Nitrate Assimilatory Genes and Their Transcriptional Regulation in a Unicellular Red Alga Cyanidioschyzon merolae: Genetic Evidence for Nitrite Reduction by a Sulfite Reductase-Like Enzyme

Abstract

Cyanidioschyzon merolae is a unicellular red alga living in acid hot springs, which is able to grow on ammonium, as well as nitrate as sole nitrogen source. Based on the complete genome sequence, proteins for nitrate utilization, nitrate transporter (NRT) and nitrate reductase (NR), were predicted to be encoded by the neighboring nuclear genes CMG018C and CMG019C, respectively, but no typical nitrite reductase (NiR) gene was found by similarity searches. On the other hand, two candidate genes for sulfite reductase (SiR) were found, one of which (CMG021C) is located next to the above-noted nitrate-related genes. Given that transcripts of CMG018C, CMG019C and CMG021C accumulate in nitrate-containing media, but are repressed by ammonium, and that SiR and NiR are structurally related enzymes, we hypothesized that the CMG021C gene product functions as an NiR in C. merolae. To test this hypothesis, we developed a method for targeted gene disruption in C. merolae. In support of our hypothesis, we found that a CMG021G null mutant in comparison with the parental strain showed decreased cell growth in nitrate-containing but not in ammonium-containing media. Furthermore, expression of CMG021C in the nirA mutant of a cyanobacterium, Leptolyngbya boryana (formerly Plectonema boryanum), could genetically complement the NiR defect. Immunofluorescent analysis indicated the localization of CMG021C in chloroplasts, and hence we propose an overall scheme for nitrate assimilation in C. merolae.

High-efficiency homologous recombination in the oil-producing alga Nannochloropsis sp.

“…One well-established example is the moss Physcomitrella patens, in which the use of HR has helped to establish this organism as a powerful model organism for studying the evolution of land plants (43). The only eukaryotic alga previously demonstrated to exhibit efficient HR is the red alga Cyanidioschyzon merolae (44,45); however, this organism is a rather divergent red alga and an extremophile that is not generally viewed as a suitable model species for investigating biofuel production by algae. In contrast, Nannochloropsis sp.…”

Translation-independent circadian control of the cell cycle in a unicellular photosynthetic eukaryote

“…The RBR genomic region with the URA insert was cut out from the vector by NotI and was transformed into C. merolae M4, a derivative of C. merolae 10D, which has a mutation in the URA gene 10 . Transformation and selection of the gene knockouts were performed as described 33 .…”

Phototaxis in the Unicellular Red Algae Cyanidioschyzon merolae and Cyanidium caldarium

“…It is possible therefore that C. merolae might use a non-conventional system for movement. Elucidating the molecular mechanism of phototaxis and gravitaxis in C. merolae using genomic information (Ohta et al 1998, Matsuzaki et al 2004, Nozaki et al 2007) and genetic techniques , Ohnuma et al 2008, 2009, Imamura et al 2009 would inform future studies of the phototaxic and gravitaxtic mechanism in primitive algae.…”