A serum-free system for culturing human placental trophoblasts

Summary We have compared hormone production by early gestation and term human placental trophoblasts cultured in Ham's F10 medium containing 10% fetal bovine serum with that by cells cultured in serum-free HB102 medium. Mean daily production of progesterone on Days 3 to 7 was approximately 25% less by both early gestation and term cells cultured in HB102 as compared to Ham's F10, but production was maintained at a stable level for at least 7 d longer than the cells in Ham's. Estradiol production from 10⁻⁶ M dehydroepiandrosterone by both early gestation and term cells was comparable in both media. Human placental lactogen production on Days 3 to 7 was 40% less by cells cultured in HB102. Human chorionic gonadotropin (hCG) output by early gestation cells was also 50% less in HB102 but term cells in HB102 produced twice as much hCG as those in Ham's F10. 3B-Hydroxysteroid dehydrogenase (3BHSD) activity in early gestation and term cells and 11B-hydroxysteroid dehydrogenase (11BHSD) activity of early gestation cultures was comparable in the two media. 11BHSD activity was decreased in the term cultures, and this decrease was more marked in Ham's than in HB102. Sulfatase and aromatase activities in the early gestation cultures were comparable in both media; sulfatase activity was comparable and aromatase activity only 20% less in the term cells cultured in HB102. These results indicate that serum-free HB102 supports differential function of human trophoblast cells and is useful for studies of placental activity for as long as 14 d in culture.     

Spermatogenesis and sperm ultrastructure in the polychaete genusOphryotrocha (Dorvilleidae)

The details of spermatogenesis and spermiogenesis are described forOphryotrocha puerilis. The ultrastructure of mature sperm is shown forO. puerilis, O. hartmanni, O. gracilis, O. diadema, O. labronica, andO. notoglandulata. Clusters of sixteen cells each are proliferated by two stem cells in each setigerous segment ofO. puerilis representing the very early stages of both oogenesis and spermatogenesis. In each spermatocyte-I cluster, the cells are interconnected by cytoplasmic bridges. Early, clusters are enveloped by peritoneal sheath cells. These transient gonad walls break down prior to meiosis. The meiotic processes may start in the clusters with the cells still interconnected, or during breakdown of the original cluster, giving rise to smaller subclusters of both spermatocytes I and spermatocytes II with various numbers of cells. Finally, spermatid tetrads are present. As spermiogenesis progresses, the tetrads disintegrate. Golgi vesicles in both spermatocytes and spermatids contain electron-dense material, presumably preacrosomal. The acrosome is formed by such vesicles. In the six species studied here, the acrosomes appear to be of a similar overall structure but are of different shape. Centrioles are usually located beneath the acrosome. The distal centriole forms the basal body of a flagellum-like cytoplasmic process. The microtubules of these flagellar equivalents do not show a normal ciliar arrangement. The flagellar equivalent appears to be non-motile. InO. hartmanni and inO. notoglandulata, a flagellar equivalent is missing. Microtubules originating from the proximal end of the distal centriole stretch to the nuclear envelope. This feature appears to be especially conspicuous inO. puerilis and inO. labronica. InO. labronica and inO. notoglandulata, bundles of microtubules paralleling the cell perimeter appear to stabilise the sperm. Various numbers of mitochondria are either randomly distributed around the nucleus or accumulate on one side, often directly under the acrosome. Parts of the present paper were presented at the 2^nd International Polychaete Conference, Copenhagen 1986 and at the 3^rd International Polychaete Conference, Long Beach, Ca. 1989.     

Differentiation of Serratia liquefaciens into swarm cells is controlled by the expression of the flhD master operon.

The velocity with which a swarming colony of Serratia liquefaciens colonizes the surface of a suitable solid substratum was controlled by modulating the expression of the flhD master operon. In liquid medium, the stimulation of flhD expression resulted in filamentous, multinucleate, and hyperflagellated cells that were indistinguishable from swarm cells isolated from the edge of a swarm colony. Thus, expression of the flhD master operon appears to play a central role in the process of swarm cell differentiation.     

Crystal structure of a bacterial family-III cellulose-binding domain: a general mechanism for attachment to cellulose.

The crystal structure of a family-III cellulose-binding domain (CBD) from the cellulosomal scaffoldin subunit of Clostridium thermocellum has been determined at 1.75 A resolution. The protein forms a nine-stranded beta sandwich with a jelly roll topology and binds a calcium ion. conserved, surface-exposed residues map into two defined surfaces located on opposite sides of the molecule. One of these faces is dominated by a planar linear strip of aromatic and polar residues which are proposed to interact with crystalline cellulose. The other conserved residues are contained in a shallow groove, the function of which is currently unknown, and which has not been observed previously in other families of CBDs. On the basis of modeling studies combined with comparisons of recently determined NMR structures for other CBDs, a general model for the binding of CBDs to cellulose is presented. Although the proposed binding of the CBD to cellulose is essentially a surface interaction, specific types and combinations of amino acids appear to interact selectively with glucose moieties positioned on three adjacent chains of the cellulose surface. The major interaction is characterized by the planar strip of aromatic residues, which align along one of the chains. In addition, polar amino acid residues are proposed to anchor the CBD molecule to two other adjacent chains of crystalline cellulose.     

Genetic variation,breeding system evolution,and conservation of the narrow sand dune endemic Stellaria arenicola and the widespread S. longipes (Caryophyllaceae)

Genetic variation was examined by electrophoresis in 14 populations of Stellaria arenicola, an endemic of the Athabasca sand dunes in northern Saskatchewan, Canada, and seven populations of S. longipes, its progenitor. Three of the 5. longipes populations were sympatric with the endemic. Populations of the endemic were found to have fewer alleles per polymorphic locus (2.21 vs. 2.37), fewer polymorphic loci (29.9 vs. 33.8), and lower genetic diversity (0.087 vs. 0.107) than populations of the progenitor. Genetic identities for all pairs of populations were high (0.932 to 1.000). The endemic had one novel allele and shared ten alleles with progenitor populations from the sand dunes that were not found in other populations of S. longipes. Populations of both species were found to partition most of their genetic variation within populations. An investigation of the multilocus outcrossing rates revealed that S. arenicola had higher rates of selling and biparental inbreeding than S. longipes. This study suggests that partial genetic isolation through a shift in the breeding system, in addition to previously reported strong directional selection, has been important in the sympatric evolution of the endemic S. arenicola. The close genetic relationship between populations of S. arenicola and S. longipes found on the Athabasca sand dunes supports the suggestion that the endemic evolved while sympatric to the gene pool of the progenitor species that is found presently in the region.     

Sequencing analysis reveals a unique gene organization in the gyrB region of Mycoplasma hominis.

The homolog of the gyrB gene, which has been reported to be present in the vicinity of the initiation site of replication in bacteria, was mapped on the Mycoplasma hominis genome, and the region was subsequently sequenced. Five open reading frames were identified flanking the gyrB gene, one of which showed similarity to that which encodes the LicA protein of Haemophilus influenzae. The organization of the genes in the region showed no resemblance to that in the corresponding regions of other bacteria sequenced so far. The gyrA gene was mapped 35 kb downstream from the gyrB gene.     

A new,pathogen-inducible gene of Arabidopsis is expressed in an ecotype-specific manner

In this study we describe a novel gene, which was isolated in an attempt to search for specific plant resistance genes of Arabidopsis against isolates of the phytopathogenic bacterium Xanthomonas campestris pv. campestris. The gene was cloned by differential screening of a genomic library of the Xcc 750-resistant ecotype Col-0, using cDNA populations derived from ecotype Col-0 and the Xcc 750-susceptible ecotype Oy-0. The isolated gene, CXc750, is differentially expressed in ecotypes of Arabidopsis thaliana. In addition, although highly expressed in uninfected plants, gene expression increases in response to pathogen attack. CXc750 potentially codes for a small, basic protein of about 10 kDa. The predicted protein product contains a potential signal leader peptide at the amino-terminal end but no ER retention sequence and no further transmembrane domain. This indicates that the gene product is transported to other compartments or out of the cell.The possible function of CXc750 as a member of the plant defense response system is discussed.     

Anthranilate synthase and chorismate mutase activities in transgenic tobacco plants overexpressing tryptophan decarboxylase fromCatharanthus roseus

TransgenicNicotiana tabacum L. Petit Havana SR1 F₁-plants expressing tryptophan decarboxylase cDNA (tdc) fromCatharanthus roseus (L.) G. Don under the control of the CaMV 35S promoter and terminator exhibited tryptophan decarboxylase (TDC) enzyme activity and accumulated tryptamine. The plants with the highest TDC activity contained 19 pkat per mg of protein. The influence of transgenic expression oftdc on the activities of anthranilate synthase (AS) and chorismate mutase (CM) were examined in 10 transgenic tobacco plants. The specific activities of these two chorismate-utilizing enzymes were not significantly affected by expression oftdc, despite their important functions as branch point enzymes in the shikimate pathway. The results indicate that the normal route of tryptophan biosynthesis in plants is sufficient to supply a considerable amount of this essential amino acid for the biosynthesis of secondary metabolites. Despite their increased tryptamine content, the growth and development of the transgenic tobacco plants expressingtdc appeared normal.     

394delTT: a Nordic cystic fibrosis mutation

In a systematic screening for mutations in the gene encoding the cystic fibrosis transmembrane regulator among Danish cystic fibrosis (CF) patients, we identified a mutation in exon 3 (394delTT); this mutation was found to be relatively common in Denmark. We therefore screened for 394delTT in Sweden and Norway, where it turned out to be the second most frequent mutation, accounting for 4% of all CF mutations. It also occurs with a high frequency in Finland, but has not been found in larger surveys of mutations in the CFTR gene. Thus, 394delTT seems to be a specific Nordic CF mutation.