@phdthesis{11945,
  abstract     = {G protein-coupled receptors (GPCRs) respond to specific ligands and regulate multiple processes ranging from cell growth and immune responses to neuronal signal transmission. However, ligands for many GPCRs remain unknown, suffer from off-target effects or have poor bioavailability. Additional challenges exist to dissect cell-type specific responses when the same GPCR is expressed on several cell types within the body. Here, we overcome these limitations by engineering DREADD-based GPCR chimeras that selectively bind their agonist clozapine-N-oxide (CNO) and mimic a GPCR-of-interest in a desired cell type.
We validated our approach with β2-adrenergic receptor (β2AR/ADRB2) and show that our chimeric DREADD-β2AR triggers comparable responses on second messenger and kinase activity, post-translational modifications, and protein-protein interactions. Since β2AR is also enriched in microglia, which can drive inflammation in the central nervous system, we expressed chimeric DREADD-β2AR in primary microglia and successfully recapitulate β2AR-mediated filopodia formation through CNO stimulation. To dissect the role of selected GPCRs during microglial inflammation, we additionally generated DREADD-based chimeras for microglia-enriched GPR65 and GPR109A/HCAR2. In a microglia cell line, DREADD-β2AR and DREADD-GPR65 both modulated the inflammatory response with a similar profile as endogenously expressed β2AR, while DREADD-GPR109A showed no impact.
Our DREADD-based approach provides the means to obtain mechanistic and functional insights into GPCR signaling on a cell-type specific level.},
  author       = {Schulz, Rouven},
  issn         = {2663-337X},
  pages        = {133},
  publisher    = {Institute of Science and Technology Austria},
  title        = {{Chimeric G protein-coupled receptors mimic distinct signaling pathways and modulate microglia function}},
  doi          = {10.15479/at:ista:11945},
  year         = {2022},
}

@article{11995,
  abstract     = {G protein-coupled receptors (GPCRs) regulate processes ranging from immune responses to neuronal signaling. However, ligands for many GPCRs remain unknown, suffer from off-target effects or have poor bioavailability. Additionally, dissecting cell type-specific responses is challenging when the same GPCR is expressed on different cells within a tissue. Here, we overcome these limitations by engineering DREADD-based GPCR chimeras that bind clozapine-N-oxide and mimic a GPCR-of-interest. We show that chimeric DREADD-β2AR triggers responses comparable to β2AR on second messenger and kinase activity, post-translational modifications, and protein-protein interactions. Moreover, we successfully recapitulate β2AR-mediated filopodia formation in microglia, an immune cell capable of driving central nervous system inflammation. When dissecting microglial inflammation, we included two additional DREADD-based chimeras mimicking microglia-enriched GPR65 and GPR109A. DREADD-β2AR and DREADD-GPR65 modulate the inflammatory response with high similarity to endogenous β2AR, while DREADD-GPR109A shows no impact. Our DREADD-based approach allows investigation of cell type-dependent pathways without known endogenous ligands.},
  author       = {Schulz, Rouven and Korkut, Medina and Venturino, Alessandro and Colombo, Gloria and Siegert, Sandra},
  issn         = {2041-1723},
  journal      = {Nature Communications},
  publisher    = {Springer Nature},
  title        = {{Chimeric GPCRs mimic distinct signaling pathways and modulate microglia responses}},
  doi          = {10.1038/s41467-022-32390-1},
  volume       = {13},
  year         = {2022},
}

@article{6521,
  abstract     = {Microglia have emerged as a critical component of neurodegenerative diseases. Genetic manipulation of microglia can elucidate their functional impact in disease. In neuroscience, recombinant viruses such as lentiviruses and adeno-associated viruses (AAVs) have been successfully used to target various cell types in the brain, although effective transduction of microglia is rare. In this review, we provide a short background of lentiviruses and AAVs, and strategies for designing recombinant viral vectors. Then, we will summarize recent literature on successful microglial transductions in vitro and in vivo, and discuss the current challenges. Finally, we provide guidelines for reporting the efficiency and specificity of viral targeting in microglia, which will enable the microglial research community to assess and improve methodologies for future studies.},
  author       = {Maes, Margaret E and Colombo, Gloria and Schulz, Rouven and Siegert, Sandra},
  issn         = {0304-3940},
  journal      = {Neuroscience Letters},
  publisher    = {Elsevier},
  title        = {{Targeting microglia with lentivirus and AAV: Recent advances and remaining challenges}},
  doi          = {10.1016/j.neulet.2019.134310},
  volume       = {707},
  year         = {2019},
}