CZON-cutter: a CRISPR-Cas9 system for multiplexed organelle imaging in a simple unicellular alga

The unicellular alga Cyanidioschyzon merolae has a simple cellular structure: each cell has one nucleus, one mitochondrion, one chloroplast, and one peroxisome. This simplicity offers unique advantages for investigating organellar proliferation and the cell cycle. Here, we describe CZON-cutter, an engineered clustered, regularly interspaced, short palindromic repeats (CRISPR)/CRISPR-associated nuclease 9 (Cas9) system for simultaneous genome editing and organellar visualization. We engineered a C. merolae strain expressing a nuclear-localized Cas9-Venus nuclease for targeted editing of any locus defined by a single guide RNA (sgRNA). We then successfully edited the algal genome and visualized the mitochondrion and peroxisome in transformants using fluorescent protein reporters with different excitation wavelengths. Fluorescent protein labeling of organelles in living transformants allows us to validate phenotypes associated with organellar proliferation and the cell cycle, even when the edited gene is essential. Combined with the exceptional biological features of C. merolae, CZON-cutter will be instrumental for investigating cellular and organellar division in a high-throughput manner.

CZON-cutter: a CRISPR-Cas9 system with multiplexed organelle imaging in a simple unicellular alga

The simple cellular structure of the unicellular alga Cyanidioschyzon merolae consists of one nucleus, one mitochondrion, one chloroplast, and one peroxisome per cell and offers unique advantages to investigate mechanisms of organellar proliferation and the cell cycle. Here, we describe an engineered clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated protein 9 (Cas9) system, CZON-cutter, for simultaneous genome editing and organellar visualization. We engineered a C. merolae strain expressing a nuclear-localized Cas9-Venus nuclease to target editing at a locus defined by a single-guide RNA (sgRNA). We then successfully edited the algal genome and visualized the mitochondrion and peroxisome in transformants by fluorescent protein reporters with different excitation wavelengths. Fluorescent protein labeling of organelles in living transformants allows validation of phenotypes associated with organellar proliferation and the cell cycle, even when the edited gene is essential. Combined with the exceptional biological features of C. merolae, CZON-cutter will be instrumental for investigating cellular and organellar division in a high-throughput manner.

PhycoCosm, a comparative algal genomics resource

Algae are a diverse, polyphyletic group of photosynthetic eukaryotes spanning nearly all eukaryotic lineages of life and collectively responsible for ∼50% of photosynthesis on Earth. Sequenced algal genomes, critical to understanding their complex biology, are growing in number and require efficient tools for analysis. PhycoCosm (https://phycocosm.jgi.doe.gov) is an algal multi-omics portal, developed by the US Department of Energy Joint Genome Institute to support analysis and distribution of algal genome sequences and other ‘omics’ data. PhycoCosm provides integration of genome sequence and annotation for >100 algal genomes with available multi-omics data and interactive web-based tools to enable algal research in bioenergy and the environment, encouraging community engagement and data exchange, and fostering new sequencing projects that will further these research goals.

Acidophilic green algal genome provides insights into adaptation to an acidic environment

Some microalgae are adapted to extremely acidic environments in which toxic metals are present at high levels. However, little is known about how acidophilic algae evolved from their respective neutrophilic ancestors by adapting to particular acidic environments. To gain insights into this issue, we determined the draft genome sequence of the acidophilic green alga Chlamydomonas eustigma and performed comparative genome and transcriptome analyses between C. eustigma and its neutrophilic relative Chlamydomonas reinhardtii. The results revealed the following features in C. eustigma that probably contributed to the adaptation to an acidic environment. Genes encoding heat-shock proteins and plasma membrane H+-ATPase are highly expressed in C. eustigma. This species has also lost fermentation pathways that acidify the cytosol and has acquired an energy shuttle and buffering system and arsenic detoxification genes through horizontal gene transfer. Moreover, the arsenic detoxification genes have been multiplied in the genome. These features have also been found in other acidophilic green and red algae, suggesting the existence of common mechanisms in the adaptation to acidic environments.

Photosynthesis and the role of plastids (kleptoplastids) in Sacoglossa (Heterobranchia, Gastropoda): a short review

Title (Bahasa Indonesia): Fotosintesis dan peran plastida [kleptoplastids] pada Sacoglossa [Heterobranchia, Gastropoda]: tinjauan singkat. In this manuscript I will give a short summary of our knowledge on photosyn-thesis in the enigmatic gastropod group Sacoglossa. Members of this group are able to sequester chloroplasts from their food algae (mainly Chlorophyta) and store them for weeks and months and it was assumed for a long time that they can use chloroplasts in a similar way as plants do. Only few sacoglossan species are able to perform photosynthesis for months, others are less effective or are not able at all. The processes involved are investigated now for a few years, but are still not clear. However we know now that many factors contribute to this enigmatic biological system. These include extrinsic (environment, origin and properties of the nutrition and the plastids) and intrinsic factors of slugs and algae (behaviour, physiological and anatomical properties). Plastids are not maintained by genes that might have originated by a horizontal gene transfer (HGT) from the algal genome into the slug genome, as was hypothesized for many years. We therefore have to focus our research now on other factors to understand what actually contributes to this unique metazoan phenomenon which is not yet understood. In this review, some of these new approaches are summarized. Dalam tulisan ini saya akan memberikan ringkasan singkat tentang fotosintesis pada gastropoda kelompok misterius Sacoglossa. Organisme anggota dari kelompok ini mampu menyerap kloroplas dari alga makanan mereka (terutama Chlorophyta) dan menyimpannya selama berminggu-minggu, bahkan berbulan-bulan, sehingga telah diasumsikan bahwa mereka dapat menggunakan kloroplas dengan cara yang sama seperti tanaman. Hanya sedikit spesies sacoglossan dapat melakukan fotosintesis selama berbulan-bulan, yang lain kurang efektif atau tidak mampu sama sekali. Proses yang terlibat diselidiki sekarang selama beberapa tahun, namun masih belum jelas. Namun kita tahu sekarang bahwa banyak faktor yang berkontribusi terhadap sistem biologis misterius ini. Ini termasuk ekstrinsik (lingkungan, asal dan sifat gizi dan plastida) dan faktor intrinsik siput dan ganggang (perilaku, fisiologis dan sifat anatomis). Plastida tidak dikelola oleh gen yang mungkin berasal oleh transfer gen horizontal (HGT) dari genom alga ke dalam genom slug, seperti yang dihipotesiskan selama bertahun-tahun. Oleh karena itu kita harus fokus penelitian kami sekarang pada faktor-faktor lain untuk memahami apa yang sebenarnya memberikan kontribusi terhadap fenomena ini metazoan unik yang belum dipahami. Dalam ulasan ini, beberapa pendekatan baru dirangkum.