Construction of a Bile-responsive Expression System in Lactobacillus plantarum

Chae, Jong Pyo; Pajarillo, Edward Alain B.; Hwang, In-Chan; Kang, Dae‐Kyung

doi:10.5851/kosfa.2018.e58

Cited by 6 publications

(7 citation statements)

References 35 publications

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“…It was combined with pUC19 by cloning into the Sac I site; the erythromycin resistance gene (GCA_000219475.3) from L. johnsonii PF01, as a selection marker, was cloned into the Kpn I site. The bile inducible promoter of the lactate dehydrogenase 1 gene (GCA_000219475.3) [ 18 ] from L. johnsonii PF01 was PCR-amplified and Sal I and Pst I sites were cloned to express the target gene. The signal peptide and cell wall anchor of the surface layer protein A (SlpA) gene (Uniprot ID P35829) from L. acidophilus ATCC 4356 were amplified using PCR and located downstream of the target gene for cell wall anchoring of the target protein.…”

Section: Methodsmentioning

confidence: 99%

“…Our previous studies have demonstrated the potential of L. plantarum SK156 as an effective host for expressing bioactive substances in the intestine owing to its bile-responsive expression system [ 18 , 19 ]. In this study, the possible antigenic determinants of the spike, membrane, and envelope proteins of SARS-CoV-2 were predicted and displayed on the surface of recombinant L. plantarum strains.…”

Section: Introductionmentioning

confidence: 99%

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Mucosal immunization with Lactiplantibacillus plantarum-displaying recombinant SARS-CoV-2 epitopes on the surface induces humoral and mucosal immune responses in mice

et al. 2023

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Background The use of probiotic lactic acid bacteria as a mucosal vaccine vector is considered a promising alternative compared to the use of other microorganisms because of its “Generally Regarded as Safe” status, its potential adjuvant properties, and its tolerogenicity to the host. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease (COVID-19), is highly transmissible and pathogenic. This study aimed to determine the potential of Lactiplantibacillus plantarum expressing SARS-CoV-2 epitopes as a mucosal vaccine against SARS-CoV-2. Results In this study, the possible antigenic determinants of the spike (S1–1, S1–2, S1–3, and S1–4), membrane (ME1 and ME2), and envelope (E) proteins of SARS-CoV-2 were predicted, and recombinant L. plantarum strains surface-displaying these epitopes were constructed. Subsequently, the immune responses induced by these recombinant strains were compared in vitro and in vivo. Most surface-displayed epitopes induced pro-inflammatory cytokines [tumor necrosis factor alpha (TNF-α and interleukin (IL)-6] and anti-inflammatory cytokines (IL-10) in lipopolysaccharide-induced RAW 264.7, with the highest anti-inflammatory to pro-inflammatory cytokine ratio in the S1–1 and S1–2 groups, followed by that in the S1–3 group. When orally administered of recombinant L. plantarum expressing SARS-CoV-2 epitopes in mice, all epitopes most increased the expression of IL-4, along with induced levels of TNF-α, interferon-gamma, and IL-10, specifically in spike protein groups. Thus, the surface expression of epitopes from the spike S1 protein in L. plantarum showed potential immunoregulatory effects, suggesting its ability to potentially circumvent hyperinflammatory states relevant to monocyte/macrophage cell activation. At 35 days post immunization (dpi), serum IgG levels showed a marked increase in the S1–1, S1–2, and S1–3 groups. Fecal IgA levels increased significantly from 21 dpi in all the antigen groups, but the boosting effect after 35 dpi was explicitly observed in the S1–1, S1–2, and S1–3 groups. Thus, the oral administration of SARS-CoV-2 antigens into mice induced significant humoral and mucosal immune responses. Conclusion This study suggests that L. plantarum is a potential vector that can effectively deliver SARS-CoV-2 epitopes to intestinal mucosal sites and could serve as a novel approach for SARS-CoV-2 mucosal vaccine development.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Mucosal immunization with Lactiplantibacillus plantarum-displaying recombinant SARS-CoV-2 epitopes on the surface induces humoral and mucosal immune responses in mice

et al. 2023

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“…A bile-responsive E. coli-Lactobacillus shuttle expression vector, pULP3-P LDH , for Lactobacillus spp. was generated by fusing pUC19 with the L. plantarum plasmid pLP27, which has been found to persist in L. plantarum for up to 80 generations without selection pressure [90].…”

Section: Vectors Properties Reference Pnz9573mentioning

confidence: 99%

Plasmid-Based Gene Expression Systems for Lactic Acid Bacteria: A Review

et al. 2022

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Lactic acid bacteria (LAB) play a very vital role in food production, preservation, and as probiotic agents. Some of these species can colonize and survive longer in the gastrointestinal tract (GIT), where their presence is crucially helpful to promote human health. LAB has also been used as a safe and efficient incubator to produce proteins of interest. With the advent of genetic engineering, recombinant LAB have been effectively employed as vectors for delivering therapeutic molecules to mucosal tissues of the oral, nasal, and vaginal tracks and for shuttling therapeutics for diabetes, cancer, viral infections, and several gastrointestinal infections. The most important tool needed to develop genetically engineered LABs to produce proteins of interest is a plasmid-based gene expression system. To date, a handful of constitutive and inducible vectors for LAB have been developed, but their limited availability, host specificity, instability, and low carrying capacity have narrowed their spectrum of applications. The current review discusses the plasmid-based vectors that have been developed so far for LAB; their functionality, potency, and constraints; and further highlights the need for a new, more stable, and effective gene expression platform for LAB.

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“…A careful design must be taken into consideration, where the selection markers for constructing vectors should not rely on antibiotic resistance selection for successful cloning, but on other selective marker genes, such as auxotrophic mutant complementation or use of the CRISPR/Cas system [ 105 ]. Once achieved, lactobacilli show promising potential not only in health promotion, immunomodulation, and tolerance [ 104 , 106 ], but also in its observed genetic stability after genetic manipulation [ 107 ].…”

Section: Bacterial Mucosal Vaccines In the Literaturementioning

confidence: 99%

Hacking Commensal Bacteria to Consolidate the Adaptive Mucosal Immune Response in the Gut–Lung Axis: Future Possibilities for SARS-CoV-2 Protection

Pereira

Kang

et al. 2022

BioTech

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Infectious diseases caused by mucosal pathogens significantly increase mortality and morbidity. Thus, the possibility to target these pathogens at their primary entry points can consolidate protective immunity. Regarding SARS-CoV-2 infection, it has been observed that the upper respiratory mucosa is highly affected and that dysregulation of resident microbiota in the gut–lung axis plays a crucial role in determining symptom severity. Thus, understanding the possibility of eliciting various mucosal and adaptive immune responses allows us to effectively design bacterial mucosal vaccine vectors. Such design requires rationally selecting resident bacterial candidates as potential host carriers, evaluating effective carrier proteins for stimulating an immune response, and combining these two to improve antigenic display and immunogenicity. This review investigated mucosal vaccine vectors from 2015 to present, where a few have started to utilize Salmonella and lactic acid bacteria (LAB) to display SARS-CoV-2 Spike S proteins or fragments. Although current literature is still lacking for its studies beyond in vitro or in vivo efficiency, decades of research into these vectors show promising results. Here, we discuss the mucosal immune systems focusing on the gut–lung axis microbiome and offer new insight into the potential use of alpha streptococci in the upper respiratory tract as a vaccine carrier.

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Construction of a Bile-responsive Expression System in Lactobacillus plantarum

Cited by 6 publications

References 35 publications

Mucosal immunization with Lactiplantibacillus plantarum-displaying recombinant SARS-CoV-2 epitopes on the surface induces humoral and mucosal immune responses in mice

Mucosal immunization with Lactiplantibacillus plantarum-displaying recombinant SARS-CoV-2 epitopes on the surface induces humoral and mucosal immune responses in mice

Plasmid-Based Gene Expression Systems for Lactic Acid Bacteria: A Review

Hacking Commensal Bacteria to Consolidate the Adaptive Mucosal Immune Response in the Gut–Lung Axis: Future Possibilities for SARS-CoV-2 Protection

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