The fission yeast Schizosaccharomyces pombe is an excellent model organism to explore cellular events owing to rich tools in genetics, molecular biology, cellular biology, and biochemistry. ...Schizosaccharomyces pombe proliferates continuously when nutrients are abundant but arrests in G1 phase upon depletion of nutrients such as nitrogen and glucose. When cells of opposite mating types are present, cells conjugate, fuse, undergo meiosis, and finally form 4 spores. This sexual differentiation process in S. pombe has been studied extensively. To execute sexual differentiation, the glucose-sensing cAMP-PKA (cyclic adenosine monophosphate-protein kinase A) pathway, nitrogen-sensing TOR (target of rapamycin) pathway, and SAPK (stress-activating protein kinase) pathway are crucial, and the MAPK (mitogen-activating protein kinase) cascade is essential for pheromone sensing. These signals regulate ste11 at the transcriptional and translational levels, and Ste11 is modified in multiple ways. This review summarizes the initiation of sexual differentiation in S. pombe based on results I have helped to obtain, including the work of many excellent researchers.
Chitosanases belong to glycoside hydrolase families 5, 7, 8, 46, 75 and 80 and hydrolyse glucosamine polymers produced by partial or full deacetylation of chitin. Herein, we determined the crystal ...structure of chitosanase from the β‐proteobacterium, Mitsuaria chitosanitabida, (McChoA) at 1.75 Å resolution; the first structure of a family 80 chitosanase. McChoA is a 34 kDa extracellular protein of 301 amino acids that fold into two (upper and lower) globular domains with an active site cleft between them. Key substrate‐binding features are conserved with family 24 lysozymes and family 46 chitosanases. The distance between catalytic residues E41 and E61 (10.8 Å) indicates an inverting type mechanism. Uniquely, three disulphide bridges and the C terminus might contribute to enzyme activity.
The fission yeast Schizosaccharomyces pombe secretes the extracellular maltase Agl1, which hydrolyzes maltose into glucose, thereby utilizing maltose as a carbon source. Whether other maltases ...contribute to efficient utilization of maltose and how Agl1 expression is regulated in response to switching of carbon sources are unknown. In this study, we show that three other possible maltases and the maltose transporter Sut1 are not required for efficient utilization of maltose. Transcription of agl1 was induced when the carbon source was changed from glucose to maltose. This was dependent on Atf1 and Pcr1, which are highly conserved transcription factors that regulate stress-responsive genes in various stress conditions. Atf1 and Pcr1 generally bind the TGACGT motif as a heterodimer. The agl1 gene lacks the exact motif, but has many degenerate TGACGT motifs in its promoter and coding region. When the carbon source was switched from glucose to maltose, Atf1 and Pcr1 associated with the promoters and coding regions of agl1, fbp1, and gpx1, indicating that the Atf1-Pcr1 heteromer binds a variety of regions in its target genes to induce their transcription. In addition, the association of Mediator with these genes was dependent on Atf1 and Pcr1. These data indicate that Atf1 and Pcr1 induce the transcription of agl1, which allows efficient utilization of extracellular maltose.
Fungal cell walls are medically important since they represent a drug target site for antifungal medication. So far there is no method to directly visualize structurally similar cell wall components ...such as α-glucan, β-glucan and mannan with high specificity, especially in a label-free manner. In this study, we have developed a Raman spectroscopy based molecular imaging method and combined multivariate curve resolution analysis to enable detection and visualization of multiple polysaccharide components simultaneously at the single cell level. Our results show that vegetative cell and ascus walls are made up of both α- and β-glucans while spore wall is exclusively made of α-glucan. Co-localization studies reveal the absence of mannans in ascus wall but are distributed primarily in spores. Such detailed picture is believed to further enhance our understanding of the dynamic spore wall architecture, eventually leading to advancements in drug discovery and development in the near future.
Anterograde vesicle transport from the endoplasmic reticulum to the Golgi apparatus is the start of protein transport through the secretory pathway, in which the transport is mediated by coat protein ...complex II (COPII)-coated vesicles. Therefore, most proteins synthesized on the endoplasmic reticulum are loaded as cargo into COPII vesicles. The COPII is composed of the small GTPase Sar1 and two types of protein complexes (Sec23/24 and Sec 13/31). Of these five COPII components, Sec24 is thought to recognize cargo that is incorporated into COPII vesicles by directly interacting with the cargo. The Arabidopsis genome encodes three types of Sec24 homologs (AtSec24A, AtSec24B, and AtSec24C). The subcellular dynamics and function of AtSec24A have been characterized. The intracellular distributions and functions of other AtSec24 proteins are not known, and the functional differences among the three AtSec24s remain unclear. Here, we found that all three AtSec24s were expressed in similar parts of the plant body and showed the same subcellular localization pattern. AtSec24B knockout plant, but not AtSec24C knockdown plant, showed mild male sterility with reduction of pollen germination. Significant decrease of AtSec24B and AtSec24C expression affected male and female gametogenesis in Arabidopsis thaliana. Our results suggested that the redundant function of AtSec24B and AtSec24C is crucial for the development of plant reproductive cells. We propose that the COPII transport is involved in male and female gametogenesis in planta.
Prenyldiphosphate synthases catalyze the reaction of allylic diphosphates with one or more isopentenyl diphosphate molecules to form compounds such as farnesyl diphosphate, used in, e.g., sterol ...biosynthesis and protein prenylation, as well as longer "polyprenyl" diphosphates, used in ubiquinone and menaquinone biosynthesis. Quinones play an essential role in electron transport and are associated with the inner mitochondrial membrane due to the presence of the polyprenyl group. In this work, we investigated the synthesis of the polyprenyl diphosphate that alkylates the ubiquinone ring precursor in Toxoplasma gondii, an opportunistic pathogen that causes serious disease in immunocompromised patients and the unborn fetus. The enzyme that catalyzes this early step of the ubiquinone synthesis is Coq1 (TgCoq1), and we show that it produces the C35 species heptaprenyl diphosphate. TgCoq1 localizes to the mitochondrion and is essential for
T. gondii growth. We demonstrate that the growth defect of a T. gondii TgCoq1 mutant is rescued by complementation with a homologous
gene or with a (C45) solanesyl diphosphate synthase from Trypanosoma cruzi (TcSPPS). We find that a lipophilic bisphosphonate (BPH-1218) inhibits T. gondii growth at low-nanomolar concentrations, while overexpression of the TgCoq1 enzyme dramatically reduced growth inhibition by the bisphosphonate. Both the severe growth defect of the mutant and the inhibition by BPH-1218 were rescued by supplementation with a long-chain (C30) ubiquinone (UQ
). Importantly, BPH-1218 also protected mice against a lethal T. gondii infection. TgCoq1 thus represents a potential drug target that could be exploited for improved chemotherapy of toxoplasmosis.
Millions of people are infected with Toxoplasma gondii, and the available treatment for toxoplasmosis is not ideal. Most of the drugs currently used are only effective for the acute infection, and treatment can trigger serious side effects requiring changes in the therapeutic approach. There is, therefore, a compelling need for safe and effective treatments for toxoplasmosis. In this work, we characterize an enzyme of the mitochondrion of T. gondii that can be inhibited by an isoprenoid pathway inhibitor. We present evidence that demonstrates that inhibition of the enzyme is linked to parasite death. In addition, the inhibitor can protect mice against a lethal dose of T. gondii. Our results thus reveal a promising chemotherapeutic target for the development of new medicines for toxoplasmosis.
A considerable portion of agricultural land in central‐east Japan has been contaminated by radioactive material, particularly radioactive Cs, due to the industrial accident at the Fukushima Daiichi ...nuclear power plant. Understanding the mechanism of absorption, translocation and accumulation of Cs+ in plants will greatly assist in developing approaches to help reduce the radioactive contamination of agricultural products. At present, however, little is known regarding the Cs+ transporters in rice. A transporter‐enriched yeast expression library was constructed and the library was screened for Cs+ transporter genes. The 1452 full length cDNAs encoding transporter genes were obtained from the Rice Genome Resource Center and 1358 clones of these transporter genes were successively subcloned into yeast expression vectors; which were then transferred into yeast. Using this library, both positive and negative selection screens can be performed, which have not been previously possible. The constructed library is an excellent tool for the isolation of novel transporter genes. This library was screened for clones that were sensitive to Cs+ using a SD‐Gal medium containing either 30 or 70 mM CsCl; resulting in the isolation of 13 Cs+ sensitive clones. 137Cs absorption experiments were conducted and confirmed that all of the identified clones were able to absorb 137Cs. A total of 3 potassium transporters, 2 ABC transporters and 1 NRAMP transporter were among the 13 identified clones.
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