Interspecies differences in the enantioselectivity of epoxide hydrolases in Cryptococcus laurentii (Kufferath) C.E. Skinner and Cryptococcus podzolicus (Bab'jeva & Reshetova) Golubev

Isolates representing Cryptococcus laurentii and Cryptococcus podzolicus, originating from soil of a heathland indigenous to South Africa, were screened for the presence of enantioselective epoxide hydrolases for 2,2-disubstituted epoxides. Epoxide hydrolase activity for the 2,2-disubstituted epoxide (+/-)-2-methyl-2-pentyl oxirane was found to be abundantly present in all isolates. The stereochemistry of the products formed by the epoxide hydrolase enzymes from isolates belonging to the two species (11 isolates representing C. laurentii and 23 isolates representing C. podzolicus) was investigated. The enantiopreferences of the epoxide hydrolases for 2,2-disubstituted epoxides of these two species were found to be opposite. All strains of C. laurentii preferentially hydrolysed the (S)-epoxides while all C. podzolicus isolates preferentially hydrolysed the (R)-epoxides of (+/-)-2,2-disubstituted epoxides. These findings indicate that the stereochemistry of the products formed from 2,2-disubstituted epoxides by the epoxide hydrolase enzymes of these yeasts should be evaluated as additional taxonomic criterion within the genus Cryptococcus. Also, the selectivity of some epoxide hydrolases originating from isolates of C. podzolicus was high enough to be considered for application in biotransformations for the synthesis of enantiopure epoxides and vicinal diols.     

Structural peculiarities of the body wall of Tubificoides benedii (Oligochaeta) and possible relations to its life in sulphidic sediments

Summary Tubificoides benedii [=Peloscolex benedeni] a ubiquitous tubificid from poorly oxygenated, often polluted coastal muds, is known to be exceptionally well adapted to sulphidic sediments. However, almost nothing is known about its structural peculiarities, such as the conspicuously papillate body surface and possible relations to its unusual ecology. As a consequence, a study of this abundant but extraordinary marine worm has been made with the use of light and electron microscopy. While many internal structures correspond to the general pattern of marine tubificids and are not mentioned here, the epidermis — cuticle complex is unusual. The thick cuticle forms numerous high leaf-shaped papillae covered by condensed, almost solid mucus caps. The intermediate furrows usually harbour many different bacteria embedded in mucus. This mucus cover is rich in precipitates containing sulphur and other xenobiotic substances. Together with the cuticular papillae it can be sloughed off in a moulting process. Epicuticular projections, usually typical of oligochaetes, are absent from most parts of the body except from the first and last segments. The epidermal cells often contain numerous extremely long and abnormally shaped mitochondria. The significance of the peculiar structure of the body wall and the distinct moulting are discussed in the light of the ecological situation of these tubificids.     

Hymenopterous EGG-pupal and larval-pupal parasitoids ofCeratitis capitata andAnastrepha spp. [Dip.: Tephritidae] in Costa Rica

Egg-pupal and larval-pupal parasitoids were recovered from less than 10% of the 16,000 tephritid puparia collected in Costa Rica from August, 1979, through April, 1980.Ceratitis capitata (Wiedemann) was attacked by 2 introduced opiineBraconidae and 2 indigenous eucoilineCynipidae Anastrepha spp. were attacked by each parasitoid species attackingC. capitata and also by 5 indigenous opiineBraconidae and 1 exoticEulophidae. Toxotrypana curvicauda Gerstaecker was attacked only by an indigenous opiineBraconidae which did not attack other tephritid species collected.     

Influence of cranial deformation on facial morphology among prehistoric South Central Andean populations

Calculating biodistances among South American populations using cranial measurements is often hindered, as many available skeletal collections exhibit deformation. Acknowledging vault modifications, researchers have sought measurements in other regions which are unaffected by deformation. In the 1970s, a set of 10 "relatively" unaffected facial measurements was identified in Argentinean crania that later became the basis of numerous South American biodistance studies. These measurements include: minimum frontal breadth, bizygomatic breadth, orbit height, orbit breadth, palate breath, palate length, upper facial height, basion-prosthion length, nasal height, and nasal breadth. Palate length was excluded from the present analysis due to considerable measurement error. The suitability of these measurements in populations other than Argentineans has not been rigorously tested. Using a sample of 350 prehistoric crania from the Museo Arqueológico San Miguel de Azapa (MASMA, Arica, Chile), this project tested the hypothesis that these measurements are unaffected by either annular or tabular deformation. Results obtained from MANOVA analysis indicate this hypothesis cannot be fully supported. Among males, only 3 of the 9 measurements are unaffected by either form of deformation (palate breadth, basion-prosthion length, and nasal breadth), while analysis of females indicates that 4 of the 9 measurements remain unaltered (minimum frontal breadth, orbit breadth, basion-prosthion length, and nasal breadth). Additionally, analogous to the vault, facial measurements display patterns consistent with the deformation applied. Two implications can be drawn from this research: 1) previous studies using these measurements must be interpreted cautiously, and 2) researchers using these measurements must explicitly test their suitability in each population.     

The b-32 protein from maize endosperm, an albumin regulated by the O2 locus: Nucleic acid (cDNA) and amino acid sequences

Summary The cDNA coding for the b-32 protein, an albumin expressed in maize endosperm cells under the control of the O2 and O6 loci, has been cloned and the complete amino acid sequence of the protein derived. A lambda gt11 cDNA library from mRNA of immature maize endosperm was screened for the expression of the b-32 protein using antibodies against the purified protein. One of the positive clones obtained was used to isolate a full-length cDNA clone. By Northern analysis, the size of the b-32 mRNA was estimated to be 1.2 kb. Hybrid-selected translation assays show that the message codes for a protein with an apparent molecular weight of 30–35 kDa. The nucleotide sequence shows that several internal repeats are present. The protein has a length of 303 amino acid residues (mol. wt. 32430 dalton) and its sequence shows the following features: no signal peptide is observable; it contains seven tryptophan residues, an amino acid absent in maize storage proteins; polar and hydrophobic residues are spread along the sequence; several pairs of basic residues are present in the N-terminal region; the secondary structure allows the prediction of two structural domains for the b-32 protein that would fold up giving rise to a globular shape. The cloning of this gene may help in understanding the role of the O2 and O6 loci in regulating the deposition of zein, the major storage protein of maize endosperm.     

Proximity of conserved U6 and U2 snRNA elements to the 5' splice site region in activated spliceosomes

Major structural changes occur in the spliceosome during its catalytic activation, which immediately precedes the splicing of pre-mRNA. Whereas changes in snRNA conformation are well documented at the level of secondary RNA-RNA interactions, little is known about the tertiary structure of this RNA-RNA network, which comprises the spliceosome's catalytic core. Here, we have used the hydroxyl-radical probe Fe-BABE, tethered to the tenth nucleotide (U(+10)) of the 5' end of a pre-mRNA intron, to map RNA-RNA proximities in spliceosomes. These studies revealed that several conserved snRNA regions are close to U(+10) in activated spliceosomes, namely (i) the U6 snRNA ACAGAG-box region, (ii) portions of the U6 intramolecular stem-loop (U6-ISL) including a nucleotide implicated in the first catalytic step (U74), and (iii) the region of U2 that interacts with the branch point. These data constrain the relative orientation of these structural elements with respect to U(+10) in the activated spliceosome. Upon conversion of the activated spliceosome to complex C, the accessibility of U6-ISL to hydroxyl-radical cleavage is altered, suggesting rearrangements after the first catalytic step.     

The lactosylceramide binding specificity of Helicobacter pylori

The possible role of glycosphingolipids as adhesion receptors for the human gastric pathogen Helicobacter pylori was examined by use of radiolabeled bacteria, or protein extracts from the bacterial cell surface, in the thin-layer chromatogram binding assay. Of several binding specificities found, the binding to lactosylceramide is described in detail here, the others being reported elsewhere. By autoradiography a preferential binding to lactosylceramide having sphingosine/phytosphingosine and 2-D hydroxy fatty acids was detected, whereas lactosylceramide having sphingosine and nonhydroxy fatty acids was consistently nonbinding. A selective binding of H. pylori to lactosylceramide with phytosphingosine and 2-D hydroxy fatty acid was obtained when the different lactosylceramide species were incorporated into liposomes, but only in the presence of cholesterol, suggesting that this selectivity may be present also in vivo . Importantly, lactosylceramide with sphingosine and hydroxy fatty acids does not bind in this assay. Furthermore, a lactosylceramide-based binding pattern obtained for different trisaccharide glycosphingolipids is consistent with the assumption that this selectivity is due to binding of a conformation of lactosylceramide in which the oxygen of the 2-D fatty acid hydroxyl group forms a hydrogen bond with the Glc hydroxy methyl group, yielding an epitope presentation different from other possible conformers. An alternative conformation that may come into consideration corresponds to the crystal structure found for cerebroside, in which the fatty acid hydroxyl group is free to interact directly with the adhesin. By isolating glycosphingolipids from epithelial cells of human stomach from seven individuals, a binding of H.pylori to the diglycosylceramide region of the non-acid fraction could be demonstrated in one of these cases. Mass spectrometry showed that the binding-active sample contained diglycosylceramides with phytosphingosine and 2-D hydroxy fatty acids with 16-24 carbon atoms in agreement with the results related above.      

The R-spondin protein family

The four vertebrate R-spondin proteins are secreted agonists of the canonical Wnt/β-catenin signaling pathway. These proteins are approximately 35 kDa, and are characterized by two amino-terminal furin-like repeats, which are necessary and sufficient for Wnt signal potentiation, and a thrombospondin domain situated more towards the carboxyl terminus that can bind matrix glycosaminoglycans and/or proteoglycans. Although R-spondins are unable to initiate Wnt signaling, they can potently enhance responses to low-dose Wnt proteins. In humans, rare disruptions of the gene encoding R-spondin1 cause a syndrome of XX sex reversal (phenotypic male), palmoplantar keratosis (a thickening of the palms and soles caused by excess keratin formation) and predisposition to squamous cell carcinoma of the skin. Mutations in the gene encoding R-spondin4 cause anonychia (absence or hypoplasia of nails on fingers and toes). Recently, leucine-rich repeat-containing G-protein-coupled receptor (Lgr)4, Lgr5 and Lgr6, three closely related orphans of the leucine-rich repeat family of G-protein-coupled receptors, have been identified as receptors for R-spondins. Lgr5 and Lgr6 are markers for adult stem cells. Because R-spondins are potent stimulators of adult stem cell proliferation in vivo and in vitro, these findings might guide the therapeutic use of R-spondins in regenerative medicine.     

Molecular evolution of a Y-chromosomal repetitive sequence family in the genus Mus

A 522-base-long Y-chromosomal sequence was isolated from a BALB/c genomic library and was designated "BF046." It is repeated about 200 times in the male genome, and a difference was detected between the Mus musculus musculus and the M. m. domesticus type Y chromosomes. BF046- related sequences were present over the entire length of the Y chromosome as visualized by in situ hybridization. Southern blot analysis against DNAs isolated from eight species in the genus Mus showed that BF046-related sequences were amplified in the Y chromosomes of three closely related species: M. musculus, M. spicilegus, and M. spretus. To gain insight into the stability of the BF046 sequence family, we isolated 18 additional clones from these three mouse species and compared their sequences. The M. musculus sequences differed from the M. spicilegus and M. spretus sequences by two indels. The remaining parts of the sequences were very similar, but both parsimony and distance-based analytical methods divided the sequences into the same four subgroups, with each species having its own subgroup(s). Thus, the Y chromosomes of M. musculus, M. spicilegus, and M. spretus can be distinguished from one another.