Stopping the swarm
Neutrophils play a major role in the early immune response and are recruited in large numbers into inflamed and infected tissues. By secreting chemoattractants that bind G protein–coupled receptors (GPCRs) on neighboring cells, neutrophils coordinate their behavior as a swarm. Less clear is how this auto-amplifying swarming activity is ultimately turned off. Kienle et al. show that desensitization of these GPCRs by the same chemoattractants by GPCR-kinase 2 (GRK2) is one way in which these swarms are shut down (see the Perspective by Rocha-Gregg and Huttenlocher). Unexpectedly, mice with GRK2-deficient neutrophils showed impaired rather than enhanced bacterial clearance. The heightened scanning ability of GRK2-deficient neutrophils may come at the cost of suboptimal phagocytosis and containment of bacteria.
Science, abe7729, this issue p. eabe7729; see also abj3065, p. 1262
Structured Abstract
INTRODUCTION
The collective behavior of cells and insects often relies on self-organizing processes. By releasing attractant signals, a few individuals can initiate the accumulation and aggregation of a whole population. Neutrophils, key players in the innate immune response, infiltrate inflamed and infected tissues in large numbers. These cells make use of such positive feedback amplification to find and kill bacteria in tissues. By secreting attractants that act through cell surface–expressed G protein–coupled receptors (GPCRs) on neighboring cells, neutrophils use this form of intercellular communication and coordinate their hunt for pathogens as a swarm. How this swarming response is terminated to avoid uncontrolled neutrophil accumulations and prevent excessive inflammation is currently unknown.
RATIONALE
The stop signals for neutrophil swarming in mammalian tissues have not yet been defined. They may be derived from cells of the surrounding inflammatory environment or from neutrophils themselves. We reasoned that the attractants released by neutrophils may become highly concentrated at sites where these cells cluster in larger numbers. It is well established that high chemoattractant concentrations can attenuate cellular responses by a process termed GPCR desensitization. We hypothesized a self-limiting mechanism for swarming: The local accumulation of the same neutrophil-expressed attractants that amplify swarming during early stages would cause desensitization of their respective GPCRs at later stages of neutrophil clustering. This led us to investigate the role of GPCR desensitization in neutrophil tissue navigation and host defense.
RESULTS
We generated mouse strains whose neutrophils were deficient in GPCR kinases (GRKs), critical enzymes for mediating the GPCR desensitization process. Of the four GRK isoforms tested, in vitro experiments identified GRK2 as the kinase necessary to desensitize GPCRs activated by swarm-released attractants (LTB4 and CXCL2). When neutrophils sense high concentrations of swarm attractants in vitro, GRK2 desensitizes the corresponding receptors to induce migration arrest. Two-photon intravital imaging of injured skin and infected lymph nodes of mice showed that GRK2 and GPCR desensitization play critical roles during neutrophil swarming in physiological tissue. At sites where swarming neutrophils accumulate and self-generate local fields of high swarm attractant concentration, GPCR desensitization was crucial to stop neutrophil migration arrest. Desensitization-resistant neutrophils moved faster and explored larger areas of lymph node tissue infected with the bacterium Pseudomonas aeruginosa. Such behavior suggested more effective bacterial sampling throughout the infected organ. Surprisingly, mice with GRK2-deficient neutrophils showed impaired rather than improved bacterial clearance. This finding could not be explained by altered antibacterial effector functions. In vitro assays for the detailed analysis of swarming behavior and bacterial growth revealed that GPCR desensitization to swarm attractants is required to induce neutrophil arrest for optimal bacterial phagocytosis and containment in swarm clusters.
CONCLUSION
We describe a cell-intrinsic stop mechanism for the self-organization of neutrophil collectives in infected tissues, which is based on sensing the local accumulation of the same cell-secreted attractants that amplify swarming during early stages. GPCR desensitization acts as a negative feedback control mechanism to stop neutrophil migration in swarm aggregates. This navigation mechanism allows neutrophils to self-limit their dynamics within forming swarms and ensures optimal elimination of bacteria. Desensitization to a self-produced activation signal as a principle of self-organization is important for immune host defense against bacteria, and likely informs other categories of collective behavior in cells and insects.
Top: Swarming neutrophils self-amplify their highly chemotactic recruitment toward sites of tissue injury or bacterial invasion by releasing attractants that act on neighboring neutrophils. Neutrophils are displayed as spheres with migration tracks (right). Bottom: The local accumulation of the same cell-secreted attractants stops neutrophils when they accumulate and form clusters, a process important for the containment of bacteria in infected tissues.
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Top: Swarming neutrophils self-amplify their highly chemotactic recruitment toward sites of tissue injury or bacterial invasion by releasing attractants that act on neighboring neutrophils. Neutrophils are displayed as spheres with migration tracks (right). Bottom: The local accumulation of the same cell-secreted attractants stops neutrophils when they accumulate and form clusters, a process important for the containment of bacteria in infected tissues.
Abstract
Neutrophils communicate with each other to form swarms in infected organs. Coordination of this population response is critical for the elimination of bacteria and fungi. Using transgenic mice, we found that neutrophils have evolved an intrinsic mechanism to self-limit swarming and avoid uncontrolled aggregation during inflammation. G protein–coupled receptor (GPCR) desensitization acts as a negative feedback control to stop migration of neutrophils when they sense high concentrations of self-secreted attractants that initially amplify swarming. Interference with this process allows neutrophils to scan larger tissue areas for microbes. Unexpectedly, this does not benefit bacterial clearance as containment of proliferating bacteria by neutrophil clusters becomes impeded. Our data reveal how autosignaling stops self-organized swarming behavior and how the finely tuned balance of neutrophil chemotaxis and arrest counteracts bacterial escape.