Loss of the TOR kinase Tor2 mimics nitrogen starvation and activates the sexual development pathway in fission yeast

Fission yeast has two TOR (target of rapamycin) kinases, namely Tor1 and Tor2. Tor1 is required for survival under stressed conditions, proper G(1) arrest, and sexual development. In contrast, Tor2 is essential for growth. To analyze the functions of Tor2, we constructed two temperature-sensitive tor2 mutants. Interestingly, at the restrictive temperature, these mutants mimicked nitrogen starvation by arresting the cell cycle in G(1) phase and initiating sexual development. Microarray analysis indicated that expression of nitrogen starvation-responsive genes was induced extensively when Tor2 function was suppressed, suggesting that Tor2 normally mediates a signal from the nitrogen source. As with mammalian and budding yeast TOR, we find that fission yeast TOR also forms multiprotein complexes analogous to TORC1 and TORC2. The raptor homologue, Mip1, likely forms a complex predominantly with Tor2, producing TORC1. The rictor/Avo3 homologue, Ste20, and the Avo1 homologue, Sin1, appear to form TORC2 mainly with Tor1 but may also bind Tor2. The Lst8 homologue, Wat1, binds to both Tor1 and Tor2. Our analysis shows, with respect to promotion of G(1) arrest and sexual development, that the loss of Tor1 (TORC2) and the loss of Tor2 (TORC1) exhibit opposite effects. This highlights an intriguing functional relationship among TOR kinase complexes in the fission yeast Schizosaccharomyces pombe.     

Evolutionarily Conserved Interaction between the Phosphoproteins and X Proteins of Bornaviruses from Different Vertebrate Species

Bornavirus, a non-segmented, negative-strand RNA viruses, is currently classified into several genetically distinct genotypes, such as Borna disease virus (BDV) and avian bornaviruses (ABVs). Recent studies revealed that bornavirus genotypes show unique sequence variability in the putative 5′ untranslated region (5′ UTR) of X/P mRNA, a bicistronic mRNA for the X protein and phosphoprotein (P). In this study, to understand the evolutionary relationship among the bornavirus genotypes, we investigated the functional interaction between the X and P proteins of four bornavirus genotypes, BDV, ABV genotype 4 and 5 and reptile bornavirus (RBV), the putative 5′ UTRs of which exhibit variation in the length. Immunofluorescence and immunoprecipitation analyses using mammalian and avian cell lines revealed that the X proteins of bornaviruses conserve the ability to facilitate the export of P from the nucleus to the cytoplasm via interaction with P. Furthermore, we showed that inter-genotypic interactions may occur between X and P among the genotypes, except for X of RBV. In addition, a BDV minireplicon assay demonstrated that the X and P proteins of ABVs, but not RBV, can affect the polymerase activity of BDV. This study demonstrates that bornaviruses may have conserved the fundamental function of a regulatory protein during their evolution, whereas RBV has evolved distinctly from the other bornavirus genotypes.     

Characterization of the cells in the repair tissue of full-thickness articular cartilage defects

 It is well established that a full-thickness articular cartilage defect is repaired with a fibrocartilaginous tissue, cells of which are derived from undifferentiated mesenchymal stem cells in the bone marrow. To characterize the repair cells biochemically, full-thickness defects were created in rabbit knee joints and the repair tissues taken at 3, 6, and 12 weeks after surgery. The repair cells were cultured and examined biochemically to investigate the effects of four exogenous growth factors with regard to the metabolism of type II collagen and proteoglycans. A significant increase of carboxy-terminal type II procollagen peptide production was observed in the conditional medium of the repair cells, especially taken at 6 weeks after surgery, in the presence of each growth factor. Glycosaminoglycan content was also increased and proteoglycan synthesis stimulated. The repair cells taken at the early stage of the repair process could originally have more activity of type II collagen synthesis, and the growth factors used could enhance the differentiation of the repair cells in vitro. Accepted: 3 November 1997     

Diurnal changes in the vertical distribution of phytoplankton in hypertrophic Lake Kasumigaura,Japan

The daily vertical migration of five species;Microcystis aeruginosa (Kütz.) Trevis,Anabaena spiroides Klebahn f.crassa (L.) Elenkin,Aphanizomenon flos-aquae (L.) Ralfs,Melosira granulata (E). Ralfs, andCoscinodiscus lacustris Grun. was studied using a close-interval water sampler on a calm summer day in Lake Kasumigaura. Many colonies ofMicrocystis were observed at the middle of the water column (approx. 1.5 m depth) in the afternoon, and at the surface in the early morning.Anabaena occurred mostly in the upper layer whileAphanizomenon tended to be uniformly distributed. The difference in migration patterns suggests thatMicrocystis is superior toAnabaena andAphanizomenon in obtaining both light and nutrients from this lake. Among diatoms,Melosira remained at the bottom of the water column throughout day and night, but Coscinodiscus was uniformly distributed.     

Early candidacy for differentiation into heterocysts in the filamentous cyanobacterium Anabaena sp. PCC 7120

The filamentous cyanobacterium Anabaena sp. PCC 7120 fixes dinitrogen facultatively. Upon depletion of combined nitrogen, about 10% of vegetative cells within the filaments differentiate terminally into nitrogen-fixing cells. The heterocyst has been studied as a model system of prokaryotic cell differentiation, with major focus on signal transduction and pattern formation. The fate of heterocyst differentiation is determined at about the eighth hour of induction (point of no return), well before conspicuous morphological or metabolic changes occur. However, little is known about how the initial heterocysts are selected after the induction by nitrogen deprivation. To address this question, we followed the fate of every cells on agar plates after nitrogen deprivation with an interval of 4 h. About 10% of heterocysts were formed without prior division after the start of nitrogen deprivation. The intensity of fluorescence of GFP in the transformants of hetR-gfp increased markedly in the future heterocysts at the fourth hour with respect to other cells. We also noted that the growing filaments consisted of clusters of four consecutive cells that we call quartets. About 75% of initial heterocysts originated from either of the two outer cells of quartets at the start of nitrogen deprivation. These results suggest that the future heterocysts are loosely selected at early times after the start of nitrogen deprivation, before the commitment. Such early candidacy could be explained by different properties of the outer and inner cells of a quartet, but the molecular nature of candidacy remains to be uncovered.     

Influence of cadmium on the distribution of the essential trace elements zinc and copper in the liver and kidneys of rats

Cd induced changes of Zn and Cd distribution in the liver and kidneys were studied in relation to Cd metallothionein (MT) synthesis. Wistar male rats were given CdCl₂ by sc injection of .8, 1.5, and 3.0 mg Cd/kg three times a week for three weeks. Cd levels of liver and kidneys increased with the increment of Cd dosage and 80–90% of Cd was found in the cytosol. The MT fractions contained 80–89% cytosolic Cd in the liver and 55–75% Cd in the kidneys. Zn concentrations in the liver increased following Cd administration, But Zn in the kidneys showed only slight increase. There was a distinct decrease of Cu concentration in the liver of the 3.0 mg group. In contrast, Cu concentrations in the kidneys increased about three times in the .8 and 1.5 mg Cd groups, but Cu in the 3.0 mg group showed only 1.5 times increase. The changes of these metal concentrations were observed mainly in the cytosol. Non-MT-Cd in the kidneys was maximum in the 1.5 mg group, but the 3.0 mg group showed significant decrease. In parallel with this decrease of Cd, Cu and Zn in the kidneys showed similar decrease. When the kidneys are injured, Zn and Cu appear to leak from this organ.     

The diversity of mycorrhizal fungi in Japanese Cephalanthera species

The diversity of mycorrhizal fungi among four Japanese Cephalanthera (Orchidaceae) species was examined at a total of seven sites, based on sequence variation in nuclear ribosomal DNA. This is the first report to detect mycorrhizal fungi in C. subaphylla. Two patterns of mycorrhizal associations were confirmed from our results. C. falcata has lower fungal specificity, associating with Russulaceae, Sebacinales and Thelephoraceae. By contrast, the three Cephalanthera species C. erecta, C. subaphylla, and C. longifolia were associated mainly with Thelephoraceae fungi. There is no large difference found in mycobionts between green and albino individuals of C. falcata and no distinct preference of Japanese Cephalanthera species for a specific fungal group of Thelephoraceae.