Regulation of the galactose-inducible lac operon and the histidine utilization operons in pts mutants of Klebsiella aerogenes.

Galactose appears to be the physiological inducer of the chromosomal lac operon in Klebsiella aerogenes. Both lactose and galactose are poor inducers in strains having a functional galactose catabolism (gal) operon, but both are excellent inducers in gal mutants. Thus the slow growth of K. aerogenes on lactose reflects the rapid degradation of the inducer. Several pts mutations were characterized and shown to affect both inducer exclusion and permanent catabolite repression. The beta-galactosidase of pts mutants cannot be induced at all by lactose, and pts mutants appear to have a permanent and constitutive inducer exclusion phenotype. In addition, pts mutants show a reduced rate of glucose metabolism, leading to slower growth on glucose and a reduced degree of glucose-mediated permanent catabolite repression. The crr-type pseudorevertants of pts mutations relieve the constitutive inducer exclusion for lac but do not restore the full level of glucose-mediated permanent catabolite repression and only slightly weaken the glucose-mediated inducer exclusion. Except for weakening the glucose-mediated permanent catabolite repression, pts and crr mutations have no effect on expression of the histidine utilization (hut) operons.     

Transfer of rpl22 to the nucleus greatly preceded its loss from the chloroplast and involved the gain of an intron.

Most chloroplast and mitochondrial proteins are encoded by nuclear genes that once resided in the organellar genomes. Transfer of most of these genes appears to have occurred soon after the endosymbiotic origin of organelles, and so little is known about the process. Our efforts to understand how chloroplast genes are functionally transferred to the nuclear genome have led us to discover the most recent evolutionary gene transfer yet described. The gene rpl22, encoding chloroplast ribosomal protein CL22, is present in the chloroplast genome of all plants examined except legumes, while a functional copy of rpl22 is located in the nucleus of the legume pea. The nuclear rpl22 gene has acquired two additional domains relative to its chloroplast ancestor: an exon encoding a putative N-terminal transit peptide, followed by an intron which separates this first exon from the evolutionarily conserved, chloroplast-derived portion of the gene. This gene structure suggests that the transferred region may have acquired its transit peptide by a form of exon shuffling. Surprisingly, phylogenetic analysis shows that rpl22 was transferred to the nucleus in a common ancestor of all flowering plants, at least 100 million years preceding its loss from the legume chloroplast lineage.     

European guidelines for quality assurance in cervical cancer screening: recommendations for collecting samples for conventional and liquid-based cytology.

The current paper presents an annex in the second edition of the European Guidelines for Quality Assurance in Cervical Cancer Screening. It provides guidance on how to make a satisfactory conventional Pap smear or a liquid-based cytology (LBC) sample. Practitioners taking samples for cytology should first explain to the woman the purpose, the procedure and how the result will be communicated. Three sampling methods are considered as acceptable for preparing conventional Pap smears: (i) the cervical broom; (ii) the combination of a spatula and an endocervical brush; and (iii) the extended tip spatula. Smear takers should take care to sample the entire circumference of the transformation zone, to quickly spread the cellular material over a glass slide, and to fix the preparation within a few seconds to avoid drying artefacts. According to local guidelines, one of these three methods may be preferred. Sampling with a cotton tip applicator is inappropriate. Similar procedures should be followed for sampling cells for LBC, but only plastic devices may be used. The collected cells should be quickly transferred into a vial with fixative liquid according to the instructions of the manufacturer of the LBC system. Subsequently, the slide or vial and the completed request form are sent to the laboratory for cytological interpretation.     

A new genus, Helgoeca gen. nov., for a nudiform choanoflagellate

A new genus, Helgoeca gen. nov., has been designated to accommodate a nudiform loricate choanoflagellate (American Type Culture Collection strain ATCC 50073) that was incorrectly attributed to the tectiform genus Acanthoecopsis (=Acanthocorbis). The first indication that this species might be nudiform came from a four-gene phylogeny of the choanoflagellates which recovered ATCC 50073 within a strongly supported monophyletic clade comprising two other nudiform taxa. Fortunately an isolate of the species in question was available from the ATCC and when observed in rapidly growing culture it was immediately apparent that this species divided with the production of 'naked' motile cells; a typically nudiform character. The beaker-shaped lorica of this species consists of an outer layer of approximately 11 longitudinal costae, which terminate anteriorly as spines, and an equal or larger number of helical costae, with a left-handed conformation, each of which terminates anteriorly adjacent to the base of a spine. The pattern of costae in this species is indistinguishable from that of Acanthocorbis nana Thomsen and for this reason A. nana has been transferred to the new genus Helgoeca gen. nov., as the type species.     

Modulation of the endocannabinoid system in viable and non-viable first trimester pregnancies by pregnancy-related hormones

Background

In early pregnancy, increased plasma levels of the endocannabinoid anandamide (AEA) are associated with miscarriage through mechanisms that might affect the developing placenta or maternal decidua.

Methods

In this study, we compare AEA levels in failed and viable pregnancies with the levels of the trophoblastic hormones (beta-human chorionic gonadotrophin (beta-hCG), progesterone (P4) and (pregnancy-associated placental protein-A (PAPP-A)) essential for early pregnancy success and relate that to the expression of the cannabinoid receptors and enzymes that modulate AEA levels.

Results

The median plasma AEA level in non-viable pregnancies (1.48 nM; n = 20) was higher than in viable pregnancies (1.21 nM; n = 25; P = 0.013), as were progesterone and beta-hCG levels (41.0 vs 51.5 ng/mL; P = 0.052 for P4 and 28,650 vs 6,560 mIU/L; P = 0.144 for beta-hCG, respectively, but were not statistically significant). Serum PAPP-A levels in the viable group were approximately 6.8 times lower than those in the non-viable group (1.82 vs 12.25 mg/L; P = 0.071), but again these differences were statistically insignificant. In the spontaneous miscarriage group, significant correlations between P4 and beta-hCG, P4 and PAPP-A and AEA and PAPP-A levels were observed. Simultaneously, immunohistochemical distributions of the two main cannabinoid receptors and the AEA-modifying enzymes, fatty acid amide hydrolase (FAAH) and N-acylphosphatidylethanolamine-phospholipase D (NAPE-PLD), changed within both the decidua and trophoblast.

Conclusions

The association of higher AEA levels with early pregnancy failure and with beta-hCG and PAPP-A, but not with progesterone concentrations suggest that plasma AEA levels and pregnancy failure are linked via a mechanism that may involve trophoblastic beta-hCG, and PAPP-A, but not, progesterone production. Although the trophoblast, decidua and embryo contain receptors for AEA, the main AEA target in early pregnancy failure remains unknown.     

Evolution of substrate recognition across a multigene family of glycosyltransferases in Arabidopsis

The complete sequence of the Arabidopsis genome enables definitive characterization of multigene families and analysis of their phylogenetic relationships. Using a consensus sequence previously defined for glycosyltransferases that use small-molecular-weight acceptors, 107 gene sequences were identified in the Arabidopsis genome and used to construct a phylogenetic tree. Screening recombinant proteins for their catalytic activities in vitro has revealed enzymes active toward physiologically important substrates, including hormones and secondary metabolites. The aim of this study has been to use the phylogenetic relationships across the entire family to explore the evolution of substrate recognition and regioselectivity of glucosylation. Hydroxycoumarins have been used as the model substrates for the analysis in which 90 sequences have been assayed and 48 sequences shown to recognize these compounds. The study has revealed activity in 6 of the 14 phylogenetic groups of the multigene family, suggesting that basic features of substrate recognition are retained across substantial evolutionary periods.     

The protistan origins of animals and fungi

Recent molecular studies suggest that Opisthokonta, the eukaryotic supergroup including animals and fungi, should be expanded to include a diverse collection of primitively single-celled eukaryotes previously classified as Protozoa. These taxa include corallochytreans, nucleariids, ministeriids, choanoflagellates, and ichthyosporeans. Assignment of many of these taxa to Opisthokonta remains uncorroborated as it is based solely on small subunit ribosomal RNA trees lacking resolution and significant bootstrap support for critical nodes. Therefore, important details of the phylogenetic relationships of these putative opisthokonts with each other and with animals and fungi remain unclear. We have sequenced elongation factor 1-alpha (EF-1alpha), actin, beta-tubulin, and HSP70, and/or alpha-tubulin from representatives of each of the proposed protistan opisthokont lineages, constituting the first protein-coding gene data for some of them. Our results show that members of all opisthokont protist groups encode a approximately 12-amino acid insertion in EF-1alpha, previously found exclusively in animals and fungi. Phylogenetic analyses of combined multigene data sets including a diverse set of opisthokont and nonopisthokont taxa place all of the proposed opisthokont protists unequivocally in an exclusive clade with animals and fungi. Within this clade, the nucleariid appears as the closest sister taxon to fungi, while the corallochytrean and ichthyosporean form a group which, together with the ministeriid and choanoflagellates, form two to three separate sister lineages to animals. These results further establish Opisthokonta as a bona fide taxonomic group and suggest that any further testing of the legitimacy of this taxon should, at the least, include data from opisthokont protists. Our results also underline the critical position of these "animal-fungal allies" with respect to the origin and early evolution of animals and fungi.     

LOW THERMAL LIMIT OF GROWTH RATE OF SYMBIODINIUM CALIFORNIUM (DINOPHYTA) IN CULTURE MAY RESTRICT THE SYMBIONT TO SOUTHERN POPULATIONS OF ITS HOST ANEMONES (ANTHOPLEURA SPP.; ANTHOZOA,CNIDARIA)1

Symbiodinium californium (#383, Banaszak et al. 1993 ) is one of two known dinoflagellate symbionts of the intertidal sea anemones Anthopleura elegantissima, A. xanthogrammica, and A. sola and occurs only in hosts at southern latitudes of the North Pacific. To investigate if temperature restricts the latitudinal distribution of S. californium, growth and photosynthesis at a range of temperatures (5°C–30°C) were determined for cultured symbionts. Mean specific growth rates were the highest between 15°C and 28°C (μ 0.21–0.26 · d^?1) and extremely low at 5, 10, and 30°C (0.02–0.03 · d^?1). Average doubling times ranged from 2.7 d (20°C) to 33 d (5, 10, and 30°C). Cells cultured at 10°C had the greatest cell volume (821 μm³) and the highest percentage of motile cells (64.5%). Growth and photosynthesis were uncoupled; light‐saturated maximum photosynthesis (P_max) increased from 2.9 pg C · cell^?1 · h^?1 at 20°C to 13.2 pg C · cell^?1 · h^?1 at 30°C, a 4.5‐fold increase. Less than 11% of daily photosynthetically fixed carbon was utilized for growth at 5, 10, and 30°C, indicating the potential for high carbon translocation at these temperatures. Low temperature effects on growth rate, and not on photosynthesis and cell morphology, may restrict the distribution of S. californium to southern populations of its host anemones.     

The Protein Tyrosine Phosphatase Rptpζ Suppresses Osteosarcoma Development in Trp53-Heterozygous Mice

Osteosarcoma (OS), a highly aggressive primary bone tumor, belongs to the most common solid tumors in growing children. Since specific molecular targets for OS treatment remain to be identified, surgical resection combined with multimodal (neo-)adjuvant chemotherapy is still the only way to help respective individuals. We have previously identified the protein tyrosine phosphatase Rptpζ as a marker of terminally differentiated osteoblasts, which negatively regulates their proliferation in vitro. Here we have addressed the question if Rptpζ can function as a tumor suppressor protein inhibiting OS development in vivo. We therefore analyzed the skeletal phenotype of mice lacking Ptprz1, the gene encoding Rptpζ on a tumor-prone genetic background, i.e. Trp53-heterozygosity. By screening a large number of 52 week old Trp53-heterozygous mice by contact radiography we found that Ptprz1-deficiency significantly enhanced OS development with 19% of the mice being affected. The tumors in Ptprz1-deficient Trp53-heterozygous mice were present in different locations (spine, long bones, ribs), and their OS nature was confirmed by undecalcified histology. Likewise, cell lines derived from the tumors were able to undergo osteogenic differentiation ex vivo. A comparison between Ptprz1-heterozygous and Ptprz1-deficient cultures further revealed that the latter ones displayed increased proliferation, a higher abundance of tyrosine-phosphorylated proteins and resistance towards the influence of the growth factor Midkine. Our findings underscore the relevance of Rptpζ as an attenuator of proliferation in differentiated osteoblasts and raise the possibility that activating Rptpζ-dependent signaling could specifically target osteoblastic tumor cells.