Search

Abstract Microorganisms can be engineered to sustainably produce a variety of products including fuels, pharmaceuticals, materials, and food. However, highly engineered strains often result in low production yield, due to undesired effects such as metabolic burden and the toxicity of intermediates. Drawing inspiration from natural ecosystems, the construction of a synthetic community with division of labor can offer advantages for bioproduction. This approach involves dividing specific tasks among community members, thereby enhancing the functionality of each member. In this study, we identify six pairs out of fifteen composed of six auxotrophs of Yarrowia lipolytica that spontaneously form robust syntrophic and synergistic communities. We characterize the stability and growth dynamics of these communities. Furthermore, we validate the existence of syntrophic interactions between two yeast species, Y. lipolytica and Saccharomyces cerevisiae, and find a strain combination, Δtrp2 and Δtrp4, forming a stable syntrophic community between two species. Subsequently, we introduce a 3-hydroxypropionic acid (3-HP) biosynthesis pathway into the syntrophic community by dividing the pathway among different strains. Our results demonstrate improved production of 3-HP in both intra- and interspecies communities compared to monocultures. Our results show the stable formation of synthetic syntrophic communities, and their potential in improving bioproduction processes.

Pedagogization can be understood as an umbrella term for increased educational action. In settler-colonial contexts, the pedagogization of traditional Indigenous games has gained traction in recent years. As noted by a number of academic studies, traditional games have been used in efforts to promote cultural connectedness, inclusion, and engagement and address social justice in physical education programs. This commentary aims to survey the pedagogization of traditional Indigenous games in Canada, Australia, and Aotearoa New Zealand. We argue, with cautious optimism, that traditional Indigenous games may act as a conduit for more culturally relevant physical education curricula and reconciliatory pedagogical training for physical education teachers.

This paper aims to extend the small but growing body of global literature on the topic of child safeguarding (CSG) policy in sport and related educational contexts. The authors, all male coaches/educators, offer 'snapshots' of the moments in time they each realised their previously unquestioned practices must change in light of shifting societal attitudes to CSG and resulting legislation. Our contributions in this space are two-fold. Firstly, we present an autoethnographic methodology that provides a lens into the challenges confronting pedagogues in sport and related educational contexts. Through this methodology, we broaden the scope of discourse that we deem necessary for future CSG policy direction and operationalisation. Secondly, we explore and include an addition to the dominant 'duality (dichotomy) of danger' narrative discourse currently reflected in the literature. We do this by proposing a trichotomy of danger framework for leaders and managers of sport and related educational contexts to consider when plotting this future landscape. Research surrounding the possible implications of CSG policy on practice is critical if educators are to navigate the 'risks' of their professions. Although there is a small and emerging body of research on this topic, we, the authors seek to redress the scarcity of research observable in the Aotearoa New Zealand context.

CRISPR gene activation and inhibition has become a powerful synthetic tool for influencing the expression of native genes for foundational studies, cellular reprograming, and metabolic engineering. Here the authors demonstrate near leak-free, inducible expression of a polycistronic array containing up to 24 gRNAs from two orthogonal CRISPR/Cas systems.

The concept of physical literacy (PL) has increasingly drawn global interest from practitioners, academics and policymakers within the fields of sport, health and physical education. Its value and appeal appear to lie in its potential to help refocus attention on the importance of human physicality and movement in enabling individuals to flourish and lead fulfilling lives Whitehead, M. (Ed.). (2010). "Physical literacy: Through the life-course. International studies in physical education and youth sport." Routledge. In this sense, it overlaps with the aims of physical education (PE) and has increasingly been seen as having an influence on future curriculum development in many Western countries. In our view, there is a need to critically question this influence, particularly in the sense that the concept is not just redefining the nomenclature of the subject area; but also its nature, focus and outcomes. This need to critically question has also become more prevalent as neoliberal technocratic agendas have pushed for clearer measures of the impact associated with government expenditure, increased individual responsibility for health outcomes and the subsequent development of measurement tools and standards. The concern here, particularly for a country like Aotearoa New Zealand, is that the current curriculum risks becoming colonised by a concept, that fails to adequately address its national values, cultural identity and broad educative aspirations.

Human milk fat has a distinctive stereoisomeric structure where palmitic acid is esterified to the middle (sn-2) position on the glycerol backbone of the triacylglycerol and unsaturated fatty acids to the outer (sn-1/3) positions. This configuration allows for more efficient nutrient absorption in the infant gut. However, the fat used in most infant formulas originates from plants, which exclude palmitic acid from the sn-2 position. Oleaginous yeasts provide an alternative source of lipids for human nutrition. However, these yeasts also exclude palmitic acid from the sn-2 position of their triacylglycerol. Here we show that Yarrowia lipolytica can be engineered to produce triacylglycerol with more than 60% of the palmitic acid in the sn-2 position, by expression of lysophosphatidic acid acyltransferases with palmitoyl-Coenzyme A specificity. The engineered Y. lipolytica strains can be cultured on glycerol, glucose, palm oil or a mixture of substrates, under nitrogen limited condition, to produce triacylglycerol with a fatty acid composition that resembles human milk fat, in terms of the major molecular species (palmitic, oleic and linoleic acids). Culture on palm oil or a mixture of glucose and palm oil produced the highest lipid titre and a triacylglycerol composition that is most similar with human milk fat. Our data show that an oleaginous yeast can be engineered to produce a human milk fat substitute (β-palmitate), that could be used as an ingredient in infant formulas.

Abstract The oleaginous yeast Yarrowia lipolytica is a potent cell factory as it is able to use a wide variety of carbon sources to convert waste materials into value-added products. Nonetheless, there are still gaps in our understanding of its central carbon metabolism. Here we present an in-depth study of Y. lipolytica hexokinase (YlHxk1), a structurally unique protein. The greatest peculiarity of YlHxk1 is a 37-amino acid loop region, a structure not found in any other known hexokinases. By combining bioinformatic and experimental methods we showed that the loop in YlHxk1 is essential for activity of this protein and through that on growth of Y. lipolytica on glucose and fructose. We further proved that the loop in YlHxk1 hinders binding with trehalose 6-phosphate (T6P), a glycolysis inhibitor, as hexokinase with partial deletion of this region is 4.7-fold less sensitive to this molecule. We also found that YlHxk1 devoid of the loop causes strong repressive effect on lipase-encoding genes LIP2 and LIP8 and that the hexokinase overexpression in Y. lipolytica changes glycerol over glucose preference when cultivated in media containing both substrates.

Yarrowia lipolytica is an important industrial microorganism used for the production of oleochemicals. The design of effective biotechnological processes with this cell factory requires an in-depth knowledge of its metabolism. Here we present a transcriptomic study of Y. lipolytica grown in the presence of glycerol and glucose, and mixture of both at different carbon to nitrogen ratios. It emerged that the transcriptomic landscape of Y. lipolytica is more sensitive to the nitrogen availability than to the utilized carbon source, as evidenced by more genes being differentially expressed in lower carbon to nitrogen ratio. Specifically, expression of hexokinase (HXK1) is significantly susceptible to changes in nitrogen concentrations. High HXK1 expression in low nitrogen seems to impact other genes which are implicated in tricarboxylic acid cycle and erythritol biosynthesis. We further show that expression of HXK1 and two genes belonging to the sugar porter family might be controlled by GATA-like zinc-finger proteins.

The development of efficient bioprocesses requires inexpensive and renewable substrates. Molasses, a by-product of the sugar industry, contains mostly sucrose, a disaccharide composed of glucose and fructose, both easily absorbed by microorganisms. Yarrowia lipolytica, a platform for the production of various chemicals, can be engineered for sucrose utilization by heterologous invertase expression, yet the problem of preferential use of glucose over fructose remains, as fructose consumption begins only after glucose depletion what significantly extends the bioprocesses. We investigated the role of hexose transporters and hexokinase (native and fructophilic) in this preference. Analysis of growth profiles and kinetics of monosaccharide utilization has proven that the glucose preference in Y. lipolytica depends primarily on the affinity of native hexokinase for glucose. Interestingly, combined overexpression of either hexokinase with hexose transporters significantly accelerated citric acid biosynthesis and enhanced pentose phosphate pathway leading to secretion of polyols (31.5 g/L vs. no polyols in the control strain). So far, polyol biosynthesis was efficient in glycerol-containing media. Moreover, overexpression of fructophilic hexokinase in combination with hexose transporters not only shortened this process to 48 h (84 h for the medium with glycerol) but also allowed to obtain 23% more polyols (40 g/L) compared to the glycerol medium (32.5 g/L).

Applying low-cost substrate is critical for sustainable bioproduction. Co-culture of phototrophic and heterotrophic microorganisms can be a promising solution as they can use CO 2 and light as feedstock. This study aimed to create a light-driven consortium using a marine cyanobacterium Synechococcus sp. PCC 7002 and an industrial yeast Yarrowia lipolytica. First, the cyanobacterium was engineered to accumulate and secrete sucrose by regulating the expression of genes involved in sucrose biosynthesis and transport, resulting in 4.0 g/L of sucrose secretion. Then, Yarrowia lipolytica was engineered to efficiently use sucrose and produce β-caryophyllene that has various industrial applications. Then, co- and sequential-culture were optimized with different induction conditions and media compositions. A maximum β-caryophyllene yield of 14.1 mg/L was obtained from the co-culture. This study successfully established an artificial light-driven consortium based on a marine cyanobacterium and Y. lipolytica, and provides a foundation for sustainable bioproduction from CO 2 and light through co-culture systems., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The global market for high-value fatty acids production, mainly omega-3/6, hydroxy fatty-acids, waxes and their derivatives, has seen strong development in the last decade. The reason for this growth was the increasing utilization of these lipids as significant ingredients for cosmetics, food and the oleochemical industries. The large demand for these compounds resulted in a greater scientific interest in research focused on alternative sources of oil production - among which microorganisms attracted the most attention. Microbial oil production offers the possibility to engineer the pathways and store lipids enriched with the desired fatty acids. Moreover, costly chemical steps are avoided and direct commercial use of these fatty acids is available. Among all microorganisms, the oleaginous yeasts have become the most promising hosts for lipid production - their efficient lipogenesis, ability to use various (often highly affordable) carbon sources, feasible large-scale cultivations and wide range of available genetic engineering tools turns them into powerful micro-factories. This review is an in-depth description of the recent developments in the engineering of the lipid biosynthetic pathway with oleaginous yeasts. The different classes of valuable lipid compounds with their derivatives are described and their importance for human health and industry is presented. The emphasis is also placed on the optimization of culture conditions in order to improve the yield and titer of these valuable compounds. Furthermore, the important economic aspects of the current microbial oil production are discussed.

The development of efficient bioprocesses requires inexpensive and renewable substrates. Molasses, a by-product of the sugar industry, contains mostly sucrose, a disaccharide composed of glucose and fructose, both easily absorbed by microorganisms. Yarrowia lipolytica , a platform for the production of various chemicals, can be engineered for sucrose utilization by heterologous invertase expression, yet the problem of preferential use of glucose over fructose remains, as fructose consumption begins only after glucose depletion what significantly extends the bioprocesses. We investigated the role of hexose transporters and hexokinase (native and fructophilic) in this preference. Analysis of growth profiles and kinetics of monosaccharide utilization has proven that the glucose preference in Y. lipolytica depends primarily on the affinity of native hexokinase for glucose. Interestingly, combined overexpression of either hexokinase with hexose transporters significantly accelerated citric acid biosynthesis and enhanced pentose phosphate pathway leading to secretion of polyols (31.5 g/L vs. no polyols in the control strain). So far, polyol biosynthesis was efficient in glycerol-containing media. Moreover, overexpression of fructophilic hexokinase in combination with hexose transporters not only shortened this process to 48 h (84 h for the medium with glycerol) but also allowed to obtain 23% more polyols (40 g/L) compared to the glycerol medium (32.5 g/L).

Yarrowia lipolytica is an important industrial microorganism used for the production of oleochemicals. The design of effective biotechnological processes with this cell factory requires an in-depth knowledge of its metabolism. Here we present a transcriptomic study of Y. lipolytica grown in the presence of glycerol and glucose, and mixture of both at different carbon to nitrogen ratios. It emerged that the transcriptomic landscape of Y. lipolytica is more sensitive to the nitrogen availability than to the utilized carbon source, as evidenced by more genes being differentially expressed in lower carbon to nitrogen ratio. Specifically, expression of hexokinase ( HXK1 ) is significantly susceptible to changes in nitrogen concentrations. High HXK1 expression in low nitrogen seems to impact other genes which are implicated in tricarboxylic acid cycle and erythritol biosynthesis. We further show that expression of HXK1 and two genes belonging to the sugar porter family might be controlled by GATA-like zinc-finger proteins., (© 2020 The Authors. Published by Elsevier B.V.)

In an era of ever-increasing energy demands, a promising technology is being developed: the use of oleaginous microorganisms such as Yarrowia lipolytica to convert waste materials into biofuels. Here, we constructed two Y. lipolytica strains that displayed both increased lipid accumulation and more efficient use of biomass-derived sugars, including glucose, fructose, galactose and inulin. The first strain, Y. lipolytica YLZ150, was derived from the French wild-type strain W29. It had inhibited triacylglycerol mobilization (∆tgl4) and β-oxidation (∆pox1-6), and it overexpressed GPD1, DGA2, HXK1, the native Leloir pathway, SUC2 from Saccharomyces cerevisiae and INU1 from Kluyveromyces marxianus. The second strain, Y. lipolytica Y4779, was derived from the Polish A-101 strain. It had inhibited β-oxidation (∆mfe2) and overexpressed GPD1, DGA1, HXK1, YHT3, SUC2 and INU1. In the first experiment, strain YLZ150 was batch-cultured in media containing different hexoses; the highest values for lipid concentration and yield of lipids from the substrate were obtained using fructose (20.3 g dm -3 and 0.14 g g -1 , respectively). In the second experiment, we grew the two strains in fed-batch cultures to examine lipid biosynthesis from inulin (a fructose polymer). For Y4779, the lipid concentration was 10.3 g dm -3 and the yield of lipids from substrate was 0.07 g g -1 ; in contrast, for YLZ150, these values were 24 g dm -3 and 0.16 g g -1 , respectively. The YLZ150 strain is thus able to efficiently exploit glucose, fructose, galactose, sucrose and inulin for lipid biosynthesis. Copyright © 2017 John Wiley & Sons, Ltd., (Copyright © 2017 John Wiley & Sons, Ltd.)