Recent advancements in the genetic engineering of microalgae

Highlights

• The genetic toolkit for engineering microalgae is currently expanding.

• New molecular tools have been utilized to produce algal-derived therapeutics.

• Microalgae production strains have been improved using evolutionary engineering.

Abstract

The development of more sustainable food, feed, and bio-products is critical to mitigating the environmental stresses facing our world today. Algae, which includes seaweeds, eukaryotic microalgae, and cyanobacteria, are a promising platform to achieving this, as they have low energy and space requirements, are safe for human and animal consumption, and can be manipulated to produce a diversity of valuable bioproducts. This review focuses on microalgae, both eukaryotic and cyanobacteria. In the past, addressing the major challenges of bringing microalgal production systems to an economically viable scale only had a relatively small genetic toolset to work with, in comparison to other microbial systems such as bacteria and yeast. Expanding the molecular tools available for genetic engineering of microalgae will lead to higher product yields, and accelerate the development of new microalgal bioproducts for commercial applications, thereby supporting the shift towards more environmentally friendly products. In this review, we highlight significant advances from recent years on the design of microalgal expression vectors, discovery of genetic regulatory elements (promoters and transcription factors), optimization of transformation methods, and development of new strain improvement techniques, all aimed at advancing microalgae to become a more efficient biomanufacturing platform. We then discuss how these tools have been applied to improving recombinant protein production, and to enhance metabolic pathway engineering.

Introduction

Microalgae, both eukaryotic and cyanobacteria taxa, are being developed as a sustainable platform for the biomanufacturing of a variety of desirable products including foods, feeds, fuels, materials, and medicines. Due to their rapid growth, efficient use of resources, and natural genetic diversity, a wide variety of algal-derived compounds can be targeted for cost-competitive production (Fig. 1). Unlike other microorganisms, including yeast and bacteria, multiple species of microalgae are safe for human consumption and have obtained GRAS (Generally Recognized as Safe) status from the US FDA, potentially reducing purification costs of algal-derived therapeutics and expanding the potential for valuable food and feed ingredients [1]. Historically, a focus of this field has been on the commercial production of biofuels from microalgae, however due to the low cost of fossil fuels, economically viable biofuels from algae have yet to be achieved at any significant scale and focus has largely shifted towards coproduct production to offset the costs [[2], [3], [4]]. In order to achieve economic viability of commodity goods produced in algae, the entire algae biomass must be valorized through the production of valuable coproducts. Many commercially desirable compounds exist naturally in microalgae, including a variety of lipids, vitamins, pigments, and the documented highly nutritional biomass itself [5], with the potential to express high value recombinant proteins using synthetic biology. Currently, the largest challenge to developing more algae bioproducts at a commercial level is bringing production from lab scale to an industrial scale. Optimization of strains, and expansion of genetic toolsets for manipulating the strains into producing high yields of target products will be instrumental in accomplishing this.

Regardless of the intended product, molecular and genetic tools offer powerful approaches to enhance the production of targeted molecules. Development of a sophisticated suite of genetic tools in microalgae is still incomplete, compared to other industrial microorganism platforms. Nevertheless, the toolsets currently available have greatly facilitated the speed and accuracy with which one can engineer algae to optimize productivity. Metabolic engineering involves targeting specific parts of metabolic pathways within cells to change the flux of the metabolites towards a desired product [6]. Recent metabolic engineering efforts in photosynthetic microorganisms have been focused on overexpressing or knocking out key genes that encode enzymes in pathways of interest. Studies involving metabolic engineering of photosynthetic microorganisms have focused on a variety of products, with many in the fatty acid or isoprenoid biosynthetic pathways. Engineering recombinant proteins in microalgae, on the other hand, involves inserting an exogenous gene encoding for the desired protein into the algal genome in a strategic way, enabling the protein to be expressed at high levels and targeted to a specific subcellular region of the algal cells. One challenge in utilizing molecular tools in algae has been the genetic difference between species, where the interspecies genetic variability limits the use of genetic tools from one species to another. However, as the cost of genome sequencing continues to decline, more microalgal strains can be easily characterized, and the tools then adapted to their genetic code. As a result, research in this field is broadening and more widely applicable genetic toolkits are being developed to enable various species of microalgae as industrial production systems. Here we review the most current research advances and discuss how these new tools are enabling us to bring algae forward as a true commercial platform.