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Yo Suzuki

Publications and source records attributed to Yo Suzuki.

8 recordsLinked to original sources
PubMed2025

Robust and highly efficient transformation method for a minimal mycoplasma cell.

UNLABELLED: Mycoplasmas have been widely investigated for their pathogenicity, as well as for genomics and synthetic biology. Conventionally, transformation of mycoplasmas was not highly efficient, and due to the low transformation efficiency, large amounts of DNA and recipient cells were required for that purpose. Here, we report a robust and highly efficient transformation method for the minimal cell JCVI-syn3B, which was created through streamlining the genome of Mycoplasma mycoides. When the growth states of JCVI-syn3B were examined in detail by focusing on such factors as pH, color, absorbance, colony forming unit, and transformation efficiency, it was found that the growth phase after the lag phase can be divided into three distinct phases, of which the highest transformation efficiency was observed during the early exponential growth phase. Notably, the transformation efficiency of up to 4.4 × 10-2 transformants per cell per microgram of plasmid DNA was obtained. A method to obtain several hundred to several thousand transformants with less than 0.2 mL of culture with approximately 1 × 107-108 cells and 10 ng of plasmid DNA was developed. Moreover, a transformation method using a frozen stock of transformation-ready cells was established. These procedures and information could simplify and enhance the transformation process of minimal cells, facilitating advanced genetic engineering and biological research using minimal cells. IMPORTANCE: Mycoplasmas are parasitic and pathogenic bacteria for many animals. They are also useful bacteria to understand the cellular process of life and for bioengineering because of their simple metabolism, small genomes, and cultivability. Genetic manipulation is crucial for these purposes, but transformation efficiency in mycoplasmas is typically quite low. Here, we report a highly efficient transformation method for the minimal genome mycoplasma JCVI-syn3B. Using this method, transformants can be obtained with only 10 ng of plasmid DNA, which is around one-thousandth of the amount required for traditional mycoplasma transformations. Moreover, a convenient method using frozen stocks of transformation-ready cells was established. These improved methods play a crucial role in further studies using minimal cells.

Transformation, Bacterial↗
PubMed2016

Design and synthesis of a minimal bacterial genome.

We used whole-genome design and complete chemical synthesis to minimize the 1079-kilobase pair synthetic genome of Mycoplasma mycoides JCVI-syn1.0. An initial design, based on collective knowledge of molecular biology combined with limited transposon mutagenesis data, failed to produce a viable cell. Improved transposon mutagenesis methods revealed a class of quasi-essential genes that are needed for robust growth, explaining the failure of our initial design. Three cycles of design, synthesis, and testing, with retention of quasi-essential genes, produced JCVI-syn3.0 (531 kilobase pairs, 473 genes), which has a genome smaller than that of any autonomously replicating cell found in nature. JCVI-syn3.0 retains almost all genes involved in the synthesis and processing of macromolecules. Unexpectedly, it also contains 149 genes with unknown biological functions. JCVI-syn3.0 is a versatile platform for investigating the core functions of life and for exploring whole-genome design.

Artificial Cells↗
PubMed2006

Genetic redundancy masks diverse functions of the tumor suppressor gene PTEN during C. elegans development.

Genetic redundancy is associated with a large percentage of genes. We investigated PTEN (phosphatase and tensin homolog deleted on chromosome 10) tumor suppressor gene functions that eluded single mutant analyses, using a Caenorhabditis elegans genome-wide screen. We show that at least 27 genes collaborate with the worm PTEN homolog daf-18 for various functions previously concealed by genetic redundancy, including embryogenesis, cuticle turnover, egg laying, and oocyte maturation. In one example, daf-18 appears to constitute a cell-autonomous germline signal that converges with a somatic gonad signal mediated by ceh-18 at a kinase inhibition. We provide evidence that daf-18 elicits some functions independent of the downstream gene daf-16.

Animals↗
PubMed2006

Dietary 5-campestenone (campest-5-en-3-one) enhances fatty acid oxidation in perfused rat liver.

The effect of dietary 5-campestenone (campest-5-en-3-one), a chemical modification product of a naturally-occurring plant sterol, campesterol, on lipid metabolism was examined using a rat liver perfusion system. Male Sprague-Dawley rats weighing about 140 g were fed a diet supplemented with or without 0.2% 5-campestenone for 14 d. 5-Campestenone feeding resulted in a marked reduction in the concentrations of serum lipids, such as triacylglycerol (TG), cholesterol, phospholipid, and free fatty acid, without influencing food intake or growth. Then, isolated livers from both groups were perfused for 4 h in the presence of an exogenous linoelaidic acid substrate. Dietary 5-campestenone markedly elevated hepatic ketone body production, while cumulative secretions of TG, cholesterol, and phospholipid by the livers of rats fed 5-campestenone were all significantly lowered as compared to those fed without the compound: the extent of the reduction was more prominent in the secretion of TG than other lipid components. In addition, the reduction of TG secretion was concomitantly accompanied by the reduced incorporation of both exogenous and endogenous fatty acids into this lipid molecule. These results suggest that dietary 5-campestenone exerts its hypotriglyceridemic effect, at least, in part through an enhanced metabolism of endogenous and exogenous fatty acids to oxidation at the expense of esterification in rat liver.

Animal Feed↗
PubMed2003

Expression of the C. elegans labial orthologue ceh-13 during male tail morphogenesis.

Hox genes are transcriptional regulators of metazoan body regionalization along the anteroposterior axis that act by specifying positional identity in differentiating cells. ceh-13, the labial orthologue in Caenorhabditis elegans, is expressed both during embryogenesis and post- embryonic development. Using GFP reporter analysis and immunocytochemistry, we discovered a spatiotemporal pattern of gene expression in the male tail during the L3 and L4 larval stages that is TGF-beta pathway-dependent. Analysis of reporter activity in transgenic animals identified a distinct promoter region driving male tail-specific ceh-13 expression. We also report the interspecies conservation of sequence motifs within this region and speculate that, in the course of evolutionary diversification, ceh-13 may have acquired new functionality while conserving its homeotic role.

Animals↗
PubMed2002

A Caenorhabditis elegans TGF-beta, DBL-1, controls the expression of LON-1, a PR-related protein, that regulates polyploidization and body length.

Using cDNA-based array analysis combined with double-stranded RNA interference (dsRNAi), we have identified yk298h6 as a target gene of Caenorhabditis elegans TGF-beta signaling. Worms overexpressing dbl-1, a TGF-beta ligand, are 16% longer than wild type. Array analysis shows yk298h6 to be one of several genes suppressed in such worms. Disruption of yk298h6 function by dsRNAi also resulted in long worms, suggesting that it is a negative regulator of body length. yk298h6 was then mapped to, and shown to be identical to, lon-1, a known gene that affects body length. lon-1 encodes a 312 amino acid protein with a motif sequence that is conserved from plants to humans. Expression studies confirm that LON-1 is repressed by DBL-1, suggesting that LON-1 is a novel downstream component of the C.elegans TGF-beta growth regulation pathway. Consistent with this, LON-1 is expressed mainly in the larval and adult hypodermis and has dose-dependent effects on body length associated with changes in hypodermal ploidy, but not hypodermal cell proliferation.

Alleles↗
PubMed2002

A cuticle collagen encoded by the lon-3 gene may be a target of TGF-beta signaling in determining Caenorhabditis elegans body shape.

The signaling pathway initiated by the TGF-beta family member DBL-1 in Caenorhabditis elegans controls body shape in a dose-dependent manner. Loss-of-function (lf) mutations in the dbl-1 gene cause a short, small body (Sma phenotype), whereas overexpression of dbl-1 causes a long body (Lon phenotype). To understand the cellular mechanisms underlying these phenotypes, we have isolated suppressors of the Sma phenotype resulting from a dbl-1(lf) mutation. Two of these suppressors are mutations in the lon-3 gene, of which four additional alleles are known. We show that lon-3 encodes a collagen that is a component of the C. elegans cuticle. Genetic and reporter-gene expression analyses suggest that lon-3 is involved in determination of body shape and is post-transcriptionally regulated by the dbl-1 pathway. These results support the possibility that TGF-beta signaling controls C. elegans body shape by regulating cuticle composition.

Animals↗