Project description:Porphyridium species are known red microalgae for producing valuable bioactive compounds such as sulfated exopolysaccharides (EPS) with diverse industrial biomed-ical applications due to their functional and rheological properties. Recent studies have investigated how abiotic stresses, particularly nitrogen deprivation, affect Porphyridium’s metabolic regulation and EPS production through transcriptomic analysis. Still, the mechanisms governing EPS biosynthesis and the involvement of carbohydrate-activated enzymes (CAZymes) remain poorly understood. This study investigated the progressive effects of nitrate consumption on unicellular red alga P. purpureum, by integrating physiological, biochemical, and transcriptomic analyses through RNA-Seq, further validated by RT-qPCR. P. purpureum displayed a gradual, phase-dependent metabolic response to progressive nitrogen stress. EPS release coincided with the decline in nitrate uptake, linking nitrogen availability to carbon redirec-tion towards polysaccharide secretion. Transcriptomic data revealed global metabolic downregulation with targeted upregulation of stress-responsive, carbohydrate catabolic, and nucleotide-sugar synthesis pathways, including upregulation of CAZyme families GT4, GT8, and GT77. Our results give insights into the coordinated nitrogen and carbon metabolic regulation underlying polysaccharide biosynthesis, while open-ing future perspectives on enzyme compartmentalization and regulatory flux distribution under nitrogen stress in P. purpureum.
Project description:Microalgae are promising production platforms for the cost-effective production of recombinant proteins. We have recently established that the red alga Porphyridium purpureum provides superior transgene expression properties, due to the episomal main- tenance of transformation vectors as multicopy plasmids in the nucleus. Here, we have explored the potential of Porphyridium to synthesize complex pharmaceutical proteins to high levels. Testing expression constructs for a candidate subunit vaccine against the hepatitis C virus (HCV), we show that the soluble HCV E2 glycoprotein can be produced in transgenic algal cultures to high levels. The antigen undergoes faithful posttranslational modification by N-glycosylation and is recognized by conformationally selective antibodies, suggesting that it adopts a proper antigenic conformation in the endoplasmic reticulum of red algal cells. We also report the experimental determina- tion of the structure of the N-glycan moiety that is attached to glycosylated proteins in Porphyridium. Finally, we demonstrate the immunogenicity of the HCV antigen produced in red algae when administered by injection as pure protein or by feeding of algal biomass.
Project description:The limited knowledge we have about red algal genomes comes from the highly specialized extremophiles, Cyanidiophyceae. Here, we describe the first genome sequence from a mesophilic, unicellular red alga, Porphyridium purpureum. The 8,355 predicted genes in P. purpureum, hundreds of which are likely to be implicated in a history of horizontal gene transfer, reside in a genome of 19.7 Mbp with 235 spliceosomal introns. Analysis of light-harvesting complex proteins reveals a nuclear-encoded phycobiliprotein in the alga. We uncover a complex set of carbohydrate-active enzymes, identify the genes required for the methylerythritol phosphate pathway of isoprenoid biosynthesis, and find evidence of sexual reproduction. Analysis of the compact, function-rich genome of P. purpureum suggests that ancestral lineages of red algae acted as mediators of horizontal gene transfer between prokaryotes and photosynthetic eukaryotes, thereby significantly enriching genomes across the tree of photosynthetic life.