State-of-the-art in engineering small molecule biosensors and their applications in metabolic engineering.

Genetically encoded biosensors are crucial for enhancing our understanding of how molecules regulate biological systems. Small molecule biosensors, in particular, help us understand the interaction between chemicals and biological processes. They also accelerate metabolic engineering by increasing screening throughput and eliminating the need for sample preparation through traditional chemical analysis. Additionally, they offer significantly higher spatial and temporal resolution in cellular analyte measurements. In this review, we discuss recent progress in in vivo biosensors and control systems-biosensor-based controllers-for metabolic engineering. We also specifically explore protein-based biosensors that utilize less commonly exploited signaling mechanisms, such as protein stability and induced degradation, compared to more prevalent transcription factor and allosteric regulation mechanism. We propose that these lesser-used mechanisms will be significant for engineering eukaryotic systems and slower-growing prokaryotic systems where protein turnover may facilitate more rapid and reliable measurement and regulation of the current cellular state. Lastly, we emphasize the utilization of cutting-edge and state-of-the-art techniques in the development of protein-based biosensors, achieved through rational design, directed evolution, and collaborative approaches.

Nitric oxide-mediated S-nitrosylation of IAA17 protein in intrinsically disordered region represses auxin signaling.

The phytohormone auxin plays crucial roles in nearly every aspect of plant growth and development. Auxin signaling is activated through the phytohormone-induced proteasomal degradation of the Auxin/INDOLE-3-ACETIC ACID (Aux/IAA) family of transcriptional repressors. Notably, many auxin-modulated physiological processes are also regulated by nitric oxide (NO) that executes its biological effects predominantly through protein S-nitrosylation at specific cysteine residues. However, little is known about the molecular mechanisms in regulating the interactive NO and auxin networks. Here, we show that NO represses auxin signaling by inhibiting IAA17 protein degradation. NO induces the S-nitrosylation of Cys-70 located in the intrinsically disordered region of IAA17, which inhibits the TIR1-IAA17 interaction and consequently the proteasomal degradation of IAA17. The accumulation of a higher level of IAA17 attenuates auxin response. Moreover, an IAA17C70W nitrosomimetic mutation renders the accumulation of a higher level of the mutated protein, thereby causing partial resistance to auxin and defective lateral root development. Taken together, these results suggest that S-nitrosylation of IAA17 at Cys-70 inhibits its interaction with TIR1, thereby negatively regulating auxin signaling. This study provides unique molecular insights into the redox-based auxin signaling in regulating plant growth and development.