Team:UCL/Effectors

Effectors

Background

The term psychobiotics describes an emerging class of mind-altering probiotics that possess the ability to produce neuroactive compounds. The molecules associated with mammalian neurotransmission that have been isolated from gut bacteria include gamma-butyric acid (GABA), serotonin, dopamine, and acetylcholine Lyte, M. and Cryan, J. Microbial Endocrinology: The Microbiota-Gut-Brain Axis in Health and Disease Advances in Experimental Medicine and Biology Volume 817, 2014, pp 221-239 [Link][1]. When secreted into intestinal lumen, these molecules can modulate the neural signalling within the enteric nervous system which communicates with brain via the vagus nerve, therefore influencing brain’s biochemistry and host’s behaviour.

“It is well recognized that some bacteria within the human GIT have the capacity to produce many neurotransmitters and neuromodulators. For example, Lactoba cillus spp. and Bifidobacterium spp. have been reported to produce GABA; Escherichia spp., and Bacillus spp. have been reported to produce norepinephrine; Streptococcus spp., Escherichia spp. and Enterococcus spp. have been reported to produce serotonin; Bacillus spp. have been reported to produce dopamine, and Lactobacillus spp. have been reported to produce acetylcholine and histamine. [...] Other bacterially-produced metabolites with proven neuroactive func- tions include short chain fatty acids (SCFAs) and long chain fatty acids such as conjugated linoleic acid (CLA). “
Microbial Endocrinology: The Microbiota-Gut-Brain Axis in Health and Disease Advances in Experimental Medicine and Biology Volume 817, 2014, pp 221-239


Probiotics are live microorganisms that, when ingested in adequate amounts, exert a health benefit on the host Report on Joint FAO/WHO Expert Consultation on Evaluation of Health and Nutritional Properties of Probiotics in Food including Powder Milk with Live Lactic Acid Bacteria, 2001. [Link][2]. The studies on mice indicate that several natural probiotic strains exhibit psychobiotic properties: treatment with Lactobacillus rhamnosus has been shown to regulate emotional behaviour and influence the mRNA expression of GABA receptor in host’s brain Bravo, Javier A, Forsythe, Paul, Chew, Marianne V, Escaravage, Emily; Savignac, Hélène M et al. Ingestion of Lactobacillus strain regulates emotional behavior and central GABA receptor expression in a mouse via the vagus nerve. Proceedings of the National Academy of Sciences of the United States of America vol. 108 (38), 2011, p. 16050-5 [Link][3], while another study has shown that administration of Bifidobacterium infantis results in increased concentration of tryptophan, a serotonergic precursor, in blood plasma and shows antidepressant properties as determined by forced swim test Desbonnet, L., Garrett, L., Clarke, G., Bienenstock, J., & Dinan, T. G. The probiotic Bifidobacteria infantis: An assessment of potential antidepressant properties in the rat. Journal of Psychiatric Research, 43(2), 2008, p. 164–174 [Link] [4]. Based on the current understanding of the role of commensal microbiota in regulation of host brain biochemistry and behaviour, we hypothesize that by engineering bacteria for precisely controlled neurochemical production we can develop probiotic strains that could potentially act as new strategies for mental health treatment.

Our constructs

The design of our constructs was based on the analysis of metabolic pathways involved in synthesis of neurotransmitter compounds, including tryptophan pathway, glutamate pathway, and acetylcholine pathway. We have decided to develop the following constructs:

Tryptophan hydroxylase constructs
Serotonin is an essential metabolite, key neurotransmitter in mood regulation and a target of vast majority of antidepressant drugs. The majority of serotonin in the body is produced by enterochromaffin cells located in the gut [ref]. Metabolomics study has revealed that serotonin levels in blood plasma are 2.8 fold higher in conventional as opposed to germ-free mice [6], suggesting that the interactions between host and gut microflora play crucial role in regulating the production of serotonin and functioning of serotonergic system. The rate-limiting step of synthesis of serotonin is catalyzed by Tryptophan hydroxylase, TPH, which converts tryptophan, an essential amino acid, into 5-hydroxytryptophan (5-HTP) [7]. For this reason, tryptophan hydroxylase as a main effector for regulation of serotonin branch of tryptophan pathway in our Mind the Gut project.

BBa_K1598001 - the "naked" construct containing the TPH1 on its own:


BBa_K1598002 - the IPTG-inducible construct containing the Lac expression
cassete (BBa_413103) and the naked TPH1 (BBa_K1598001):


BBa_K1598003 - the constitutively expressed TPH1:

Modelling

Cloning