial vaccine that overproduces HMBPP would elicit long lasting Vγ2Vδ2 T cell immunity by mimicking a natural infection. Therefore, we metabolically engineered the avirulent aroA− Salmonella enterica serovar Typhimurium SL7207 strain by deleting the gene for LytB (the downstream enzyme from HMBPP) and functionally complementing for this loss with genes encoding mevalonate pathway enzymes. LytB− Salmonella SL7207 had high HMBPP levels, infected human cells as efficiently as the wild-type bacteria, and stimulated large ex vivo expansions of Vγ2Vδ2 T cells from human donors. Importantly, vaccination of a rhesus monkey with live lytB− Salmonella SL7207 stimulated a prolonged expansion of Vγ2Vδ2 T cells without significant side effects or anergy induction. These studies provide proof-of-principle that metabolic engineering can be used to derive live bacterial vaccines that boost Vγ2Vδ2 T cell immunity. Similar engineering of metabolic pathways to produce lipid Ags or B vitamin metabolite Ags could be used to derive live bacterial vaccine for other unconventional T cells that recognize nonpeptide Ags.

Emerging role of γδ T cells in vaccine‐mediated protection from infectious diseases

“…31,32 Similarly, in pigs, cd T cells proliferated following vaccination with BCG. [36][37][38] cd T cells also expand following salmonella vaccination in chickens and macaques 39,40 or following salmonella infection in humans. Similarly to the described response to Mtb, human cd T-cell populations, in particular the Vc9Vd2 subset, expand following leptospirosis infection.…”

“…Similarly to the described response to Mtb, human cd T-cell populations, in particular the Vc9Vd2 subset, expand following leptospirosis infection. 40,42 Following subclinical Listeria monocytogenes infection, Vc9Vd2 T cells expanded, trafficked to the lungs and intestinal mucosa and evolved into effector cells producing IFNc, TNFa, Il-4, Il-17 and/or perforin. [36][37][38] cd T cells also expand following salmonella vaccination in chickens and macaques 39,40 or following salmonella infection in humans.…”

“…34,35 In leptospirosis vaccination studies in cattle, IFNc-producing cd T cells expressing the WC1 co-receptor expand postvaccination and upon in vitro restimulation. [36][37][38] cd T cells also expand following salmonella vaccination in chickens and macaques 39,40 or following salmonella infection in humans. 41 Furthermore, following salmonella or listeria vaccination in macaques, cd T cells displaying Vc9Vd2 were the major T-cell subset proliferating.…”

Resistance-resistant antibiotics

“…Thus, an IspH inhibitor will kill bacteria in a direct fashion, in addition to boosting innate immunity via γδ T cell expansion. As one proof-of-concept, a Salmonella ispH − mutant (complemented with genes from the mevalonate pathway) stimulated a profound expansion of Vγ2Vδ2 T cells in monkeys without side-effects or anergy induction[84]. IspH inhibitors should have the same effect, in addition to inhibiting bacterial cell wall biosynthesis.…”

Harnessing the antibacterial and immunological properties of mucosal-associated invariant T cells in the development of novel oral vaccines against enteric infections

“…One other exciting prospect is that bacteria could be engineered to over produce a MAIT cell agonist such as rRL-6-CH 2 OH, thus allowing for live attenuated bacterial vaccine strains to produce their own adjuvant. This proof of concept has been realized in Salmonella vaccine strains engineered to over produce the gd T cell activator HMB-PP [81], and could offer an alternative solution to the chemical synthesis method required for rRL-6-CH 2 OH production.…”