Towards yeast as a tool for in vivo continuous evolution of TRPV1 modulators
(2020) KMBM05 20201Applied Microbiology
- Abstract
- The TRPV1 receptor (transient receptor potential vanilloid 1) is a nonspecific membrane-bound cation channel, which plays an important role in nociception and pain transmission, the nerve-response to possibly dangerous stimuli. It is known that TRPV1 can be modulated by a variety of stimuli, amongst which heat, protons and chemical compounds, such as capsaicin, are the most known activators. This makes the TRPV1 receptor an interesting target for drugs that seek to regulate and reduce pain. However, many of the chemical compounds that are used to target this receptor, e.g. for the purpose of pain-relief, also suffer the risk of severe side effects. In this project, a first step was made in investigating the possibility of applying in vivo... (More)
- The TRPV1 receptor (transient receptor potential vanilloid 1) is a nonspecific membrane-bound cation channel, which plays an important role in nociception and pain transmission, the nerve-response to possibly dangerous stimuli. It is known that TRPV1 can be modulated by a variety of stimuli, amongst which heat, protons and chemical compounds, such as capsaicin, are the most known activators. This makes the TRPV1 receptor an interesting target for drugs that seek to regulate and reduce pain. However, many of the chemical compounds that are used to target this receptor, e.g. for the purpose of pain-relief, also suffer the risk of severe side effects. In this project, a first step was made in investigating the possibility of applying in vivo continuous evolution (or ICE), as an emerging method, for novel drug discovery and development. The work was focused on finding and evaluating an ideal selection system for receptor modulation in a yeast strain overexpressing the TRPV1 receptor. By investigating different combinations of growth conditions, such as temperature, pH, external calcium levels, as well as looking into the level of TRPV1 expression, the optimal landscape for TRPV1 modulating conditions was explored. Furthermore, the effect of supplementing the growth media with the known modulators capsaicin (agonist) and capsazepine (antagonist) was also investigated. Throughout the experiments, cell growth and viability was monitored and internal calcium fluorescence was analyzed by flow-cytometry. It was found that the condition combinations tested affected yeast strains with different expression levels of TRPV1 differently. Additionally, the yeast strains overexpressing the receptor had lower growth in all growth conditions and increased internal calcium uptake in most conditions, compared to the control. The expression level of TRPV1, as well as the addition of chemical modulators in the media was also shown to have an effect on the growth and internal calcium levels. The findings in the factors affecting TRPV1 modulation provide a stepping stone in constructing an optimal selection system. (Less)
- Popular Abstract
- Throughout the history of mankind, drug discovery and development have played a crucial role for public health and establishing a higher standard of living. Its importance shines even brighter in times like these, where a global pandemic plagues our world. Therefore, it is important to constantly push this frontier of drug development forward, towards newer and more efficient methods.
In this Master Thesis, the development of in vivo continuous evolution, or ICE, as method for the drug discovery of capsaicinoids using baker’s yeast, was investigated.
Capsaicinoids are of great interest, as they modulate the TRPV1 receptor, which plays a role in the transmission of pain, and have been used for the treatment of just pain,... (More) - Throughout the history of mankind, drug discovery and development have played a crucial role for public health and establishing a higher standard of living. Its importance shines even brighter in times like these, where a global pandemic plagues our world. Therefore, it is important to constantly push this frontier of drug development forward, towards newer and more efficient methods.
In this Master Thesis, the development of in vivo continuous evolution, or ICE, as method for the drug discovery of capsaicinoids using baker’s yeast, was investigated.
Capsaicinoids are of great interest, as they modulate the TRPV1 receptor, which plays a role in the transmission of pain, and have been used for the treatment of just pain, inflammation or even cancer. However, due to potentially severe side-effects of using these pharmaceutical compounds, new and improved versions are needed. But how can one generate these new compounds without losing the beneficial effects? The answer could be ICE.
ICE is, as the name suggests, a continuous system, which occurs in cycles. It consists of three parts: First you generate a genetic variation within a population of organisms (hence, in vivo) to get a wide and diverse population, in this project, yeast cells. Following this, the population is screened and a small subpopulation of cells with desirable traits are selected. In this thesis, this would be strains of yeast producing suitable capsaicinoids. Lastly, the selected population is amplified, which occurs naturally in a growth based ICE-system.
By providing some fundamental understanding about the environmental effects on the TRPV1 receptor, for the growth based system, as well as investigating the effects of known capsaicinoids on the receptor, when added to the growth media, this thesis provides future research on the topic with a good base for new research questions and experiments. (Less)
Please use this url to cite or link to this publication:
http://lup.lub.lu.se/student-papers/record/9017654
- author
- Marthin, Malte LU
- supervisor
- organization
- alternative title
- Mot jäst som ett verktyg för in vivo kontinuerlig evolution av TRPV1 reglerande molekyler
- course
- KMBM05 20201
- year
- 2020
- type
- H2 - Master's Degree (Two Years)
- subject
- language
- English
- id
- 9017654
- date added to LUP
- 2020-06-15 14:11:12
- date last changed
- 2020-06-16 08:28:10
@misc{9017654,
abstract = {{The TRPV1 receptor (transient receptor potential vanilloid 1) is a nonspecific membrane-bound cation channel, which plays an important role in nociception and pain transmission, the nerve-response to possibly dangerous stimuli. It is known that TRPV1 can be modulated by a variety of stimuli, amongst which heat, protons and chemical compounds, such as capsaicin, are the most known activators. This makes the TRPV1 receptor an interesting target for drugs that seek to regulate and reduce pain. However, many of the chemical compounds that are used to target this receptor, e.g. for the purpose of pain-relief, also suffer the risk of severe side effects. In this project, a first step was made in investigating the possibility of applying in vivo continuous evolution (or ICE), as an emerging method, for novel drug discovery and development. The work was focused on finding and evaluating an ideal selection system for receptor modulation in a yeast strain overexpressing the TRPV1 receptor. By investigating different combinations of growth conditions, such as temperature, pH, external calcium levels, as well as looking into the level of TRPV1 expression, the optimal landscape for TRPV1 modulating conditions was explored. Furthermore, the effect of supplementing the growth media with the known modulators capsaicin (agonist) and capsazepine (antagonist) was also investigated. Throughout the experiments, cell growth and viability was monitored and internal calcium fluorescence was analyzed by flow-cytometry. It was found that the condition combinations tested affected yeast strains with different expression levels of TRPV1 differently. Additionally, the yeast strains overexpressing the receptor had lower growth in all growth conditions and increased internal calcium uptake in most conditions, compared to the control. The expression level of TRPV1, as well as the addition of chemical modulators in the media was also shown to have an effect on the growth and internal calcium levels. The findings in the factors affecting TRPV1 modulation provide a stepping stone in constructing an optimal selection system.}},
author = {{Marthin, Malte}},
language = {{eng}},
note = {{Student Paper}},
title = {{Towards yeast as a tool for in vivo continuous evolution of TRPV1 modulators}},
year = {{2020}},
}