Periodic Reporting for period 4 - ARTISYM (Artificial endosymbiosis)
Période du rapport: 2021-07-01 au 2021-12-31
Living organisms have acquired new functionalities by uptake and integration of species to create symbiotic life-forms. This process of endosymbiosis has intrigued scientists over the years, albeit mostly from an evolution biology perspective. With the advance of chemical and synthetic biology, our ability to create molecular-life-like systems has increased tremendously, which enables us to build cell and organelle-like structures. However, these advances have not been taken to a level to study comprehensively if endosymbiosis can be applied to non-living systems or to integrate living with non-living matter. The aim of the research described in the ARTISYM project is to establish the field of artificial endosymbiosis. Two lines of research will be followed. First, we will incorporate artificial organelles in living cells to design hybrid cells with acquired functionality. This investigation is scientifically of great interest, as it will show us how to introduce novel compartmentalized pathways into living organisms. It also serves an important societal goal, as with these compartments dysfunctional cellular processes can be corrected. We will follow both a transient and a permanent approach. With the transient route biodegradable nanoreactors are introduced to supply living cells temporarily with novel function. Functionality is permanently introduced using genetic engineering to express protein-based nanoreactors in living cells, or via organelle transplantation of healthy mitochondria in diseased living cells. Secondly I aim to create artificial cells with the ability to perform endosymbiosis; the uptake and presence of artificial organelles in synthetic vesicles allows them to dynamically respond to their environment. Responses that are envisaged are shape changes, motility, and growth and division. Furthermore, the incorporation of natural organelles in liposomes provides
biocatalytic cascades with the necessary cofactors to function in an artificial cell.
biocatalytic cascades with the necessary cofactors to function in an artificial cell.
Our research is aimed to creating life-like systems that are reminiscent of living cells, and to add new functionality to living cells in order to make hybrid cells.
Regarding the latter, we have succeeded in constructing nanosized polymer capsules, of which the membrane is composed of a biodegradable polymer building block. Due to the unique composition of the polymer, the membrane was semipermeable for small molecules. Enzymes entrapped in the capsule could therefore still convert their substrate, and the subsequent product could leave the nanoreactor. We have been able to introduce these nanoreactors in living cells, in which they operated as artificial organelles. The reactors were able to protect cells from patients against oxidative stress. This is a form of enzyme replacement therapy that would allow a much more efficient usage of the enzymes, as they can still perform their function in the cell while being protected against factors that normally degrade the enzymes quickly.
In our artificial cell research, we have developed a unique platform that can mimic both the cell’s cytosol as well as the cell’s membrane. Until now protocell systems that were developed could only mimic either one of the two aspects. With our system, we now have the ability to study biological processes in a microenvironment that is highly reminiscent of a eukaryotic cell. The platform has the same type of crowdedness as what proteins experience when they are positioned in a living cell’s cytoplasm. Furthermore, the membrane offers stability and protection, and at the same time allows the exchange of molecules with the outside environment. As a result, these artificial cells can communicate with each other.
One of the hallmarks of life is out-of-equilibrium behavior: systems need constant input to be pushed into a transient, active state. We have developed artificial organelles with this specific behavior. We have constructed polymer membranes of which the permeability could be temporarily switched from an open to a closed state either by pH or by biological components such as ATP. Because of intrinsic elements in the system the organelles were autonomously switched back to their original state. We could demonstrate this phenomenon for organelles that showed substrate conversion and also motility.
Regarding the latter, we have succeeded in constructing nanosized polymer capsules, of which the membrane is composed of a biodegradable polymer building block. Due to the unique composition of the polymer, the membrane was semipermeable for small molecules. Enzymes entrapped in the capsule could therefore still convert their substrate, and the subsequent product could leave the nanoreactor. We have been able to introduce these nanoreactors in living cells, in which they operated as artificial organelles. The reactors were able to protect cells from patients against oxidative stress. This is a form of enzyme replacement therapy that would allow a much more efficient usage of the enzymes, as they can still perform their function in the cell while being protected against factors that normally degrade the enzymes quickly.
In our artificial cell research, we have developed a unique platform that can mimic both the cell’s cytosol as well as the cell’s membrane. Until now protocell systems that were developed could only mimic either one of the two aspects. With our system, we now have the ability to study biological processes in a microenvironment that is highly reminiscent of a eukaryotic cell. The platform has the same type of crowdedness as what proteins experience when they are positioned in a living cell’s cytoplasm. Furthermore, the membrane offers stability and protection, and at the same time allows the exchange of molecules with the outside environment. As a result, these artificial cells can communicate with each other.
One of the hallmarks of life is out-of-equilibrium behavior: systems need constant input to be pushed into a transient, active state. We have developed artificial organelles with this specific behavior. We have constructed polymer membranes of which the permeability could be temporarily switched from an open to a closed state either by pH or by biological components such as ATP. Because of intrinsic elements in the system the organelles were autonomously switched back to their original state. We could demonstrate this phenomenon for organelles that showed substrate conversion and also motility.
Within the Artisym project we have created a unique artificial cell platform and life-like out-of- equilibrium operating artificial organelles. Both elements have not been reported before and will form the basis for the next phase of development within the project.
First of all, we will combine the artificial organelles and cells in a multicompartment system with an architecture similar as to be found in a eukaryotic cell. Secondly, we expect these systems to be functional in communication processes between artificial cells and between living cells and artificial ones. The biological process operated from within one artificial cell will trigger a process in a neighbouring system. As a third element we will include motility as life like behavior in these artificial cell systems; artificial cells and organelles will be attracted by a fuel-sending cell in a chemotactic fashion. Ensemble movement of artificial cells will be designed, also in an out-of-equilibrium mode. Finally, natural organelles that are light-driven will be incorporated to execute/initiate biological processes within artificial cells.
Our hybrid cell work will be focused on the development and incorporation of artificial organelles that are key to cell survival or that can be used for bio-orthogonal chemistry. Furthermore, artificial organelles and cells will be explored for biomedical applications, in enzyme replacement strategies and as artificial antigen presenting cells.
First of all, we will combine the artificial organelles and cells in a multicompartment system with an architecture similar as to be found in a eukaryotic cell. Secondly, we expect these systems to be functional in communication processes between artificial cells and between living cells and artificial ones. The biological process operated from within one artificial cell will trigger a process in a neighbouring system. As a third element we will include motility as life like behavior in these artificial cell systems; artificial cells and organelles will be attracted by a fuel-sending cell in a chemotactic fashion. Ensemble movement of artificial cells will be designed, also in an out-of-equilibrium mode. Finally, natural organelles that are light-driven will be incorporated to execute/initiate biological processes within artificial cells.
Our hybrid cell work will be focused on the development and incorporation of artificial organelles that are key to cell survival or that can be used for bio-orthogonal chemistry. Furthermore, artificial organelles and cells will be explored for biomedical applications, in enzyme replacement strategies and as artificial antigen presenting cells.