Identification of the bud emergence gene BEM4 and its interactions with rho-type GTPases in Saccharomyces cerevisiae.

The Rho-type GTPase Cdc42p is required for cell polarization and bud emergence in Saccharomyces cerevisiae. To identify genes whose functions are linked to CDC42, we screened for (i) multicopy suppressors of a Ts- cdc42 mutant, (ii) mutants that require multiple copies of CDC42 for survival, and (iii) mutations that display synthetic lethality with a partial-loss-of-function allele of CDC24, which encodes a guanine nucleotide exchange factor for Cdc42p. In all three screens, we identified a new gene, BEM4. Cells from which BEM4 was deleted were inviable at 37 degrees C. These cells became unbudded, large, and round, consistent with a model in which Bem4p acts together with Cdc42p in polarity establishment and bud emergence. In some strains, the ability of CDC42 to serve as a multicopy suppressor of the Ts- growth defect of deltabem4 cells required co-overexpression of Rho1p, which is an essential Rho-type GTPase necessary for cell wall integrity. This finding suggests that Bem4p also affects Rho1p function. Bem4p displayed two-hybrid interactions with Cdc42p, Rho1p, and two of the three other known yeast Rho-type GTPases, suggesting that Bem4p can interact with multiple Rho-type GTPases. Models for the role of Bem4p include that it serves as a chaperone or modulates the interaction of these GTPases with one or more of their targets or regulators.     

Sex allocation of three solitary ectoparasitic wasp species on bean weevil larvae: Sex ratio change with host quality and local mate competition

Sex ratio manipulation by ovipositing females was surveyed in 3 solitary ectoparastic wasp species,Dinarmus basalis (Pteromalidae),Anisopteromalus calanrae (Pteromalidae), andHeterospilus prosopidis (Braconidae), that parasitize azuki bean weevil (Callosobruchus chinensis (L) (Coleoptera: Buruchidae)) larvae within azuki beans (Vigna angularis). Variables were local mate competition (LMC) and host quality (HQ). We used host age as a measure of host quality (from 9-to 16-day-old hosts), changed the number of ovipositing females to control the level of local mate competition (1 female and 10 females), and examined oviposition patterns of the wasps. The offspring sex ratios (proportion of females) of the 3 wasp species respond qualitatively same to HQ and LMC. The common qualitative tendency among the 3 species is an increase of sex ratios increase with host age. In the process of changing the sex ratio (9–13-day-old) 3 wasp species respond only to HQ. In the hosts that end development in size (14–16-day-old) wasps respond to LMC. The response of sex ratio change to LMC in the old host ageclasses are different among the 3 species. In the situation that there exists LMC (10 females) sex ratios are the same among the 3 wasps. However, the sex ratios in no LMC (single female) are heterogeneous among the 3 wasps.     

Developmental expression of extracellular matrix components in intramuscular connective tissue of bovine semitendinosus muscle

 We have investigated the expression patterns of extracellular matrix components in intramuscular connective tissue during the development of bovine semitendinosus muscle by means of indirect immunofluorescence techniques. Types I, III, V, and VI collagen and fibronectin were located in the endomysium and the perimysium. Type IV collagen, laminin, and heparan sulfate proteoglycans (PGs) were exclusively located in the endomysium, and dermatan sulfate PGs existed only in the perimysium. The localization of these components in the intramuscular connective tissue of semitendinosus muscle remained unchanged throughout prenatal and postnatal growth of cattle, suggesting that they are essential for forming and maintaining structures of the endomysium and perimysium in bovine semitendinosus muscle. On the other hand, decorin was undetectable in the endomysium of neonates, although other matrix components were already expressed. It was expressed slightly in the endomysium of 2-month-old calves, and clearly detectable in the endomysium of cattle more than 6 months old. Chondroitin sulfate PGs were barely detectable in the perimysium of fetuses and neonatal calves, and progressively appeared during postnatal development of the muscle. It seems likely that these PGs are closely related to the postnatal development of the endomysium and perimysium. Accepted: 30 October 1996     

Biosynthesis and biological activity of β-(5-methylisoxazolin-3-on-2-yl)alanine in higher plants

Enzyme preparations from Leucaena seedlings catalysed the formation of β-(5-methylisoxazolin-3-on-2-yl)alanine (MIA) by using 3-hydroxy-5-methylisoxazole (HMI) and O-acetyl-L-serine. Some properties of this enzyme are described. The β-substituted alanine synthases from Pisum and Citrullus seedlings could not catalyse the synthesis of MIA. The phytotoxic effect of HMI on rice seedlings is reduced by alanylation.     

Distribution of body weight, blood insulin and lipid levels in the SMXA recombinant inbred strains and the QTL analysis.

In the SMXA recombinant inbred (RI) strains, we measured body weight, blood insulin and lipid (triglyceride, total cholesterol and phospholipid) levels in each strain. In the five traits, mean values of substrains varied remarkably and showed a continuous spectrum of distribution, suggesting control by multiple genes at distinct loci for each trait. We also screened for quantitative trait loci (QTLs) involved in the five traits. Suggestive QTLs for body weight (Chromosomes 1 and 6), insulin (Chromosomes 1, 3, 10 and 17), triglyceride (Chromosomes 4 and 11) and phospholipid (Chromosome 18) levels were detected. The SMXA RI strains are unique tools for analyzing genetic factors that influence body weight, blood insulin and lipids levels.     

A VPE family supporting various vacuolar functions in plants

Vacuolar processing enzyme (VPE) is a cysteine protease that has substrate specificity toward Asn and Asp residues, and found in various eukaryotic organisms including higher plants and mammals. Plant VPEs are separated into three subfamilies: seed-type, vegetative-type and uncharacterized-type. VPE was originally identified as a protease responsible for the maturation of seed storage proteins, and recent research has shown that it is a key protease responsible for the maturation of various vacuolar proteins not only in maturating cotyledons, but also in vegetative tissues. Thus, the VPE-mediated processing system is important for various vacuolar functions in the plant. Vegetative-type VPEs are expressed during senescence or pathogen-induced hypersensitive response. A VPE-deficiency abolished programmed cell death during hypersensitive response in tobacco leaves after TMV infection. This suggests that vegetative-type VPEs are involved in vacuolar-organized programmed cell death.     

A defect in glyoxysomal fatty acid beta-oxidation reduces jasmonic acid accumulation in Arabidopsis.

The final steps of jasmonic acid (JA) biosynthesis are thought to involve peroxisomal beta-oxidation, but this has not been directly demonstrated. The last and key step in fatty acid beta-oxidation is catalyzed by 3-ketoacyl-CoA thiolase (KAT) (EC 2.3.1.16). A mutant of Arabidopsis thaliana ecotype Landsberg erecta, which lacks a functional KAT protein and is defective in glyoxysomal fatty acid beta-oxidation has been reported. In this study, the mutant was found to accumulate reduced level of JA in both its wounded cotyledons and leaves, while only the cotyledons accumulate 3-oxo-2-(pent-2'-enyl)-cyclopentane-1-octanoic acid (OPC-8:0). This indicates that a defect in one of the thiolase isoenzymes impairs beta-oxidation of OPC-8:0 to JA. The mutant had sufficient thiolase activity for the synthesis of JA in the unwounded but not in the wounded tissues. Activities of the enzymes in the JA pathway that catalyze the steps, which precede beta-oxidation were not altered by the mutation in a thiolase protein. Thus, reduced levels of JA in the wounded tissues of the mutant were attributed to the defect in a thiolase protein.     

Comparison of analytical methods for profiling <Emphasis Type="Italic">N-</Emphasis> and <Emphasis Type="Italic">O-</Emphasis>linked glycans from cultured cell lines

The Human Disease Glycomics/Proteome Initiative (HGPI) is an activity in the Human Proteome Organization (HUPO) supported by leading researchers from international institutes and aims at development of disease-related glycomics/glycoproteomics analysis techniques. Since 2004, the initiative has conducted three pilot studies. The first two were N- and O-glycan analyses of purified transferrin and immunoglobulin-G and assessed the most appropriate analytical approach employed at the time. This paper describes the third study, which was conducted to compare different approaches for quantitation of N- and O-linked glycans attached to proteins in crude biological samples. The preliminary analysis on cell pellets resulted in wildly varied glycan profiles, which was probably the consequence of variations in the pre-processing sample preparation methodologies. However, the reproducibility of the data was not improved dramatically in the subsequent analysis on cell lysate fractions prepared in a specified method by one lab. The study demonstrated the difficulty of carrying out a complete analysis of the glycome in crude samples by any single technology and the importance of rigorous optimization of the course of analysis from preprocessing to data interpretation. It suggests that another collaborative study employing the latest technologies in this rapidly evolving field will help to realize the requirements of carrying out the large-scale analysis of glycoproteins in complex cell samples.     

Scanning for a unified model for translational repression by microRNAs

MicroRNAs (miRNAs) silence target mRNAs by inhibiting translation and subsequently initiating mRNA decay. The mechanism by which miRNAs silence translation is still poorly understood, with a number of competing models proposed. In this issue of The EMBO Journal, Kuzuo?lu‐Öztürk et al ( 2016 ) investigated miRNA silencing in human and insect cells. Their data support a model whereby miRNAs inhibit translation initiation. However, in contrast to several recent reports, their data suggest that translational inhibition is independent of 43S ribosomal subunit scanning, eIF4A translation factor activity, and 5′UTR secondary structure.