Conservation of the Native Orchids Through Seedling Culture and Reintroduction—A Singapore Experience

Singapore is located near the equator, off the southern tip of the Malay Peninsula. The whole country consists mostly of lowland. It has many interesting types of natural habitats such as primary rain forest, freshwater swamp forest, mangroves, secondary forests, shrub, grasslands, and urban parks and fields. The climate is equatorial with relatively uniform temperature and high humidity. Unfortunately, many of the natural habitats and the native orchids which thrive there have disappeared due to habitat destruction. Some 226 species of native orchids have been recorded in Singapore. However, of these 178 are considered to be extinct, and only five are common. The orchid conservation programme aims to monitor existing species, explore ways to conserve their germplasm, and increase their numbers in natural, semi-natural, and urban environments through ex-situ seedling culture and subsequent re-introduction into appropriate habitats, including roadside trees, parks and natural areas. In the first phase of the programme, we have successfully propagated and carried out experiments of re-introduction on five species of native orchids, namely, Grammatophyllum speciosum, Bulbophyllum vaginatum, Bulbophyllum membranaceum, Cymbidium finlaysonianum and Cymbidium bicolor. Survival percentages 8-yr after the reintroduction events ranged from 10 to 95 for G. speciosum, the target species of the earliest re-introduction experiments. Size of the seedlings at reintroduction, host trees, and relative humidity seemed to play significant roles in the success rate of the reintroductions.     

QTL mapping and phenotypic variation for root architectural traits in maize (Zea mays L.)

Key message

QTL were identified for root architectural traits in maize.

Abstract

Root architectural traits, including the number, length, orientation, and branching of the principal root classes, influence plant function by determining the spatial and temporal domains of soil exploration. To characterize phenotypic patterns and their genetic control, three recombinant inbred populations of maize were grown for 28 days in solid media in a greenhouse and evaluated for 21 root architectural traits, including length, number, diameter, and branching of seminal, primary and nodal roots, dry weight of embryonic and nodal systems, and diameter of the nodal root system. Significant phenotypic variation was observed for all traits. Strong correlations were observed among traits in the same root class, particularly for the length of the main root axis and the length of lateral roots. In a principal component analysis, relationships among traits differed slightly for the three families, though vectors grouped together for traits within a given root class, indicating opportunities for more efficient phenotyping. Allometric analysis showed that trajectories of growth for specific traits differ in the three populations. In total, 15 quantitative trait loci (QTL) were identified. QTL are reported for length in multiple root classes, diameter and number of seminal roots, and dry weight of the embryonic and nodal root systems. Phenotypic variation explained by individual QTL ranged from 0.44 % (number of seminal roots, NyH population) to 13.5 % (shoot dry weight, OhW population). Identification of QTL for root architectural traits may be useful for developing genotypes that are better suited to specific soil environments.     

Virus-Mediated shRNA Knockdown of Prodynorphin in the Rat Nucleus Accumbens Attenuates Depression-Like Behavior and Cocaine Locomotor Sensitization

Dynorphins, endogenous opioid peptides that arise from the precursor protein prodynorphin (Pdyn), are hypothesized to be involved in the regulation of mood states and the neuroplasticity associated with addiction. The current study tested the hypothesis that dynorphin in the nucleus accumbens (NAcc) mediates such effects. More specifically, we examined whether knockdown of Pdyn within the NAcc in rats would alter the expression of depressive-like and anxiety-like behavior, as well as cocaine locomotor sensitization. Wistar rats were injected with adeno-associated viral (AAV) vectors encoding either a Pdyn-specific short hairpin RNA (AAV-shPdyn) or a scrambled shRNA (AAV-shScr) as control. Four weeks later, rats were tested for anxiety-like behavior in the elevated plus maze test and depressive-like behavior in the forced swim test (FST). Finally, rats received one daily injection of saline or cocaine (20 mg/kg, i.p.), followed by assessment of locomotion for 4 consecutive days. Following 3 days of abstinence, the rats completed 2 additional daily cocaine/saline locomotor trials. Pdyn knockdown in the NAcc led to a significant reduction in depressive-like behavior in the FST, but had no effect on anxiety-like behavior in the elevated plus maze. Pdyn knockdown did not alter baseline locomotor behavior, the locomotor response to acute cocaine, or the initial sensitization of the locomotor response to cocaine over the first 4 cocaine treatment days. However, following 3 days abstinence the locomotor response to the cocaine challenge returned to their original levels in the AAV-shPdyn rats while remaining heightened in the AAV-shScr rats. These results suggest that dynorphin in a very specific area of the nucleus accumbens contributes to depressive-like states and may be involved in neuroadaptations in the NAcc that contribute to the development of cocaine addiction as a persistent and lasting condition.     

Degradation of cyclin A does not require its phosphorylation by CDC2 and cyclin-dependent kinase 2

Many cyclins are degraded by the ubiquitination/proteasome pathways involving the anaphase-promoting complex and SCF complexes. These degradations are frequently dependent on phosphorylation by cyclin-dependent kinases (CDKs), providing a self-limiting mechanism for CDK activity. Here we present evidence from in vitro and in vivo assay systems that the degradation of human cyclin A can be inhibited by kinase-inactive mutants of CDK2 and CDC2. One obvious interpretation of these results is that like other cyclins, CDK-dependent phosphorylation of the cyclin A may be involved in cyclin A degradation. Our data indicated that CDK2 can phosphorylate cyclin A on Ser-154. Site-directed mutagenesis of Ser-154 abolished the phosphorylation by recombinant CDK2 in vitro and the majority of cyclin A phosphorylation in the cell. Activation of CDK2 and binding to SKP2 or p27(KIP1) were not affected by the phosphorylation of Ser-154. Surprising, in marked contrast to cyclin E, where phosphorylation of Thr-380 by CDK2 is required for proteolysis, degradation of cyclin A was not affected by Ser-154 phosphorylation. It is likely that the stabilization of cyclin A by the kinase-inactive CDKs was mainly due to a cell cycle effect. These data suggest an important difference between the regulation of cyclin A and cyclin E.     

Full engagement of liganded maltose‐binding protein stabilizes a semi‐open ATP‐binding cassette dimer in the maltose transporter

MalFGK₂ is an ATP‐binding cassette (ABC) transporter that mediates the uptake of maltose/maltodextrins into Escherichia coli. A periplasmic maltose‐binding protein (MBP) delivers maltose to the transmembrane subunits (MalFG) and stimulates the ATPase activity of the cytoplasmic nucleotide‐binding subunits (MalK dimer). This MBP‐stimulated ATPase activity is independent of maltose for purified transporter in detergent micelles. However, when the transporter is reconstituted in membrane bilayers, only the liganded form of MBP efficiently stimulates its activity. To investigate the mechanism of maltose stimulation, electron paramagnetic resonance spectroscopy was used to study the interactions between the transporter and MBP in nanodiscs and in detergent. We found that full engagement of both lobes of maltose‐bound MBP unto MalFGK₂ is facilitated by nucleotides and stabilizes a semi‐open MalK dimer. Maltose‐bound MBP promotes the transition to the semi‐open state of MalK when the transporter is in the membrane, whereas such regulation does not require maltose in detergent. We suggest that stabilization of the semi‐open MalK₂ conformation by maltose‐bound MBP is key to the coupling of maltose transport to ATP hydrolysis in vivo, because it facilitates the progression of the MalK dimer from the open to the semi‐open conformation, from which it can proceed to hydrolyze ATP.     

Thrombospondin-1 interacts with Trypanosoma cruzi surface calreticulin to enhance cellular infection

Trypanosoma cruzi causes Chagas disease, which is a neglected tropical disease that produces severe pathology and mortality. The mechanisms by which the parasite invades cells are not well elucidated. We recently reported that T. cruzi up-regulates the expression of thrombospondin-1 (TSP-1) to enhance the process of cellular invasion. Here we characterize a novel TSP-1 interaction with T. cruzi that enhances cellular infection. We show that labeled TSP-1 interacts specifically with the surface of T. cruzi trypomastigotes. We used TSP-1 to pull down interacting parasite surface proteins that were identified by mass spectrometry. We also show that full length TSP-1 and the N-terminal domain of TSP-1 (NTSP) interact with T. cruzi surface calreticulin (TcCRT) and other surface proteins. Pre-exposure of recombinant NTSP or TSP-1 to T. cruzi significantly enhances cellular infection of wild type mouse embryo fibroblasts (MEF) compared to the C-terminal domain of TSP-1, E3T3C1. In addition, blocking TcCRT with antibodies significantly inhibits the enhancement of cellular infection mediated by the TcCRT-TSP-1 interaction. Taken together, our findings indicate that TSP-1 interacts with TcCRT on the surface of T. cruzi through the NTSP domain and that this interaction enhances cellular infection. Thus surface TcCRT is a virulent factor that enhances the pathogenesis of T. cruzi infection through TSP-1, which is up-regulated by the parasite.     

Germination niches and seed persistence of tropical epiphytic orchids in an urban landscape

Journal of Plant Research - Urbanisation has contributed to significant biodiversity loss, yet, urban areas can facilitate biodiversity conservation. For instance, there is evidence of urban trees...     

Extraepitopic compensatory substitutions partially restore fitness to simian immunodeficiency virus variants that escape from an immunodominant cytotoxic-T-lymphocyte response

Selection for escape mutant immunodeficiency viruses by cytotoxic T lymphocytes (CTL) has been well characterized and may be associated with disease progression. CTL epitopes accrue escape mutations at different rates in vivo. Interestingly, certain high-frequency CTL do not select for escape until the chronic phase of infection. Here we show that mutations conferring escape from immunodominant CTL directed against an epitope in the viral Gag protein are strongly associated with extraepitopic mutations in gag in vivo. The extraepitopic mutations partially restore in vitro replicative fitness of viruses bearing the escape mutations. Constraints on epitope sequences may therefore play a role in determining the rate of escape from CTL responses in vivo.