Brianna Dalesandro

Infect Immun. 2022 Mar 8:e0005622. doi: 10.1128/iai.00056-22. Online ahead of print.

ABSTRACT

Infections caused by the Gram-positive bacterium Staphylococcus aureus remain a significant health threat globally. The production of bicomponent pore-forming leukocidins plays an important role in S. aureus pathogenesis. Transcriptionally, these toxins are primarily regulated by the Sae and Agr regulatory systems. However, the posttranslational regulation of these toxins is largely unexplored. In particular, one of the leukocidins, LukAB, has been shown to be both secreted into the extracellular milieu and associated with the bacterial cell envelope. Here, we report that a major cell wall hydrolase, autolysin (Atl), controls the sorting of LukAB from the cell envelope to the extracellular milieu, an effect independent of transcriptional regulation. By influencing the sorting of LukAB, Atl modulates S. aureus cytotoxicity toward primary human neutrophils. Mechanistically, we found that the reduction in peptidoglycan cleavage and increased LukAB secretion in the atl mutant can be reversed through the supplementation of exogenous mutanolysin. Altogether, our study revealed that the cell wall hydrolase activity of Atl and the cleavage of peptidoglycan play an important role in controlling the sorting of S. aureus toxins during secretion.

PMID:35258336 | DOI:10.1128/iai.00056-22

Diseases. 2022 Jan 31;10(1):9. doi: 10.3390/diseases10010009.

ABSTRACT

D-amino acids may play key roles for specific physiological functions in different organs including the brain. Importantly, D-amino acids have been detected in several neurological disorders such as schizophrenia, amyotrophic lateral sclerosis, and age-related disorders, reflecting the disease conditions. Relationships between D-amino acids and neurophysiology may involve the significant contribution of D-Serine or D-Aspartate to the synaptic function, including neurotransmission and synaptic plasticity. Gut-microbiota could play important roles in the brain-function, since bacteria in the gut provide a significant contribution to the host pool of D-amino acids. In addition, the alteration of the composition of the gut microbiota might lead to schizophrenia. Furthermore, D-amino acids are known as a physiologically active substance, constituting useful biomarkers of several brain disorders including schizophrenia. In this review, we wish to provide an outline of the roles of D-amino acids in brain health and neuropsychiatric disorders with a focus on schizophrenia, which may shed light on some of the superior diagnoses and/or treatments of schizophrenia.

PMID:35225861 | PMC:PMC8883943 | DOI:10.3390/diseases10010009

Biomolecules. 2022 Feb 12;12(2):300. doi: 10.3390/biom12020300.

ABSTRACT

Ubiquitylation and ISGylation are protein post-translational modifications (PTMs) and two of the main events involved in the activation of pattern recognition receptor (PRRs) signals allowing the host defense response to viruses. As with similar viruses, SARS-CoV-2, the virus causing COVID-19, hijacks these pathways by removing ubiquitin and/or ISG15 from proteins using a protease called PLpro, but also by interacting with enzymes involved in ubiquitin/ISG15 machinery. These enable viral replication and avoidance of the host immune system. In this review, we highlight potential points of therapeutic intervention in ubiquitin/ISG15 pathways involved in key host-pathogen interactions, such as PLpro, USP18, TRIM25, CYLD, A20, and others that could be targeted for the treatment of COVID-19, and which may prove effective in combatting current and future vaccine-resistant variants of the disease.

PMID:35204803 | DOI:10.3390/biom12020300

ACS Chem Biol. 2022 Feb 25. doi: 10.1021/acschembio.1c00808. Online ahead of print.

ABSTRACT

Senescent cells undergo a permanent cell cycle arrest and drive a host of age-related pathologies. Recent transgenic mouse models indicate that removing cells expressing the senescence marker p16^Ink4a (p16) can increase median lifespan and delay the onset of many aging phenotypes. However, identifying and eliminating native human cells expressing p16 has remained a challenge. We hypothesize that senescent cells display peptides derived from p16 in major histocompatibility complex (MHC)-peptide complexes on the cell surface that could serve as targetable antigens for antibody-based biologics. Using Fab-phage display technology, we generated antibodies that bind to a p16 MHC-peptide complex from the human leukocyte antigen (HLA) allele HLA-B*35:01. When converted to single-chain Fab chimeric antigen receptor (CAR) constructs, these antibodies can recognize naturally presented p16 MHC-peptide complexes on the surface of cells and activate Jurkat cells. Furthermore, we developed antibodies against predicted p16 MHC-peptide complexes for HLA-A*02:01 that specifically recognize their respective antigen on the surface of cells. These tools establish a platform to survey the surface of senescent cells and provide a potential novel senolytic strategy.

PMID:35212540 | DOI:10.1021/acschembio.1c00808

J Mol Biol. 2022 Feb 23:167513. doi: 10.1016/j.jmb.2022.167513. Online ahead of print.

ABSTRACT

We have previously developed a universal chimeric antigen receptor (CAR), which recognizes dinitrophenyl (DNP) and can redirect T and NK cells to target cancer and HIV antigens using DNP-conjugated antibodies as adaptor molecules. However, the DNP-antibody conjugates are generated by random modification, which may not be optimal for this modular system. Here, we report the development of enhanced adaptor molecules by site-specific DNP modification. We use the genetic code expansion technology to generate single-chain fragment variable (scFv) antibodies with site-specific DNP. We compare four anti-CD19 scFv mutants and find that the one with DNP at the flexible peptide linker between V_H and V_L is the most effective in redirecting anti-DNP CAR-T cells against CD19+ cells. The other three mutants are ineffective in doing so due to reduced DNP exposure or abrogated CD19 binding. We also use the anti-CD22 scFv as another model adaptor molecule and again find that the peptide linker is ideal for DNP derivatization. Our approach can potentially be used to design enhanced adaptor molecules to redirect the DNP-mediated universal CAR against other tumor antigens.

PMID:35218770 | DOI:10.1016/j.jmb.2022.167513

Microorganisms. 2022 Jan 24;10(2):264. doi: 10.3390/microorganisms10020264.

ABSTRACT

Pseudomonas aeruginosa is a common, opportunistic bacterial pathogen among patients with cystic fibrosis, asthma, and chronic obstructive pulmonary disease. During the course of these diseases, l-ornithine, a non-proteinogenic amino acid, becomes more abundant. P. aeruginosa is chemotactic towards other proteinogenic amino acids. Here, we evaluated the chemotaxis response of P. aeruginosa towards l-ornithine. Our results show that l-ornithine serves as a chemoattractant for several strains of P. aeruginosa, including clinical isolates, and that the chemoreceptors involved in P. aeruginosa PAO1 are PctA and PctB. It seems likely that P. aeruginosa's chemotactic response to l-ornithine might be a common feature and thus could potentially contribute to pathogenesis processes during colonization and infection scenarios.

PMID:35208720 | DOI:10.3390/microorganisms10020264

RSC Med Chem. 2021 Nov 9;13(1):72-78. doi: 10.1039/d1md00262g. eCollection 2022 Jan 27.

ABSTRACT

The recruitment of antibody naturally present in human blood stream at the surface of cancer cells have been proved a promising immunotherapeutic strategy to fight cancer. Antibody recruiting molecules (ARMs) combining tumor and antibody binding modules have been developed for this purpose, however the formation of the interacting complex with both antibody and cell is difficult to optimize to stimulate immune-mediated cytotoxicity. To circumvent this limitation, we report herein a more direct approach combining cell metabolism of azido-sugar and bio-orthogonal click chemistry to conjugate at the cell glycocalyx structurally well-defined glycodendrimers as antibody binding module (ABM). We demonstrate that this strategy allows not only the recruitment of natural antibody at the surface of isolated cells or solid tumor models but also activate a cytotoxic response with human serum as unique source of immune effectors.

PMID:35211675 | PMC:PMC8792828 | DOI:10.1039/d1md00262g

Front Microbiol. 2022 Feb 2;12:708580. doi: 10.3389/fmicb.2021.708580. eCollection 2021.

ABSTRACT

Bacterial persisters are a sub-population of phenotypic variants that tolerate high concentrations of antibiotics within the genetically homogeneous cells. They resume division upon the removal of drugs. Bacterial persistence is one of major causes of antibiotic treatment failure and recurrent infection. Cell dormancy, triggered by toxin/antitoxin pair, (p)ppGpp, SOS response and ATP levels, is known to be the mechanistic basis for persistence. However, recent studies have demonstrated that bacteria with active metabolism can maintain persistence by lowering intracellular antibiotic concentration via an efflux pump. Additionally, others and our work have showed that cell wall deficient bacteria (CWDB), including both L-form and spheroplasts that produced by β-lactam antibiotics, are associated with antibiotic persistence. They are not dormant cells as their cell walls have been completely damaged. In this review, we discuss the various types of persisters and highlight the contribution of non-walled bacteria on bacterial persistence.

PMID:35185807 | PMC:PMC8847742 | DOI:10.3389/fmicb.2021.708580

ACS Chem Biol. 2022 Nov 18;17(11):3036-3046. doi: 10.1021/acschembio.1c00855. Epub 2022 Feb 16.

NO ABSTRACT

PMID:35170962 | DOI:10.1021/acschembio.1c00855

Sci Rep. 2022 Feb 8;12(1):2097. doi: 10.1038/s41598-022-06109-7.

ABSTRACT

The disaccharide trehalose is essential for viability of Mycobacterium tuberculosis, which synthesizes trehalose de novo but can also utilize exogenous trehalose. The mycobacterial cell wall encompasses two permeability barriers, the cytoplasmic membrane and the outer mycolic acid-containing mycomembrane. The ABC transporter LpqY-SugA-SugB-SugC has previously been demonstrated to mediate the specific uptake of trehalose across the cytoplasmic membrane. However, it is still unclear how the transport of trehalose molecules across the mycomembrane is mediated. In this study, we harnessed the antimycobacterial activity of the analogue 6-azido trehalose to select for spontaneous resistant M. tuberculosis mutants in a merodiploid strain harbouring two LpqY-SugA-SugB-SugC copies. Mutations mediating resistance to 6-azido trehalose mapped to the proline-proline-glutamate (PPE) family member PPE51 (Rv3136), which has recently been shown to be an integral mycomembrane protein involved in uptake of low-molecular weight compounds. A site-specific ppe51 gene deletion mutant of M. tuberculosis was unable to grow on trehalose as the sole carbon source. Furthermore, bioorthogonal labelling of the M. tuberculosis Δppe51 mutant incubated with 6-azido trehalose corroborated the impaired internalization. Taken together, the results indicate that the transport of trehalose and trehalose analogues across the mycomembrane of M. tuberculosis is exclusively mediated by PPE51.

PMID:35136132 | PMC:PMC8826857 | DOI:10.1038/s41598-022-06109-7

PLoS Pathog. 2019 Feb 11;15(2):e1007571. doi: 10.1371/journal.ppat.1007571. eCollection 2019 Feb.

ABSTRACT

Bacterial pathogens have evolved strategies that enable them to invade tissues and spread within the host. Enterococcus faecalis is a leading cause of local and disseminated multidrug-resistant hospital infections, but the molecular mechanisms used by this non-motile bacterium to penetrate surfaces and translocate through tissues remain largely unexplored. Here we present experimental evidence indicating that E. faecalis generates exopolysaccharides containing β-1,6-linked poly-N-acetylglucosamine (polyGlcNAc) as a mechanism to successfully penetrate semisolid surfaces and translocate through human epithelial cell monolayers. Genetic screening and molecular analyses of mutant strains identified glnA, rpiA and epaX as genes critically required for optimal E. faecalis penetration and translocation. Mechanistically, GlnA and RpiA cooperated to generate uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) that was utilized by EpaX to synthesize polyGlcNAc-containing polymers. Notably, exogenous supplementation with polymeric N-acetylglucosamine (PNAG) restored surface penetration by E. faecalis mutants devoid of EpaX. Our study uncovers an unexpected mechanism whereby the RpiA-GlnA-EpaX metabolic axis enables production of polyGlcNAc-containing polysaccharides that endow E. faecalis with the ability to penetrate surfaces. Hence, targeting carbohydrate metabolism or inhibiting biosynthesis of polyGlcNAc-containing exopolymers may represent a new strategy to more effectively confront enterococcal infections in the clinic.

PMID:30742693 | PMC:PMC6386517 | DOI:10.1371/journal.ppat.1007571

Front Immunol. 2022 Jan 19;12:811381. doi: 10.3389/fimmu.2021.811381. eCollection 2021.

ABSTRACT

Infection caused by antibiotic-resistant microorganisms (ARMs) has been declared a global threat to public health. Polymeric nanoparticles (PNPs) formed by antimicrobial peptides (AMPs) and synthetic PNPs against ARM infections are emerging. PNPs are also considered to be a promising natural biological preservative that prevents microbial spoilage through food processing and preservation. We engineered CNMs, a novel nanocomposite antibacterial agent based on chitosan nanoparticles and AMP microcin J25. In this study, we aimed to evaluate the comprehensive antimicrobial activity, potential antimicrobial mechanism, and anti-inflammatory activity of CNMs. We demonstrated that CNMs harbor excellent bactericidal activity against clinical foodborne pathogens and ARMs. CNMs caused fast mortality against different growth phases of tetracycline (Tet)-resistant enterotoxigenic E. coli (ETEC) and significantly killed Tet-resistant ETEC in food biological environments. Mechanistically, CNMs have the ability to bind lipopolysaccharides (LPS), neutralize endotoxin, and promote diaphragm permeability by damaging the cell membrane. CNMs did not cause mouse RAW264.7 cell cytotoxicity. Notably, CNMs significantly reduced the cytotoxicity of RAW264.7 macrophages induced by LPS. The LPS-induced inflammatory response was significantly ameliorated by CNMs by reducing the levels of nitric oxide and proinflammatory cytokines, including tumor necrosis factor α, interleukin (IL)-6, IL-8, IL-1β, Toll-like receptor 4, and nuclear factor κB (NF-κB), in LPS-challenged RAW264.7 macrophages. CNMs downregulated the NF-κB and mitogen-activated protein kinase signaling pathways, thereby inhibiting inflammatory responses upon LPS stimulation. Taken together, CNMs could be applied as effective antimicrobial/anti-inflammatory agents with lower cytotoxicity in food, medicine, and agriculture to prevent bacterial contamination and infection, respectively.

PMID:35126369 | PMC:PMC8807516 | DOI:10.3389/fimmu.2021.811381