Biology subjects

Johannsen, M.

Publications and source records attributed to Johannsen, M..

3 recordsLinked to original sources

bioRxiv2026

Heterologous Reconstitution of 4-Amidopentadienoate Biosynthesis Identifies the PKS Machinery Responsible for Warhead Formation

The 4-amidopentadienoate (APD) motif is a rare and bioactive pharmacophore found in the cyclic lipodepsipeptide natural products BE-43547, rakicidins, microtermolides, and related compounds. Although several biosynthetic gene clusters associated with this family have been identified, the enzymatic origin of the APD warhead has remained unknown. Here, we investigated APD formation using NRPS engineering to install the unusual APD-associated polyketide synthase (PKS) module from the BE-43547 biosynthetic pathway into heterologous nonribosomal peptide synthetase/polyketide synthase (NRPS/PKS) assembly lines expressed in Escherichia coli. The engineered hybrids produced the designed lipopeptide containing the APD moiety, demonstrating that the core biosynthetic machinery is sufficient for APD installation and that no trans-acting enzymes are required. Total synthesis of the target compound enabled structural validation by co-elution, MS/MS analysis, and NMR spectroscopy. Production titers of 59 ug/L were achieved in the heterologous host. Systematic mutagenesis further revealed that the conserved dehydratase His-Asp catalytic dyad is essential for APD formation, whereas an additional highly conserved histidine is dispensable. These findings establish the biosynthetic origin of the APD warhead and provide a foundation for incorporating this privileged pharmacophore into engineered peptide scaffolds.

biochemistry↗

bioRxiv2026

Glutamate transporter xCT is important for cGAS-dependent interferon responses to DNA and to HSV-1

Metabolic reprogramming is a key component of antiviral immunity, yet how metabolite transport regulates innate immune signaling remains incompletely understood. Here, we show that infection with herpes simplex virus 1 (HSV-1) and stimulation with cytosolic DNA induce the cellular export of glutamate via the xCT (SLC7A11) transporter and that inhibition of xCT reduces cellular resistance to viral replication. Mechanistically, xCT inhibition impaired cGAS-STING signaling by reducing DNA-induced cGAMP production, thereby diminishing type I interferon (IFN/{beta}) responses and downstream induction of interferon-stimulated genes. Interestingly, modulating intracellular glutamate levels through inhibition of other glutamate pathways, e.g., glutaminolysis or glutamate import, also affected cellular IFN responses, suggesting that glutamate is a central control knob for DNA sensing. Finally, we demonstrate that HSV-1 suppresses xCT expression via a mechanism dependent on the immediate early viral protein ICP27, thereby promoting viral replication by limiting cGAS-dependent IFN induction. Together, these findings identify xCT-dependent glutamate transport as a critical metabolic regulator of cGAS-STING-mediated antiviral immunity.

immunology↗

bioRxiv2023

T-cell derived extracellular vesicles prime macrophages for improved STING based cancer immunotherapy

A key phenomenon in cancer is the establishment of a highly immunosuppressive tumor microenvironment (TME). Despite advances in immunotherapy, where the purpose is to induce tumor recognition and hence hereof tumor eradication, the majority of patients applicable for such treatment still fail to respond. It has been suggested that high immunological activity in the tumor is essential for achieving effective response to immunotherapy, which therefore have led to exploration of strategies that triggers inflammatory pathways. Here activation of the stimulator of interferon genes (STING) signaling pathway has been considered an attractive target, as it is a potent trigger of pro-inflammatory cytokines and type I and III interferons. However, immunotherapy combined with targeted STING agonists has not yielded sustained clinical remission in humans. This suggest a need for exploring novel adjuvants to improve the innate immunological efficacy. Here, we demonstrate that extracellular vesicles (EVs), derived from activated CD4+ T cells (T-EVs), sensitizes macrophages to elevate STING activation, mediated by IFN{gamma} carried on the T-EVs. Our work support that T-EVs can disrupt the immune suppressive environment in the tumor by reprogramming macrophages to a pro-inflammatory phenotype, and priming them for a robust immune response towards STING activation.

immunology↗