The conserved U.G pair in the 5'' splice site duplex of a group I intron is required in the first but not the second step of self-splicing.

Group I self-splicing introns have a 5' splice site duplex (P1) that contains a single conserved base pair (U.G). The U is the last nucleotide of the 5' exon, and the G is part of the internal guide sequence within the intron. Using site-specific mutagenesis and analysis of the rate and accuracy of splicing of the Tetrahymena thermophila group I intron, we found that both the U and the G of the U.G pair are important for the first step of self-splicing (attack of GTP at the 5' splice site). Mutation of the U to a purine activated cryptic 5' splice sites in which a U.G pair was restored; this result emphasizes the preference for a U.G at the splice site. Nevertheless, some splicing persisted at the normal site after introduction of a purine, suggesting that position within the P1 helix is another determinant of 5' splice site choice. When the U was changed to a C, the accuracy of splicing was not affected, but the Km for GTP was increased by a factor of 15 and the catalytic rate constant was decreased by a factor of 7. Substitution of U.A, U.U, G.G, or A.G for the conserved U.G decreased the rate of splicing by an even greater amount. In contrast, mutation of the conserved G enhanced the second step of splicing, as evidenced by a trans-splicing assay. Furthermore, a free 5' exon ending in A or C instead of the conserved U underwent efficient ligation. Thus, unlike the remainder of the P1 helix, which functions in both the first and second steps of self-splicing, the conserved U.G appears to be important only for the first step.     

SPECIFIC CHALONE INHIBITION OF THE REGENERATION OF THE JB-1 ASCITES TUMOUR STUDIED BY FLOW MICROFLUOROMETRY

The variation in the DNA distribution in the JB-1 and the Lla₂ ascites tumour was investigated by means of flow microfluorometry (FMF) in the plateau stage and during the initiation of the regeneratave growth induced by percutaneous aspiration. The study showed that a considerable influx of cells with G₁ DNA content into the S phase occurred in both tumours about 10 hr after aspiration. In the JB-1 tumour, these initial regenerative changes could be reversibly blocked by injections of cell-free plateau JB-1 ascitic fluid or an ultrafiltrate of this ascites. In contrast to these observations no delay in the regenerative changes was observed in the Lla₂ tumour after treatment with JB-1 ascites or the ultrafiltrate. The study supports the assumption of a specific growth regulation of the JB-1 ascites tumour and emphasizes the suitability of FMF analyses in cell-kinetic studies in which short-term fluctuations take place in the distribution of cells with different DNA content.     

Structural and functional significance of association of a low molecular weight chalone from JB-1 ascites tumours with high molecular weight factors

Abstract. The cell-specific inhibitory (chalone) activity of JB-1 ascites tumour cell proliferation has been purified using five different procedures. By combining (1) molecular weight estimations based on ultrafiltration and gel chromatography, and (2) partitioning in organic solvent and ion exchangers, it is concluded that the active factor associates, in a complex manner, with various other components involving both hydrophobic and ionic forces. The active factor appears to be a slightly acidic, hydrophobic peptide (molecular weight 500–1000 D). When assessing the activity in vivo , it appears to be highly dependent on associated serum factors. Thus, the chalone studied appears to interact both structurally and functionally with various associated factors which affect its physicochemical behaviour and biological activity.     

Src regulates distinct pathways for cell volume control through Vav and phospholipase Cgamma

Cell volume recovery in response to swelling requires reorganization of the cytoskeleton and fluid efflux. We have previously shown that electrolyte and fluid efflux via K+ and Cl- channels is controlled by swelling-induced activation of phospholipase Cgamma (PLCgamma). Recently, integrin engagement has been suggested to trigger responses to swelling through activation of Rho family GTPases and Src kinases. Because both PLCgamma and Rho GTPases can be regulated by Src during integrin-mediated cytoskeletal reorganization, we sought to identify swelling-induced Src effectors. Upon hypotonic challenge, Src was rapidly activated in transient plasma membrane protrusions, where it colocalized with Vav, an activator of Rho GTPases. Inhibition of Src with PP2 attenuated phosphorylation of Vav. PP2 also attenuated phosphorylation of PLCgamma, and inhibited swelling-mediated activation of K+ and Cl- channels and cell volume recovery. These findings suggest that swelling-induced Src regulates cytoskeletal dynamics, through Vav, and fluid efflux, through PLCgamma, and thus can coordinate structural reorganization with fluid balance to maintain cellular integrity.     

Chondroitin sulfate A-adhering Plasmodium falciparum-infected erythrocytes express functionally important antibody epitopes shared by multiple variants

Acquired protection from Plasmodium falciparum placental malaria, a major cause of maternal, fetal, and infant morbidity, is mediated by IgG specific for the P. falciparum erythrocyte membrane protein 1 variant VAR2CSA. This protein enables adhesion of P. falciparum-infected erythrocytes to chondroitin sulfate A in the intervillous space. Although interclonal variation of the var2csa gene is lower than that among var genes in general, VAR2CSA-specific Abs appear to target mainly polymorphic epitopes. This has raised doubts about the feasibility of VAR2CSA-based vaccines. We used eight human monoclonal IgG Abs from affinity-matured memory B cells of P. falciparum-exposed women to study interclonal variation and functional importance of Ab epitopes among placental and peripheral parasites from East and West Africa. Most placental P. falciparum isolates were labeled by several mAbs, whereas peripheral isolates from children were essentially nonreactive. The mAb reactivity of peripheral isolates from pregnant women indicated that some were placental, whereas others had alternative sequestration foci. Most of the mAbs were comparable in their reactivity with bound infected erythrocytes (IEs) and recombinant VAR2CSA and interfered with IE and/or VAR2CSA binding to chondroitin sulfate A. Pair-wise mAb combinations were more inhibitory than single mAbs, and all of the mAbs together was the most efficient combination. Each mAb could opsonize IEs for phagocytosis, and a combination of the eight mAbs caused phagocytosis similar to that of plasma IgG-opsonized IEs. We conclude that functionally important Ab epitopes are shared by the majority of polymorphic VAR2CSA variants, which supports the feasibility of VAR2CSA-based vaccines against placental malaria.     

The Density of Knobs on Plasmodium falciparum-Infected Erythrocytes Depends on Developmental Age and Varies among Isolates

Background

The virulence of Plasmodium falciparum malaria is related to the parasite’s ability to evade host immunity through clonal antigenic variation and tissue-specific adhesion of infected erythrocytes (IEs). The P. falciparum erythrocyte membrane protein 1 (PfEMP1) family expressed on dome-shaped protrusions called knobs on the IE surface is central to both. Differences in receptor specificity and affinity of expressed PfEMP1 are important for IE adhesiveness, but it is not known whether differences in the number and size of the knobs on which the PfEMP1 proteins are expressed also play a role. Therefore, the aim of this study was to provide detailed information on isolate- and time-dependent differences in knob size and density.

Methodology/Principal Findings

We used atomic force microscopy to characterize knobs on the surface of P. falciparum-infected erythrocytes. Fourteen ex vivo isolates from Ghanaian children with malaria and 10 P. falciparum isolates selected in vitro for expression of a particular PfEMP1 protein (VAR2CSA) were examined. Knob density increased from ∼20 h to ∼35 h post-invasion, with significant variation among isolates. The knob density ex vivo, which was about five-fold higher than following long-term in vitro culture, started to decline within a few months of culture. Although knob diameter and height varied among isolates, we did not observe significant time-dependent variation in these dimensions.

Conclusions/Significance

The density of knobs on the P. falciparum-IE surface depends on time since invasion, but is also determined by the infecting isolate in a time-independent manner. This is the first study to quantitatively evaluate knob densities and dimensions on different P. falciparum isolates, to examine ex vivo isolates from humans, and to compare ex vivo and long-term in vitro-cultured isolates. Our findings contribute to the understanding of the interaction between P. falciparum parasites and the infected host.     

Molecular phylogeny of the cosmopolitan aquatic plant genus Limosella (Scrophulariaceae) with a particular focus on the origin of the Australasian L. curdieana

Journal of Plant Research - Limosella is a small aquatic genus of Scrophulariaceae of twelve species, of which one is distributed in northern circumpolar regions, two in southern circumpolar...     

Trade in Palm Products in North-Western South America

More than 200 scientific publications and Internet sources dealing with trade in palm products in north-western South America are reviewed. We focus on value chains, trade volumes, prices, and recent developments for some of the most important raw materials derived from native palms. Trade in palm products takes place at local, regional, national, and international levels. For local communities and individual households palm products may play a key role as the most important or only source of cash income. Most of these palm products are inadequately or not at all captured in trade statistics at the local and regional economic levels. Only products such as vegetable ivory and palm heart are monitored statistically, mainly because they are exported. Most raw materials derived from palms are extracted from the wild, and mainly by destructive harvesting. Reduced availability and rising prices on local and regional markets reflect incipient resource depletion. Only in vegetable ivory more or less sustainable wild harvesting methods prevail. Palm heart is increasingly being harvested from orchards and non-sustainable exploitation of wild populations is loosing ground. The international market for native palm oils and pulp (esp. Euterpe oleracea or açaí) is currently served almost exclusively from Brazil. Due to low oil contents and high production costs palm oils are currently used mainly for cosmetics. Based on their content of protein, starch, tocols, and carotenoids palm fruits have high nutritional value and represent a considerable potential for the development of functional foods, food supplements and animal fodder. Palms could undoubtedly play a more important role in the socio-economic development of north-western South America. Sustainability and marketing potential of palm products are negatively affected by the low income obtained by primary producers which often represents no more than 0.01–3% of the retail value. Poor governance, insecurity of land tenure and unequal sharing of profits endanger a sustainable long-term development of these valuable resources.