Depression is a long-term mental health condition causing uncontrollable negative emotions, sleep disturbances, physical problems, and even thoughts of death. Severe depression is not only a personal problem but also has significant impacts on interpersonal relationships and society.
According to a global investigation launched by the World Health Organization (WHO) in 2023, 2.8 million people are suffering from depression worldwide, and the number of people with depression grows year by year (World Health Organization, 2023). Similarly, a survey conducted by the American Institute of Public Opinion indicated that more and more people are experiencing negative emotions, as shown in Figure 1 (Gallup, 2023). According to their report, the percentage of the global population experiencing negative emotions rose 9% from 2006 to 2022.
▲ Figure 1: Negative Experience Index Stable, 2022
Moreover, a study on elderly depression patients indicated that the reduction of negative emotions could significantly ease the non-emotional symptoms of depression and, consequently, decrease the negative impacts of depression (Arslanoglou E., et al. 2019). Hence, this year, the CCU-Taiwan iGEM team chose to tackle the problem of long-term negative emotions, which in turn will relieve depression and its serious consequences.
To start, we interviewed Min-Xiang Zhang, a clinical psychologist specializing in psychotherapy for depression and stress management, and conducted street surveys to better understand the issues surrounding negative emotions. The psychologist noted that while many think they are familiar with their emotions, they often lack true awareness. Our surveys also revealed a preference for more convenient methods to manage stress and alleviate negative emotions. Based on these insights, we developed a comprehensive approach focusing on two key areas: increasing emotional awareness and providing more convenient and accessible ways to enhance mood.
Recognizing and understanding emotions is vital for mental well-being, and the lack of awareness is reflected in the limited number of tools readily accessible to help individuals detect and understand their emotions.
To make emotional awareness more accessible, we developed a stress-detecting and managing app (Figure 2). No need for wearable devices; stress can be measured simply by placing a finger on the phone's flashlight and camera. By shining light through the fingertip and observing the subtle color changes caused by blood flow, we can measure Heart Rate Viability (HRV) to help people to detect their current emotional state.
▲ Figure 2: Stress-detecting app
Get to know more about our app at “Software”!
To further educate the public about emotional awareness and help people alleviate these negative feelings, we held an “emotion awareness” seminar, open lab, free hug events, and elderly activities.
Dive into “Human Practices” to discover how we held these events!
To address negative emotions in a way that's both easy and accessible, we created a solution that integrates seamlessly into daily life—specifically, through something Taiwanese people consume regularly: daily beverages. On average, Taiwanese people consume up to 6.7 cups of beverage per week (Health Promotion Administration, 2018), making this a natural platform for our innovation. This led to the development of SERENE Beverages, featuring a probiotic-infused edible film that allows the probiotics to produce 5-HTP directly in the gut.
5-Hydroxytryptophan (5-HTP) is a natural compound that serves as a precursor to serotonin, a neurotransmitter crucial for regulating mood, sleep, and appetite (Maffei, M. E, 2021). Supplements containing 5-HTP are often used to alleviate symptoms of depression, anxiety, and insomnia by increasing serotonin levels in the brain (Turner, E. H., et al. 2005).
Our project aims to revolutionize serotonin supplementation by genetically modifying the probiotic Nissle 1917 (EcN) to produce 5-HTP directly in the intestine. The 5-HTP produced will travel from the intestines to the brain through the bloodstream and be converted into serotonin. By enhancing 5-HTP synthesis locally in the gut, our genetically modified EcN could provide a reliable and accessible option for improving mental well-being.
In the serotonin biosynthesis pathway, tryptophan hydroxylase (TPH) catalyzes the formation of 5-HTP from L-tryptophan (Wang H, et al., 2018). TPH incorporates one atom of oxygen from molecular oxygen into the substrate and reduces the other atom to water. The two electrons required for reduction are supplied by oxidation of tetrahydrobiopterin (BH4) into BH4-4a-carbinolamine. The regeneration of BH4 from BH4-4a-carbinolamine is mediated by pterin-4a-carbinol-amine dehydratase (PCD) and dihydropteridine reductase (DHPR) (Figure 3) (Pribat et al. 2010).
▲ Figure 3: Serotonin synthesis pathway with BH4 regeneration system
Accordingly, we will express the monomeric human Tryptophan Hydroxylase 1 (m-hTPH1) protein, the human pterin-4a-carbinol-amine dehydratase (hPCBD1) protein, and the human q-dihydropteridine reductase (hQDPR) protein through polycistronic expression vector pST39 in EcN.
▲ Figure 4: Gene block for expressing 5-HTP to form three fusion proteins
To enhance 5-HTP production, we applied a DNA scaffold to bring m-hTPH1, hPCDB1, and hQDPR enzymes into close physical proximity. Accordingly, we fused zinc finger binding domains Zif268, PBSII, and ZFa with the m-hTPH1, hPCDB1, and hQDPR enzymes, respectively (Figure 4) (Raschellà G, et al., 2017). The corresponding binding motif for each zinc finger binding domain will be located on the DNA scaffold (Figure 5) (DeLisa M. P., et al., 2012).
▲ Figure 5: DNA binding platform to close physical proximity
To generate sufficient DNA scaffolds in EcN for assembly, we applied the rolling circle replication (RCR) mechanism to produce circular single-stranded DNA (ssDNA) containing sequences complementary to the binding motifs. The resulting circular ssDNA will automatically form the DNA scaffold through sequence complementation (Figure 6). To regulate the DNA scaffold formation, we applied the protocatechuate (PCA) induced promoter pPCA to control the expression of RepA (Figure 7), the replication initiation protein in RCR. Together, this will build a PCA-promoted 5-HTP biosynthesis pathway in EcN.
▲ Figure 6: The rolling circle replication (RCR) is applied to generate ssDNA containing sequences complementary to the zinc finger binding motifs, allowing the fusion proteins to bind to the DNA scaffold.
▲ Figure 7: Gene block for enhancing 5-HTP production through PCA-regulatory promoter
To enhance the accessibility of our probiotics, we devised an innovative approach of combining them with daily beverages by creating a water-soluble film to encase our probiotic powder underneath the sealing films of the beverage tea cups.
After conducting experiments with multiple materials, we've chosen sodium caseinate as our main ingredient, which has a relatively fast dissolution rate and does not affect the color of beverages. (Read more at “Application”)
The application of the edible film to the sealing film will be carried out by automated machinery with rollers and nozzles. (Discover more at “Implementation”). This novel delivery method integrates our product seamlessly into daily routines, making mental health support more convenient and accessible.
The core ambition of the CCU-Taiwan is to tackle the growing problem of negative emotions by providing a comprehensive approach. We genetically modify probiotic EcN to produce 5-HTP in the intestine, providing a sustainable and accessible alternative to current supplements. Our approach optimizes 5-HTP synthesis using a DNA-mediated biosynthesis pathway and integrates this solution into daily life with an innovative edible beverage film.
In addition to the synthetic biology strategy, we developed an app to help people monitor their stress levels. We promoted mental well-being and society connection through free hugs and activities for the elderly. With our project, we wish to target negative emotions through 5-HTP-producing probiotics and offer a comprehensive, accessible solution that significantly impacts global mental health.