Review Bacterial Signaling to the Nervous System through Toxins and Metabolites Nicole J. Yang and Isaac M. Chiu Department of Microbiology and Immunobiology, Division of Immunology, Harvard Medical School, Boston, MA 02115, USA Correspondence to Isaac M. Chiu: [email protected] http://dx.doi.org/10.1016/j.jmb.2016.12.023 Edited by Jenal Urs Abstract Mammalian hosts interface intimately with commensal and pathogenic bacteria. It is increasingly clear that molecular interactions between the nervous system and microbes contribute to health and disease. Both commensal and pathogenic bacteria are capable of producing molecules that act on neurons and affect essential aspects of host physiology. Here we highlight several classes of physiologically important molecular interactions that occur between bacteria and the nervous system. First, clostridial neurotoxins block neurotransmission to or from neurons by targeting the SNARE complex, causing the characteristic paralyses of botulism and tetanus during bacterial infection. Second, peripheral sensory neurons—olfactory chemosensory neurons and nociceptor sensory neurons—detect bacterial toxins, formyl peptides, and lipopolysaccharides through distinct molecular mechanisms to elicit smell and pain. Bacteria also damage the central nervous system through toxins that target the brain during infection. Finally, the gut microbiota produces molecules that act on enteric neurons to influence gastrointestinal motility, and metabolites that stimulate the “gut–brain axis” to alter neural circuits, autonomic function, and higher-order brain function and behavior. Furthering the mechanistic and molecular understanding of how bacteria affect the nervous system may uncover potential strategies for modulating neural function and treating neurological diseases. © 2017 Elsevier Ltd. All rights reserved. Introduction metabolism, temperature control, mood, behavior, and cognition. In this review, we focus on two molecular Mammals host an incredibly complex community classes of microbial signals that regulate the nervous of commensal bacteria, with an estimated 10 trillion system: bacterial toxins and metabolites. organism residents in an adult human gut [1]. From the perspective of the host, the nervous Increasing evidence suggests that microbes residing system provides a rapidly responsive and robust in the gut, respiratory tract, genitourinary tract, and mechanism to detect bacterial cues and coordinate other barrier tissues actively participate in shaping and the appropriate defensive response. For example, maintaining our physiology during development and peripheral sensory neurons densely innervate host homeostasis—almost as an extra “organ” [2].In barrier tissues, and are thus well positioned to detect contrast, pathogenic bacteria have developed molec- unwanted microbes and microbial products when ular strategies to survive within hosts, damaging infection occurs [3,4]. Alternatively, in health, the physiological function and fitness through secreted gastrointestinal tract is densely inhabited by commen- toxins and metabolites. However, despite these sal bacteria, which easily outnumber local host cells differences, commensal and pathogenic bacteria by orders of magnitude [1]. As the gut microbiota is share a common incentive to influence host physiology also an active producer of metabolites, its chemical for their benefit. In this aspect, the nervous system is a signature allows the host nervous system to sample desirable target as a master regulator of host function. the status of gut bacterial communities and health. By signaling to the nervous system, bacteria are An improved mechanistic understanding of how granted a handle to influence a broad range of complex bacterial molecules act on the nervous system could physiology, including motor coordination, sensation, yield improved therapeutics for treating neurological 0022-2836/© 2017 Elsevier Ltd. All rights reserved. J Mol Biol (2017) 429, 587–605 588 Bacterial toxins and metabolites modulate neurons diseases, as well as research tools for perturbing glutamate and γ-aminobutyric acid (GABA) stored and analyzing the nervous system. Botulinum within synaptic vesicles. Certain bacterial pathogens neurotoxin (BoNT) is a prime example where the have evolved toxins that block this vesicular release. toxin's ability to silence neurotransmission is cur- Depending on the type of neuron affected (e.g., motor rently utilized for the treatment of migraine and or sensory neuron), inhibiting neurotransmitter re- muscle spasticity [5]. The receptor-binding subunit of lease can cause diverging physiological conse- cholera toxin (CT) is widely used for retrograde quences, such as flaccid paralysis observed with tracing of neuronal connections as an experimental BoNT or spastic paralysis observed with tetanus tool [6]. Identifying novel molecular interactions neurotoxin (TeNT). Here we discuss the specific through which bacteria act on the nervous system, molecular mechanisms through which these bacterial and characterizing known interactions in greater toxins block neurotransmission (Fig. 1). detail could highlight additional molecular pathways to be targeted or utilized. In the case of infectious Botulinum neurotoxins disease, a better understanding of pathogenic mechanisms involving neuron–microbe interactions BoNTs and TeNT together compose the clostridial could also lead to novel antimicrobial approaches. family of neurotoxins. BoNTs are the causative toxin As such, here we highlight the molecular mecha- for botulism in humans, which initially presents with nisms through which commensal and pathogenic blurred vision, difficulty swallowing and speaking, bacteria signal to the host central and peripheral and muscle weakening, progressing to descending nervous systems. Here, we focus mainly on recent flaccid paralysis [8]. Among the known clostridial work showing direct microbe–neuron molecular inter- toxins, BoNTs are the most potent [9], and in the actions, where bacterial molecules bind specifically or wild, amounts below the lethal dose are sufficient to non-specifically to neurons to alter their biology and paralyze a host and effectively terminate its survival subsequent host physiology. We also introduce exam- [10]. Consequently, BoNTs play a central role in ples of indirect interactions where bacterial molecules facilitating bacterial spread as cadavers provide an act on an intermediary cell type such as endocrine or organic-rich, anaerobic environment that allows immune cells, which in turn produce neurochemicals toxigenic clostridial species to proliferate [10]. and immune mediators that affect neurons. First, we BoNTs are produced by multiple strains of clostridia, discuss neurotoxins that specifically inhibit the vesicu- including Clostridium botulinum, Clostridium butyricum, lar release of neurotransmitters. Next, we introduce and Clostridium baratii. BoNTs are a genetically examples of bacterial molecules that affect smell and diverse group that can be classified into seven pain through their action on sensory neurons. In serotypes (BoNT/A–BoNT/G) based on immunoreac- primitive organisms such as Caenorhabditis elegans, tivity. An additional serotype (BoNT/H) was proposed, pathogenic bacteria are detected by chemosensation, but recent studies suggest that it is a hybrid of BoNT which leads to avoidance behavior [7]. Recent work serotypes A and F [11]. shows that similar mechanisms are also present in BoNTs are synthesized as a single-chain precursor, mammals, whose vomeronasal olfactory neurons and which is cleaved by clostridial or host proteases to pain-sensing nociceptor neurons respond to bacterial yield a heavy chain (HC) and light chain (LC) that effectors such as pore-forming toxins, N-formyl pep- compose the mature toxin. The two chains are linked tides, and various pathogen-associated molecular by a disulfide bridge and held together by a “belt patterns (PAMPs). Bacteria also act on the intrinsic region,” a loop that extends from the HC and wraps neurons of the enteric nervous system (ENS) to around the LC [12–14]. In instances of food-borne or regulate gut motility through structural molecules such inhalation botulism where the toxin is first present in as polysaccharide A (PSA) or metabolites such as the lumen of the gut or respiratory tract, the HC short-chain fatty acids (SCFAs). Bacteria-induced mediates the transcytosis of the toxin across epithelial changes in gut motility could facilitate their colonization cells [15–17]. Subsequently, the intact holotoxin can or maintenance of gut microflora composition. Lastly, enter the general circulation and accumulate at nerve we explore the increasingly complex area of research terminals [18]. of the gut–brain axis, where neurotransmitters, toxins, The selective binding of BoNTs to nerve terminals is and metabolites produced by the gut microbiota possible by their recognition of two independent host affect neural circuits, behavior, and central nervous receptors, which synergistically increase apparent system (CNS) function, with implications in neurolog- affinity [19]. Initial BoNT binding is to membrane poly- ical disorders such as autism. sialogangliosides (PSGs), mediated by the C terminal domain of the HC [20–22]. Subsequent binding occurs through proteinaceous receptors, mediated by Bacterial blockade of neurotransmission a separate site within the C terminal domain of the HC. The synaptic vesicle protein SV2 serves as the Neurons transmit signals to each other at synapses