Search

Microalgae are unicellular photosynthetic organisms that can be grown in artificial systems to capture CO2, release oxygen, use nitrogen- and phosphorus-rich wastes, and produce biomass and bioproducts of interest including edible biomass for space exploration. In the present study, we report a metabolic engineering strategy for the green alga Chlamydomonas reinhardtii to produce high-value proteins for nutritional purposes. Chlamydomonas reinhardtii is a species approved by the U.S. Food and Drug Administration (FDA) for human consumption, and its consumption has been reported to improve gastrointestinal health in both murine models and humans. By utilizing the biotechnological tools available for this green alga, we introduced a synthetic gene encoding a chimeric protein, zeolin, obtained by merging the γ-zein and phaseolin proteins, in the algal genome. Zein and phaseolin are major seed storage proteins of maize (Zea mays) and bean (Phaseolus vulgaris) that accumulate in the endoplasmic reticulum (ER) and storage vacuoles, respectively. Seed storage proteins have unbalanced amino acid content, and for this reason, need to be complemented with each other in the diet. The chimeric recombinant zeolin protein represents an amino acid storage strategy with a balanced amino acid profile. Zeolin protein was thus efficiently expressed in Chlamydomonas reinhardtii; thus, we obtained strains that accumulate this recombinant protein in the endoplasmic reticulum, reaching a concentration up to 5.5 fg cell-1, or secrete it in the growth medium, with a titer value up to 82 µg/L, enabling the production of microalga-based super-food.

In eukaryotes, many proteins contain an N-terminal signal peptide that allows their translocation into the endoplasmic reticulum followed by secretion outside the cell according to the classical secretory system. However, an increasing number of secreted proteins lacking the signal peptide sequence are emerging. These proteins, secreted in several alternative ways collectively known as unconventional protein secretion (UPS) pathways, exert extracellular functions including cell signaling, immune modulation, as well as moonlighting activities different from their well-described intracellular functions. Pathways for UPS include direct transfer across the plasma membrane, secretion from endosomal/multivesicular body-related components, release within plasma membrane-derived microvesicles, or use of elements of autophagy. In this review we describe the mammals and plants UPS pathways identified so far highlighting commonalities and differences.

Abstract  Human Papillomavirus (HPV) is the causal agent of cervical cancer, one of the most common causes of death for women. The major capsid L1 protein self-assembles in Virus Like Particles (VLPs), which are highly immunogenic and suitable for vaccine production. In this study, a plastid transformation approach was assessed in order to produce a plant-based HPV-16 L1 vaccine. Transplastomic plants were obtained after transformation with vectors carrying a chimeric gene encoding the L1 protein either as the native viral (L1v gene) or a synthetic sequence optimized for expression in plant plastids (L1pt gene) under control of plastid expression signals. The L1 mRNA was detected in plastids and the L1 antigen accumulated up to 1.5% total leaf proteins only when vectors included the 5′-UTR and a short N-terminal coding segment (Downstream Box) of a plastid gene. The half-life of the engineered L1 protein, determined by pulse-chase experiments, is at least 8 h. Formation of immunogenic VLPs in chloroplasts was confirmed by capture ELISA assay using antibodies recognizing conformational epitopes and by electron microscopy. [ABSTRACT FROM AUTHOR]