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1. The macroscopic limit to synchronization of cellular clocks in single cells of Neurospora crassa

   https://doi.org/10.1038/s41598-022-10612-2  

   Cheong, Jia Hwei; Qiu, Xiao; Liu, Yang; Al-Omari, Ahmad; Griffith, James; Schüttler, Heinz-Bernd; Mao, Leidong; Arnold, Jonathan (April 2022, Scientific Reports)

   Abstract We determined the macroscopic limit for phase synchronization of cellular clocks in an artificial tissue created by a “big chamber” microfluidic device to be about 150,000 cells or less. The dimensions of the microfluidic chamber allowed us to calculate an upper limit on the radius of a hypothesized quorum sensing signal molecule of 13.05 nm using a diffusion approximation for signal travel within the device. The use of a second microwell microfluidic device allowed the refinement of the macroscopic limit to a cell density of 2166 cells per fixed area of the device for phase synchronization. The measurement of averages over single cell trajectories in the microwell device supported a deterministic quorum sensing model identified by ensemble methods for clock phase synchronization. A strong inference framework was used to test the communication mechanism in phase synchronization of quorum sensing versus cell-to-cell contact, suggesting support for quorum sensing. Further evidence came from showing phase synchronization was density-dependent. 

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2. Quorum sensing in single cells of Neurospora crassa

   Qiu, Xiao; Cheong, Jia; Liu, Yang; Schuttler, Bernd; Mao, L; Arnold, Jonathan (October 2021, MicroTAS2021)

   The use of a microwell microfluidic device allows separating single cells and tracking single cells data. The measurement of single cell fluorescent intensity trajectories in the microwell device supported a deterministic quorum sensing model identified by ensemble methods for clock phase synchronization. A strong inference framework was used to test the communication mechanism in phase synchronization of quorum sensing versus cell-to-cell contact, and the results lent support for quorum sensing. 

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3. Orthogonal Surface Acoustic Wave (SAW) Sensor for Cancer Biomarker Detection with Accelerated Binding Kinetics

   https://doi.org/10.1109/SENSORS52175.2022.9967354  

   Huang, Yuqi; Boucher, Maelys; Evans-Nguyen, Theresa; Bhethanabotla, Venkat R. (October 2022, 2022 IEEE Sensors)

   A device incorporating both Rayleigh wave and shear horizontal surface acoustic waves is made on a ST-Quartz substrate. The Rayleigh wave induced microfluidic mixing shows effects on accelerating the binding kinetics of real-time sensing between antibody and antigen, which is measured by phase change from the shear horizontal surface acoustic wave direction on the ST-Quartz. Preliminary results on this device show shortened response time and enhanced phase signal when the binding is accelerated by microfluidic streaming from the Rayleigh wave. The device can be fabricated using a low cost, single step photolithography method and can be combined with a small electronic sensor for data readout, which allows for a variety of surface-based biomarker detections on a portable platform. In this work, detection of Carcinoembryonic antigen (CEA) binding with functionalized capture antibody is studied to show the effects of mass loading amplification due to Rayleigh wave microfluidic streaming. 

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4. Antiactivators prevent self-sensing in Pseudomonas aeruginosa quorum sensing

   https://doi.org/10.1073/pnas.2201242119  

   Smith, Parker; Schuster, Martin (June 2022, Proceedings of the National Academy of Sciences)

   Quorum sensing is described as a widespread cell density-dependent signaling mechanism in bacteria. Groups of cells coordinate gene expression by secreting and responding to diffusible signal molecules. Theory, however, predicts that individual cells may short-circuit this mechanism by directly responding to the signals they produce irrespective of cell density. In this study, we characterize this self-sensing effect in the acyl-homoserine lactone quorum sensing system of Pseudomonas aeruginosa . We show that antiactivators, a set of proteins known to affect signal sensitivity, function to prevent self-sensing. Measuring quorum-sensing gene expression in individual cells at very low densities, we find that successive deletion of antiactivator genes qteE and qslA produces a bimodal response pattern, in which increasing proportions of constitutively induced cells coexist with uninduced cells. Comparing responses of signal-proficient and -deficient cells in cocultures, we find that signal-proficient cells show a much higher response in the antiactivator mutant background but not in the wild-type background. Our results experimentally demonstrate the antiactivator-dependent transition from group- to self-sensing in the quorum-sensing circuitry of P. aeruginosa . Taken together, these findings extend our understanding of the functional capacity of quorum sensing. They highlight the functional significance of antiactivators in the maintenance of group-level signaling and experimentally prove long-standing theoretical predictions. 

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5. Structural determinants driving homoserine lactone ligand selection in the Pseudomonas aeruginosa LasR quorum-sensing receptor

   https://doi.org/10.1073/pnas.1817239116  

   McCready, Amelia R.; Paczkowski, Jon E.; Henke, Brad R.; Bassler, Bonnie L. (December 2018, Proceedings of the National Academy of Sciences)

   Quorum sensing is a cell–cell communication process that bacteria use to orchestrate group behaviors. Quorum sensing is mediated by signal molecules called autoinducers. Autoinducers are often structurally similar, raising questions concerning how bacteria distinguish among them. Here, we use thePseudomonas aeruginosaLasR quorum-sensing receptor to explore signal discrimination. The cognate autoinducer, 3OC₁₂homoserine lactone (3OC₁₂HSL), is a more potent activator of LasR than other homoserine lactones. However, other homoserine lactones can elicit LasR-dependent quorum-sensing responses, showing that LasR displays ligand promiscuity. We identify mutants that alter which homoserine lactones LasR detects. Substitution at residue S129 decreases the LasR response to 3OC₁₂HSL, while enhancing discrimination against noncognate autoinducers. Conversely, the LasR L130F mutation increases the potency of 3OC₁₂HSL and other homoserine lactones. We solve crystal structures of LasR ligand-binding domains complexed with noncognate autoinducers. Comparison with existing structures reveals that ligand selectivity/sensitivity is mediated by a flexible loop near the ligand-binding site. We show that LasR variants with modified ligand preferences exhibit altered quorum-sensing responses to autoinducers in vivo. We suggest that possessing some ligand promiscuity endows LasR with the ability to optimally regulate quorum-sensing traits. 

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