Beyond natural biology: rewiring cellular networks to study innate immunity

Abstract

Within immune cells, microbial and self-ligands trigger pattern recognition receptors (PRRs) to nucleate and activate the signaling organelles of the immune system. Much work in this area has derived from observational biology of natural innate immune signaling. More recently, synthetic biology approaches have been used to rewire and study innate immune networks. By utilizing controllable chemical or optogenetic inputs, rearranging protein building blocks, or engineering signal recording circuits, synthetic biology-based techniques complement and inform studies of natural immune pathway operation. In this review, we describe recent synthetic biology-based approaches that have uncovered new insights into PRR signaling, virus-host interactions, and systemic cytokine responses.

Figure 1.. The complexity of natural signaling networks.

Many different plasma membrane and intracellular PRRs promote complex signaling networks. These networks contain activating and inhibitory signals that interact across different pathways to generate molecular crosstalk. Outcomes of PRR signaling pathways include gene expression, metabolic reprogramming, protein degradation, membrane trafficking, cytokine secretion, and cell rupture.

Figure 2.. Synthetic biology approaches rewire the inputs, processing, and outputs of innate immune cells.

Innate immune cells function as information processors that convert inputs, such as microbial products, into outputs, such as cytokines. Synthetic biology approaches reengineer each of these steps to disentangle complex networks and study components of innate immune signaling. (A) Chemical oligomerization of FKBP or light-induced oligomerization of CRY2 allows for controllable SMOC assembly and signaling. (B) The functional domains of different proteins can be rearranged to generated chimeric proteins with new capacities. For example, the pLxIS domain of STING can be appended to MyD88, the transmembrane domain of TMEM192 can replace the transmembrane domain of STING, or the human TIR domain of TLR2 can replace the TIR domain of invertebrate TLRs. (C) Innate immune outputs such as gene expression, ROS products, or Mg²⁺ signaling can be recorded through rewired memory networks. For example, signals that induce gene expression can be engineered to induce crRNAs that direct dCas12a-ABE8e fusion proteins to introduce permanent mutations in an endogenous locus. Additionally, ROS products can induce memory switches in bacteria to be permanently turned on. TM = transmembrane domain. ABE8e = evolved adenine base editor.