A 100%-complete sequence reveals unusually simple genomic features in the hot-spring red alga Cyanidioschyzon merolae

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116

Plant cells sense environmental nitrogen levels and alter their gene expression accordingly to survive; however, the underlying regulatory mechanisms still remains to be elucidated. Here, we identified and characterized a transcription factor that is responsible for expression of nitrogen assimilation genes in a unicellular red alga Cyanidioschyzon merolae . DNA microarray and Northern blot analyses revealed that transcript of the gene encoding CmMYB1, an R2R3-type MYB transcription factor, increased 1 h after nitrogen depletion. The CmMYB1 protein started to accumulate after 2 h and reached a peak after 4 h after nitrogen depletion, correlating with the expression of key nitrogen assimilation genes, such as CmNRT , CmNAR , CmNIR , CmAMT , and CmGS . Although the transcripts of these nitrogen assimilation genes were detected in nitrate-grown cells, they disappeared upon the addition of preferred nitrogen source such as ammonium or glutamine, suggesting the presence of a nitrogen catabolite repression (NCR) mechanism. The nitrogen depletion-induced gene expression disappeared in a CmMYB1 -null mutant, and the mutant showed decreased cell viability after exposure to the nitrogen-depleted conditions compared with the parental strain. Chromatin immunoprecipitation analysis demonstrated that CmMYB1 specifically occupied these nitrogen-responsive promoter regions only under nitrogen-depleted conditions, and electrophoretic mobility shift assays using crude cell extract revealed specific binding of CmMYB1, or a complex containing CmMYB1, to these promoters. Thus, the presented results indicated that CmMYB1 is a central nitrogen regulator in C. merolae .

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confidence: 99%

R2R3-type MYB transcription factor, CmMYB1, is a central nitrogen assimilation regulator in Cyanidioschyzon merolae

Imamura

Kanesaki

Ohnuma

et al. 2009

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116

“…After completion of both ODR and NDR, division of the plastid, mitochondrion, and nucleus occurs sequentially and is followed by cell division (7). The complete nucleotide sequences of the 3 genomes of C. merolae have been determined (8)(9)(10)(11), with the result that many tools are now available or are under development for genomics analysis. In this study, we have identified the chemical signal that coordinates ODR and NDR in C. merolae and have shown that this mechanism is also working in flowering plants.…”

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confidence: 99%

Tetrapyrrole signal as a cell-cycle coordinator from organelle to nuclear DNA replication in plant cells

Kobayashi

Kanesaki

Tanaka

et al. 2009

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102

Eukaryotic cells arose from an ancient endosymbiotic association of prokaryotes, with plant cells harboring 3 genomes as the remnants of such evolution. In plant cells, plastid and mitochondrial DNA replication [organelle DNA replication (ODR)] occurs in advance of the subsequent cell cycles composed of nuclear DNA replication (NDR) and cell division. However, the mechanism by which replication of these genomes with different origins is coordinated is largely unknown. Here, we show that NDR is regulated by a tetrapyrrole signal in plant cells, which has been suggested as an organelle-to-nucleus retrograde signal. In synchronized cultures of the primitive red alga Cyanidioschyzon merolae, specific inhibition of A-type cyclin-dependent kinase (CDKA) prevented NDR but not ODR after onset of the cell cycle. In contrast, inhibition of ODR by nalidixic acid also resulted in inhibition of NDR, indicating a strict dependence of NDR on ODR. The requirement of ODR for NDR was bypassed by addition of the tetrapyrrole intermediates protoporphyrin IX (ProtoIX) or Mg-ProtoIX, both of which activated CDKA without inducing ODR. This scheme was also observed in cultured tobacco cells (BY-2), where inhibition of ODR by nalidixic acid prevented CDKA activation and NDR, and these inhibitions were circumvented by Mg-ProtoIX without inducing ODR. We thus show that tetrapyrrole-mediated organelle-nucleus replicational coupling is an evolutionary conserved process among plant cells.Cyanidioschyzon merolae ͉ retrograde signal ͉ tobacco BY-2 T he origin of eukaryotic cells is still unclear, but it is now generally accepted that mitochondria and plastids arose from endosymbiosis of rickettsia-like ␣-proteobacteria and cyanobacteria-like photosynthetic bacteria, respectively (1). Many of the genes of these endosymbionts, including those for DNA replication and maintenance of genomic integrity, were subsequently lost or transferred to the nuclear genome [endosymbiotic gene transfer (EGT)]. According to current dogma, the transfer of genes of endosymbiotic origin to the nucleus that are required for organelle DNA replication (ODR) has resulted in loss of the independence of the cell cycles of the endosymbionts and in their integration into the eukaryotic control system that is mediated largely by cyclins and cyclin-dependent kinase (CDK). However, coordination of cell-cycle events such as DNA replication would have been essential for establishing integrity of eukaryotic cells, at least during the early stages of endosymbiotic association. Although there is little evidence for discrete cell-cycle control of ODR in animal and fungal cells, studies with algae and flowering plants have shown that ODR precedes the subsequent cell proliferation cycles composed of nuclear DNA replication (NDR) and cell division (2-4).C. merolae is an unicellular red alga, each cell of which contains 1 plastid, 1 mitochondrion, and 1 nucleus. The advantage of studying nuclear and organelle proliferation cycles in C. merolae is that the cultures can be highly synchro...

“…S2F). Dnm1 is one of only two dynamins (Dnm1 and Dnm2) encoded in the C. merolae genome (7,11). Dnm1 and Dnm2 are important components of the mitochondrion-dividing (MD) machinery (9,14,15) and plastiddividing (PD) machinery (16,17), respectively.…”

Section: Resultsmentioning

confidence: 99%

“…Samples were analyzed by a peptide mass fingerprinting search using a MALDI-TOF-MS (AXIMA-TOF 2 ; Shimadzu), based on the FASTA file distributed by the C. merolae Genome Project (7,11).…”

Section: Methodsmentioning

confidence: 99%

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Single-membrane–bounded peroxisome division revealed by isolation of dynamin-based machinery

Imoto

Kuroiwa

Yoshida

et al. 2013

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Peroxisomes (microbodies) are ubiquitous single-membrane-bounded organelles and fulfill essential roles in the cellular metabolism. They are found in virtually all eukaryotic cells and basically multiply by division. However, the mechanochemical machinery involved in peroxisome division remains elusive. Here, we first identified the peroxisome-dividing (POD) machinery. We isolated the POD machinery from Cyanidioschyzon merolae, a unicellular red alga containing a single peroxisome. Peroxisomal division in C. merolae can be highly synchronized by light/dark cycles and the microtubuledisrupting agent oryzalin. By proteomic analysis based on the complete genome sequence of C. merolae, we identified a dynamin-related protein 3 (DRP3) ortholog, CmDnm1 (Dnm1), that predominantly accumulated with catalase in the dividing-peroxisome fraction. Immunofluorescence microscopy demonstrated that Dnm1 formed a ring at the division site of the peroxisome. The outlines of the isolated dynamin rings were dimly observed by phasecontrast microscopy and clearly stained for Dnm1. Electron microscopy revealed that the POD machinery was formed at the cytoplasmic side of the equator. Immunoelectron microscopy showed that the POD machinery consisted of an outer dynamin-based ring and an inner filamentous ring. Down-regulation of Dnm1 impaired peroxisomal division. Surprisingly, the same Dnm1 serially controlled peroxisomal division after mitochondrial division. Because genetic deficiencies of Dnm1 orthologs in multiperoxisomal organisms inhibited both mitochondrial and peroxisomal proliferation, it is thought that peroxisomal division by contraction of a dynaminbased machinery is universal among eukaryotes. These findings are useful for understanding the fundamental systems in eukaryotic cells.