Exploring ORFan Domains in Giant Viruses: Structure of Mimivirus Sulfhydryl Oxidase R596

The mimivirus genome contains many genes that lack homologs in the sequence database and are thus known as ORFans. In addition, mimivirus genes that encode proteins belonging to known fold families are in some cases fused to domain-sized segments that cannot be classified. One such ORFan region is present in the mimivirus enzyme R596, a member of the Erv family of sulfhydryl oxidases. We determined the structure of a variant of full-length R596 and observed that the carboxy-terminal region of R596 assumes a folded, compact domain, demonstrating that these ORFan segments can be stable structural units. Moreover, the R596 ORFan domain fold is novel, hinting at the potential wealth of protein structural innovation yet to be discovered in large double-stranded DNA viruses. In the context of the R596 dimer, the ORFan domain contributes to formation of a broad cleft enriched with exposed aromatic groups and basic side chains, which may function in binding target proteins or localization of the enzyme within the virus factory or virions. Finally, we find evidence for an intermolecular dithiol/disulfide relay within the mimivirus R596 dimer, the first such extended, intersubunit redox-active site identified in a viral sulfhydryl oxidase.     

Biosynthetic incorporation of unnatural sialic acids into polysialic acid on neural cells

In this study we demonstrate that polysialyltransferases are capable of accepting unnatural substrates in terminally differentiated human neurons. Polysialyltransferases catalyze the glycosylation of the neural cell adhesion molecule (NCAM) with polysialic acid (PSA). The unnatural sialic acid analog, N-levulinoyl sialic acid (SiaLev), was incorporated into cell surface glycoconjugates including PSA by the incubation of cultured neurons with the metabolic precursor N-levulinoylmannosamine (ManLev). The ketone group within the levulinoyl side chain of SiaLev was then used as a chemical handle for detection using a biotin probe. The incorporation of SiaLev residues into PSA was demonstrated by protection from sialidases that can cleave natural sialic acids but not those bearing unnatural N-acyl groups. The presence of SiaLev groups on the neuronal cell surface did not impede neurite outgrowth or significantly affect the distribution of PSA on neuronal compartments. Since PSA is important in neural plasticity and development, this mechanism for modulating PSA structure might be useful for functional studies.     

The relationship between intra–guild diet overlap and breeding in owls in Israel

Even though intra-guild predators frequently prey on the same species, it is unclear whether diet overlap between two predators is a source of interspecific competition or whether predators simply use the same abundant prey resource. We measured the extent to which the diets of barn owls (Tyto alba) and long-eared owls (Asio otus) in Israel overlap and examined whether yearly differences in diet overlap correlate with barn owl breeding success. Pianka’s index of niche overlap was positively related to barn owl population size but not to its breeding success. The number of breeding barn owls was higher when long-eared owls consumed more rodents, suggesting that diet overlap most likely increased when rodents became more abundant. Therefore, in Israel, when these two owl species prey more often on rodents, their diets are more similar and interspecific competition is reduced. Unlike sympatric populations in Europe, in years when rodents are less abundant in Israel long-eared owls switch to hunting alternative prey (e.g., birds), perhaps to avoid competition with barn owls.     

Screening marine fungi for inhibitors of the C4 plant enzyme pyruvate phosphate dikinase: unguinol as a potential novel herbicide candidate

A total of 2,245 extracts, derived from 449 marine fungi cultivated in five types of media, were screened against the C(4) plant enzyme pyruvate phosphate dikinase (PPDK), a potential herbicide target. Extracts from several fungal isolates selectively inhibited PPDK. Bioassay-guided fractionation of one isolate led to the isolation of the known compound unguinol, which inhibited PPDK with a 50% inhibitory concentration of 42.3 +/- 0.8 muM. Further kinetic analysis revealed that unguinol was a mixed noncompetitive inhibitor of PPDK with respect to the substrates pyruvate and ATP and an uncompetitive inhibitor of PPDK with respect to phosphate. Unguinol had deleterious effects on a model C(4) plant but no effect on a model C(3) plant. These results indicate that unguinol inhibits PPDK via a novel mechanism of action which also translates to an herbicidal effect on whole plants.     

Retinoic acid induces neuronal differentiation of embryonal carcinoma cells by reducing proteasome-dependent proteolysis of the cyclin-dependent inhibitor p27.

Retinoic acid (RA) treatment of embryonal carcinoma cell line NTERA-2 clone D1 (NT2/D1) induces growth arrest and terminal differentiation along the neuronal pathway. In the present study, we provide a functional link between RA and p27 function in the control of neuronal differentiation in NT2/D1 cells. We report that RA enhances p27 expression, which results in increased association with cyclin E/cyclin-dependent kinase 2 complexes and suppression of their activity; however, antisense clones, which have greatly reduced RA-dependent p27 inducibility (NT2-p27AS), continue to synthesize DNA and are unable to differentiate properly in response to RA as determined by lack of neurite outgrowth and by the failure to modify surface antigens. As to the mechanism involved in RA-dependent p27 upregulation, our data support the concept that RA reduces p27 protein degradation through the ubiquitin/proteasome-dependent pathway. Taken together, these findings demonstrate that in embryonal carcinoma cells, p27 expression is required for growth arrest and proper neuronal differentiation.     

The importance of micro-habitat in the breeding of Barn Owls Tyto alba

Capsule?Habitat parameters associated with 706 Barn Owl (Tyto alba) nesting boxes in Israel were analysed. Pairs bred in 259 of the boxes. The intensity of agricultural practices at nestbox sites were shown to have only a weak effect on aspects of Barn Owl breeding in this region.     

Molecular characterization of the Pseudomonas aeruginosa serotype O5 (PAO1) B-band lipopolysaccharide gene cluster

Pseudomonas aeruginosa co-expresses A-band lipopolysaccharide (LPS), a homopolymer of rhamnose, and B-band LPS, a heteropolymer with a repeating unit of 2–5 sugars which is the serotype-specific antigen. The gene clusters for A- and B-band biosynthesis in P. aeruginosa O5 (strain PAO1) have been cloned previously. Here we report the DNA sequence and molecular analysis of the B-band O-antigen biosynthetic cluster. Sixteen open reading frames (ORFs) thought to be involved in synthesis of the O5 O antigen were identified, including wzz ( rol ), wzy ( rfc ), and wbpA – wbpN . A further 3 ORFs not thought to be involved with LPS synthesis were identified ( hisH , hisF , and uvrB ). Most of the wbp genes are found only in serotypes O2, O5, O16, O18, and O20, which form a chemically and structurally related O-antigen serogroup. In contrast, wbpM and wbpN are common to all 20 serotypes of P. aeruginosa. Although wbpM is not serogroup-specific, knockout mutations confirmed it is necessary for O5 O-antigen biosynthesis. A novel insertion sequence, IS 1209 , is present at the junction between the serogroup-specific and non-specific regions. We have predicted the functions of the proteins encoded in the wbp cluster based on their homologies to those in the databases, and provide a proposed pathway of P. aeruginosa O5 O-antigen biosynthesis.