Drosophila rolling blackout displays lipase domain-dependent and -independent endocytic functions downstream of dynamin

Drosophila temperature-sensitive rolling blackout (rbo(ts) ) mutants display a total block of endocytosis in non-neuronal cells and a weaker, partial defect at neuronal synapses. RBO is an integral plasma membrane protein and is predicted to be a serine esterase. To determine if lipase activity is required for RBO function, we mutated the catalytic serine 358 to alanine in the G-X-S-X-G active site, and assayed genomic rescue of rbo mutant non-neuronal and neuronal phenotypes. The rbo(S358A) mutant is unable to rescue rbo null 100% embryonic lethality, indicating that the lipase domain is critical for RBO essential function. Likewise, the rbo(S358A) mutant cannot provide any rescue of endocytic blockade in rbo(ts) Garland cells, showing that the lipase domain is indispensable for non-neuronal endocytosis. In contrast, rbo(ts) conditional paralysis, synaptic transmission block and synapse endocytic defects are all fully rescued by the rbo(S358A) mutant, showing that the RBO lipase domain is dispensable in neuronal contexts. We identified a synthetic lethal interaction between rbo(ts) and the well-characterized dynamin GTPase conditional shibire (shi(ts1)) mutant. In both non-neuronal cells and neuronal synapses, shi(ts1); rbo(ts) phenocopies shi(ts1) endocytic defects, indicating that dynamin and RBO act in the same pathway, with dynamin functioning upstream of RBO. We conclude that RBO possesses both lipase domain-dependent and scaffolding functions with differential requirements in non-neuronal versus neuronal endocytosis mechanisms downstream of dynamin GTPase activity.     

Global Drivers and Tradeoffs of Three Urban Vegetation Ecosystem Services

Our world is increasingly urbanizing which is highlighting that sustainable cities are essential for maintaining human well-being. This research is one of the first attempts to globally synthesize the effects of urbanization on ecosystem services and how these relate to governance, social development and climate. Three urban vegetation ecosystem services (carbon storage, recreation potential and habitat potential) were quantified for a selection of a hundred cities. Estimates of ecosystem services were obtained from the analysis of satellite imagery and the use of well-known carbon and structural habitat models. We found relationships between ecosystem services, social development, climate and governance, however these varied according to the service studied. Recreation potential was positively related to democracy and negatively related to population. Carbon storage was weakly related to temperature and democracy, while habitat potential was negatively related to democracy. We found that cities under 1 million inhabitants tended to have higher levels of recreation potential than larger cities and that democratic countries have higher recreation potential, especially if located in a continental climate. Carbon storage was higher in full democracies, especially in a continental climate, while habitat potential tended to be higher in authoritarian and hybrid regimes. Similar to other regional or city studies we found that the combination of environment conditions, socioeconomics, demographics and politics determines the provision of ecosystem services. Results from this study showed the existence of environmental injustice in the developing world.     

Characterization of Influenza Virus Neuraminidases: Peptide Changes Associated with Antigenic Divergence Between Early and Late N2 Neuraminidases

Neuraminidases (EC 3.2.1.18) of 1957, 1960, and 1969 influenza virus strains were isolated after proteolytic digestion of viral hemagglutinin. Each neuraminidase was recovered with a final yield of about 15% and had similar specific activities. Immunization of rabbits with the neuraminidases elicited monospecific neuraminidase antibodies, with no antibodies to viral hemagglutinin. Further evidence of purity was the existence of only a single component, about 50,000 daltons in size, when reduced neuraminidase preparations were examined by sodium dodecyl sulfate acrylamide gel electrophoresis. However, storage of neuraminidase in solution resulted in the appearance of slightly smaller degradation products. Preparations of each neuraminidase were denatured under reducing conditions, and exposed sulfhydryl residues were blocked by reaction with (14)C-iodoacetamide. After tryptic digestion, peptide maps were prepared for the neuraminidases, and the (14)C-labeled cysteinyl peptides were then identified by autoradiography. About 20 peptides were present, in agreement with the number predicted from amino acid analysis for neuraminidase subunits of only one type. The 1957 and 1960 neuraminidases exhibited a small antigenic divergence from each other, and maps of their cysteinyl peptides appeared to be identical. The 1969 neuraminidase exhibited considerable antigenic divergence from the other two neuraminidases, and maps of 1969 neuraminidase peptides revealed two major and several minor differences from the other maps. Thus, antigenic divergence between the neuraminidases of Asian and Hong Kong influenza viruses is associated with a small number of changes in the primary structure of the neuraminidase subunit.     

Functional and regulatory analysis of the <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> CAX2 cation transporter

The vacuolar sequestration of metals is an important metal tolerance mechanism in plants. The Arabidopsis thaliana vacuolar transporters CAX1 and CAX2 were originally identified in a Saccharomyces cerevisiae suppression screen as Ca²⁺/H⁺ antiporters. CAX2 has a low affinity for Ca²⁺ but can transport other metals including Mn²⁺ and Cd²⁺. Here we demonstrate that unlike cax1 mutants, CAX2 insertional mutants caused no discernable morphological phenotypes or alterations in Ca²⁺/H⁺ antiport activity. However, cax2 lines exhibited a reduction in vacuolar Mn²⁺/H⁺ antiport and, like cax1 mutants, reduced V-type H⁺-ATPase (V-ATPase) activity. Analysis of a CAX2 promoter -glucoronidase (GUS) reporter gene fusion confirmed that CAX2 was expressed throughout the plant and strongly expressed in flower tissue, vascular tissue and in the apical meristem of young plants. Heterologous expression in yeast identified an N-terminal regulatory region in CAX2, suggesting that Arabidopsis contains multiple cation/H⁺ antiporters with shared regulatory features. Furthermore, despite significant variations in morphological and biochemical phenotypes, cax1 and cax2 lines both significantly alter V-ATPase activity, hinting at coordinate regulation among transporters driven by H⁺ gradients and the V-ATPase.     

Neophilia, innovation and social learning: a study of intergeneric differences in callitrichid monkeys

In a comparative study of neophilia, innovation and social attentiveness we exposed individuals in seven callitrichid species, from three genera, to novel extractive foraging tasks. The results revealed consistently shorter response latencies, higher levels of successful and unsuccessful manipulation, and greater attentiveness to the task and to conspecifics inLeontopithecus (lion tamarins) than in both Saguinus (tamarins) and Callithrix (marmosets). This is consistent with the hypothesis that species dependent upon manipulative and explorative foraging tend to be less neophobic and more innovative than other species. Furthermore, Callithrix appeared to be less neophobic than Saguinus; ifCallithrix is regarded as the greater specialist, this result is inconsistent with the hypothesis that neophobia is associated with foraging specialization. We consider the relevance of our findings to taxonomic relationships, and to technical and Machiavellian intelligence hypotheses and discuss the implications for captive breeding and reintroduction strategies.Copyright 2003 Published by Elsevier Science Ltd on behalf of The Association for the Study of Animal Behaviour.      

Don't shoot the (second) messenger: endomembrane transporters and binding proteins modulate cytosolic Ca2+ levels

Ca(2+) signal transduction requires the meticulous regulation of cytosolic Ca(2+) levels. Endomembrane Ca(2+) transporters and binding proteins are important components in partitioning these Ca(2+) signals to mediate cellular activity. Recently, many of these proteins have been characterized and mutant analysis suggests that these transporters form a network. Future attempts to manipulate plant Ca(2+) signaling must address all aspects of this complex.     

Insertional mutants: a foundation for assessing gene function

The abundance of Arabidopsis insert mutants portends the day when null alleles in every gene will be obtained. Once these are created, all plant scientists can become geneticists. However, this brief technical highlight of genetic concepts cautions against ascribing gene function based exclusively on phenotypic analysis of null alleles.     

Mechanism of N-terminal autoinhibition in the Arabidopsis Ca(2+)/H(+) antiporter CAX1

Regulation of Ca(2+)/H(+) antiporters may be an important function in determining the duration and amplitude of cytosolic Ca(2+) oscillations. Previously the Arabidopsis Ca(2+)/H(+) transporter, CAX1 (cation exchanger 1), was identified by its ability to suppress yeast mutants defective in vacuolar Ca(2+) transport. Recently, a 36-amino acid N-terminal regulatory region on CAX1 has been identified that inhibits CAX1-mediated Ca(2+)/H(+) antiport. Here we show that a synthetic peptide designed against the CAX1 36 amino acids inhibited Ca(2+)/H(+) transport mediated by an N-terminal-truncated CAX1 but did not inhibit Ca(2+) transport by other Ca(2+)/H(+) antiporters. Ca(2+)/H(+) antiport activity measured from vacuolar-enriched membranes of Arabidopsis root was also inhibited by the CAX1 peptide. Through analyzing CAX chimeric constructs the region of interaction of the N-terminal regulatory region was mapped to include 7 amino acids (residues 56-62) within CAX1. The CAX1 N-terminal regulatory region was shown to physically interact with this 7-amino acid region by yeast two-hybrid analysis. Mutagenesis of amino acids within the N-terminal regulatory region implicated several residues as being essential for regulation. These findings describe a unique mode of antiporter autoinhibition and demonstrate the first detailed mechanisms for the regulation of a Ca(2+)/H(+) antiporter from any organism.