Pasquale Stano

A Possible Route to Prebiotic Vesicle Reproduction

Spherical bounded structures such as those formed by surfactant aggregates (mostly micelles and vesicles), with an inside that is chemically and physically different from the outside medium, can be seen as primitive cell models. As such, they are fundamental structures for the theory of autopoiesis as originally formulated by Varela and Maturana. In particular, since self-reproduction is a very important feature of minimal cellular life, the study of self-reproduction of micelles and vesicles represents a quite challenging bio-mimetic approach. Our laboratory has put much effort in recent years into implementing self-reproduction of vesicles as models for self-reproduction of cellular bounded structures, and this article is a further contribution in this direction. In particular, we deal with the so-called matrix effect of vesicles, related to the fact that when fresh surfactant is added to an aqueous solution containing preformed vesicles of a very narrow size distribution, the newly formed vesicles (instead of being polydisperse, as is usually the case) have dimensions very close to those of the preformed ones. In practice, this corresponds to a mechanism of reproduction of vesicles of the same size. In this article, the matrix effect is re-elaborated in the perspective of the origin of life, and in particular in terms of the prebiotic mechanisms that might permit the growth and reproduction of vesicles. The data are analyzed by dynamic light scattering with a new program that permits the calculation of the number-weighted size distribution. It is shown that, on adding a stoichiometric amount of oleate micelles to preformed oleate vesicles extruded at 50 and 100 nm, the final distribution contains about twice the initial number of particles, centered around 50 and 100 nm. The same holds when oleate is added to preformed phospholipid liposomes. By contrast, when the same amount of oleate is added to an aqueous solution (as a control experiment), a very broad distribution ranging between 20 and 1000 nm is obtained. The data can then be seen as a kind of reproduction of the same size vesicles, and the argument is advanced that this may correspond to a simple prebiotic mechanism of vesicle multiplication in prebiotic times, when only physical forces might be responsible for the basic mechanisms of early protocell growth and division. Preliminary data also show that repeated addition of oleate maintains the same basic initial features, and that surfactants other than oleate also respect the reproductive mode of the matrix effect.

Recent Theoretical Approaches to Minimal Artificial Cells

Minimal artificial cells (MACs) are self-assembled chemical systems able to mimic the behavior of living cells at a minimal level, i.e. to exhibit self-maintenance, self-reproduction and the capability of evolution. The bottom-up approach to the construction of MACs is mainly based on the encapsulation of chemical reacting systems inside lipid vesicles, i.e. chemical systems enclosed (compartmentalized) by a double-layered lipid membrane. Several researchers are currently interested in synthesizing such simple cellular models for biotechnological purposes or for investigating origin of life scenarios. Within this context, the properties of lipid vesicles (e.g., their stability, permeability, growth dynamics, potential to host reactions or undergo division processes…) play a central role, in combination with the dynamics of the encapsulated chemical or biochemical networks. Thus, from a theoretical standpoint, it is very important to develop kinetic equations in order to explore first—and specify later—the conditions that allow the robust implementation of these complex chemically reacting systems, as well as their controlled reproduction. Due to being compartmentalized in small volumes, the population of reacting molecules can be very low in terms of the number of molecules and therefore their behavior becomes highly affected by stochastic effects both in the time course of reactions and in occupancy distribution among the vesicle population. In this short review we report our mathematical approaches to model artificial cell systems in this complex scenario by giving a summary of three recent simulations studies on the topic of primitive cell (protocell) systems.

Synthesis and Investigation of Tryptophan–Amphotericin B Conjugates

Anchors aweigh! The synthesis of tryptophan–amphotericin B conjugates (see figure) is described. The membrane‐anchoring effect of tryptophane was thus combined with the pore‐formation effect of amphotericin B leading to high channel activity in sterol‐free liposomes.

The atypical retinoid E -3-(3’-Adamantan-1-yl-4’-methoxybiphenyl-4-yl)-2-propenoic acid (ST1898) displays comedolytic activity in the rhino mouse model

Retinoids represent the first-line therapy for the treatment of acne vulgaris. Their effect is comedolytic and anti-comedogenic, and associates with hyperplasia and deregulated differentiation of the epidermis, and decreased inflammation. We here tested the comedolytic effect of the novel atypical retinoid E-3-(3-Adamantan-1-yl-4-methoxybiphenyl-4-yl)-2-propenoic acid (ST1898) in the rhino mouse, as a model of comedogenic acne, and compared this effect to that of adapalene (Differin® gel), as reference compound. Topical administration of 0.1% ST1898 for three weeks exerted a comedolytic effect comparable to that of adapalene 0.1%. In ST1898-treated mice, epidermal hyperplasia was significantly reduced and the expression of keratinocyte differentiation markers was less perturbed compared to adapalene-treated animals. Moreover, keratin 6, which stains activated keratinocytes, was strongly and uniformly induced in interfollicular epidermis of adapalene-treated mice, while only faintly and focally expressed in ST1898-treated ones. Our data indicate that ST1898 has strong comedolytic activity but modest topical side effects.

New and Unexpected Insights on the Formation of Protocells from a Synthetic Biology Approach: The Case of Entrapment of Biomacromolecules and Protein Synthesis Inside Vesicles

In this chapter we present our “semi-synthetic” approach to the construction of minimal living cells.

Interfacing Synthetic Cells with Biological Cells: An Application of the Synthetic Method

The “synthetic method” is the methodological approach that guides current scientific attempts of understanding natural processes by the construction of hardware, software, and/or wetware models fro...

Chapter 11:Chemical synthetic biology projects: never born biopolymers and synthetic cells

“Chemical” synthetic biology can be defined as a branch of synthetic biology focused on the synthesis of chemical structures alternative to those present in nature. Here we present two chemical synthetic biology projects, namely (1) the Never Born Biopolymers and (2) the Synthetic Minimal Cells. The first project aims at identifying and constructing biopolymers like nucleic acids and proteins that do not exist in nature and that display biological-like functions. The goal of the second project is instead focused on the assembly of cell-like structures, based on liposomes, that behave like simple cells. The concepts and the experimental approaches concerning these projects are shortly summarized and discussed.

On the Construction of Minimal Cell Models in Synthetic Biology and Origins of Life Studies

In this chapter we describe the concept of minimal living cells, defined as synthetic or semi-synthetic cells having the minimal and sufficient number of components to be endowed with the main biological properties of living cells. The construction of minimal cells starting from isolated compounds is an issue in synthetic biology, origins of life studies, and biotechnology. We start by discussing the different concepts underlining the three above-mentioned fields, by comparing the different viewpoints and highlighting common perspectives. We focus on the first two approaches, firstly describing our recent investigation on the construction of semi-synthetic minimal cells (developed in the Synthcells project), based on the use of liposomes as cell models. A short review of most relevant studies in the field is also given. The emphasis is then shifted to more basic biophysical aspects that emerged from these studies and that can significantly contribute to the understanding of the origins of primitive cells. In particular, we report the unexpected finding of spontaneous self-concentration of proteins and other solutes inside lipid vesicles. This recent discovery gives rise to a several theoretical and experimental implications that are shortly discussed. As a conclusion, we comment on the state-of-the-art in the field, next developments, and future challenges, and highlight how this research may contribute to improve our understanding of life.