Site-Specific Structural Variations Accompanying Tubular Assembly of the HIV-1 Capsid Protein

The 231-residue capsid (CA) protein of human immunodeficiency virus type 1 (HIV-1) spontaneously self-assembles into tubes with a hexagonal lattice that is believed to mimic the surface lattice of conical capsid cores within intact virions. We report the results of solid-state nuclear magnetic resonance (NMR) measurements on HIV-1 CA tubes that provide new information regarding changes in molecular structure that accompany CA self-assembly, local dynamics within CA tubes, and possible mechanisms for the generation of lattice curvature. This information is contained in site-specific assignments of signals in two- and three-dimensional solid-state NMR spectra, conformation-dependent ¹⁵N and ¹³C NMR chemical shifts, detection of highly dynamic residues under solution NMR conditions, measurements of local variations in transverse spin relaxation rates of amide ¹H nuclei, and quantitative measurements of site-specific ¹⁵N–¹⁵N dipole–dipole couplings. Our data show that most of the CA sequence is conformationally ordered and relatively rigid in tubular assemblies and that structures of the N-terminal domain (NTD) and the C-terminal domain (CTD) observed in solution are largely retained. However, specific segments, including the N-terminal β-hairpin, the cyclophilin A binding loop, the inter-domain linker, segments involved in intermolecular NTD–CTD interactions, and the C-terminal tail, have substantial static or dynamical disorder in tubular assemblies. Other segments, including the 3₁₀-helical segment in CTD, undergo clear conformational changes. Structural variations associated with curvature of the CA lattice appear to be localized in the inter-domain linker and intermolecular NTD–CTD interface, while structural variations within NTD hexamers, around local 3-fold symmetry axes, and in CTD–CTD dimerization interfaces are less significant.     

Aminoadamantanes containing monoterpene-derived fragments as potent tyrosyl-DNA phosphodiesterase 1 inhibitors

The ability of a number of nitrogen-containing compounds that simultaneously carry the adamantane and monoterpene moieties to inhibit Tdp1, an important enzyme of the DNA repair system, is studied. Inhibition of this enzyme has the potential to overcome chemotherapeutic resistance of some tumor types. Compound (+)-3c synthesized from 1-aminoadamantane and (+)-myrtenal, and compound 4a produced from 2-aminoadamantane and citronellal were found to be most potent as they inhibited Tdp1 with IC₅₀ values of 6 and 3.5 µM, respectively. These compounds proved to have low cytotoxicity in colon HCT-116 and lung A-549 human tumor cell lines (CC₅₀ > 50 µM). It was demonstrated that compound 4a at 10 µM enhanced cytotoxicity of topotecan, a topoisomerase 1 poison in clinical use, against HCT-116 more than fivefold and to a lesser extent of 1.5 increase in potency for A-549.     

Functional role of phenylacetic acid from metapleural gland secretions in controlling fungal pathogens in evolutionarily derived leaf-cutting ants

Fungus-farming ant colonies vary four to five orders of magnitude in size. They employ compounds from actinomycete bacteria and exocrine glands as antimicrobial agents. Atta colonies have millions of ants and are particularly relevant for understanding hygienic strategies as they have abandoned their ancestors'' prime dependence on antibiotic-based biological control in favour of using metapleural gland (MG) chemical secretions. Atta MGs are unique in synthesizing large quantities of phenylacetic acid (PAA), a known but little investigated antimicrobial agent. We show that particularly the smallest workers greatly reduce germination rates of Escovopsis and Metarhizium spores after actively applying PAA to experimental infection targets in garden fragments and transferring the spores to the ants'' infrabuccal cavities. In vitro assays further indicated that Escovopsis strains isolated from evolutionarily derived leaf-cutting ants are less sensitive to PAA than strains from phylogenetically more basal fungus-farming ants, consistent with the dynamics of an evolutionary arms race between virulence and control for Escovopsis, but not Metarhizium. Atta ants form larger colonies with more extreme caste differentiation relative to other attines, in societies characterized by an almost complete absence of reproductive conflicts. We hypothesize that these changes are associated with unique evolutionary innovations in chemical pest management that appear robust against selection pressure for resistance by specialized mycopathogens.     

Sequences of cDNAs for human salivary and pancreatic α-amylases

The nucleotide sequences of the cloned human salivary and pancreatic α-amylase cDNAs correspond to the continuous mRNA sequences of 1768 and 1566 nucleotides, respectively. These include all of the amino acid coding regions. Salivary cDNA contains 200 bp in the 5′-noncoding region and 32 in the 3′-noncoding region. Pancreatic cDNA contains 3 and 27 bp of 5′- and 3′-noncoding regions, respectively. The nucleotide sequence humology of the two cDNAs is 96% in the coding region, and the predicted amino acid sequences are 94% homologous.Comparison of the sequences of human α-amylase cDNAs with those previously obtained for mouse α-amylase genes (Hagenbuchle et al., 1980; Schibler et al., 1982) showed the possibility of gene conversion between the two genes of human α-amylase.     

Geographic isolation and evolution of Mediterranean endemic Cyclamen: insights from chloroplast trnL (UAA) intron sequence variation

 In this study we construct a phylogenetic hypothesis for the relatedness among disjunct subspecies of Cyclamen repandum and its two allopatric congeners, C. creticum and C. balearicum in order to examine the evolutionary divergence of currently isolated populations across the western Mediterranean. The most parsimonious phylogenetic tree obtained from sequencing the cpDNA trnL (UAA) intron suggests a major phylogeographic divide in southern Greece between two clades. The first clade comprises samples of C. repandum subsp. peloponnesiacum (from the Peloponnese) and C. creticum (from Crete). The second comprises samples of C. repandum subsp. repandum (from Croatia, Italy, southern France, Corsica, Sardinia and Sicily), C. repandum subsp. rhodense (from Rhodes and Kos) and C. balearicum (from the Balearic Islands and southern France). These data suggest that C. creticum has evolved in allopatry from C. repandum subsp. peloponnesiacum and that C. balearicum and C. repandum ssp. rhodense have diverged from C. repandum subsp. repandum at its western and eastern distribution limits. At one small site on Corsica, a population of C. repandum may have introgressed with relictual populations of C. balearicum. These divergence patterns illustrate how a phylogenetic perspective can be used to better understand the evolution of endemism in the Mediterranean flora. Received February 19, 2001 Accepted August 22, 2001     

Pore-forming Activity of the Escherichia coli Type III Secretion System Protein EspD

Enterohemorrhagic Escherichia coli is a causative agent of gastrointestinal and diarrheal diseases. Pathogenesis associated with enterohemorrhagic E. coli involves direct delivery of virulence factors from the bacteria into epithelial cell cytosol via a syringe-like organelle known as the type III secretion system. The type III secretion system protein EspD is a critical factor required for formation of a translocation pore on the host cell membrane. Here, we show that recombinant EspD spontaneously integrates into large unilamellar vesicle (LUV) lipid bilayers; however, pore formation required incorporation of anionic phospholipids such as phosphatidylserine and an acidic pH. Leakage assays performed with fluorescent dextrans confirmed that EspD formed a structure with an inner diameter of ∼2.5 nm. Protease mapping indicated that the two transmembrane helical hairpin of EspD penetrated the lipid layer positioning the N- and C-terminal domains on the extralumenal surface of LUVs. Finally, a combination of glutaraldehyde cross-linking and rate zonal centrifugation suggested that EspD in LUV membranes forms an ∼280–320-kDa oligomeric structure consisting of ∼6–7 subunits.     

Retrograde Traffic Out of the Yeast Vacuole to the TGN Occurs via the Prevacuolar/Endosomal Compartment

A large number of trafficking steps occur between the last compartment of the Golgi apparatus (TGN) and the vacuole of the yeast Saccharomyces cerevisiae. To date, two intracellular routes from the TGN to the vacuole have been identified. Carboxypeptidase Y (CPY) travels through a prevacuolar/endosomal compartment (PVC), and subsequently on to the vacuole, while alkaline phosphatase (ALP) bypasses this compartment to reach the same organelle. Proteins resident to the TGN achieve their localization despite a continuous flux of traffic by continually being retrieved from the distal PVC by virtue of an aromatic amino acid–containing sorting motif. In this study we report that a hybrid protein based on ALP and containing this retrieval motif reaches the PVC not by following the CPY sorting pathway, but instead by signal-dependent retrograde transport from the vacuole, an organelle previously thought of as a terminal compartment. In addition, we show that a mutation in VAC7, a gene previously identified as being required for vacuolar inheritance, blocks this trafficking step. Finally we show that Vti1p, a v-SNARE required for the delivery of both CPY and ALP to the vacuole, uses retrograde transport out of the vacuole as part of its normal cellular itinerary.     

Mechanisms and development of self–other distinction in dyads and groups

This opinion piece offers a commentary on the four papers that address the theme of the development of self and other understanding with a view to highlighting the important contribution of developmental research to understanding of mechanisms of social cognition. We discuss potential mechanisms linking self–other distinction and empathy, implications for grouping motor, affective and cognitive domains under a single mechanism, applications of these accounts for joint action and finally consider self–other distinction in group versus dyadic settings.     

Detection of Raillietina saudiae from the domestic pigeon in Saudi Arabia through 18S and 28S rDNA genes

Raillietina saudiae is a well-studied avian gastrointestinal parasite belonging to the family Davaineidae and is the most prevalent cyclophyllid tapeworm infecting pigeon in Saudi Arabia. The present study considered as a complementary analysis of Al-Quraishy et al. (2019; Parasitol Int 71 , 59–72) with molecular studies for two ribosomal DNA genes employed for precise recognition of this Raillietina species. The annotated partial 18S and 28S rDNA gene regions were found to be 888 and 900 bp long that utilized further to elucidate their genetic relationships at species level using maximum likelihood method. The query sequence of R. saudiae is well aligned and placed within the Davaineidae family, with the same clade of all species of Raillietina that well separated from other cyclophyllidean cestodes especially taeniid and hymenolepid species. Sequence data recorded the monophyly of Raillietina species. The current phylogeny supports the usage of the partial 18S and 28S rDNA genes as reliable markers for phylogenetic reconstructions.     

Genetics and Mechanism of Resistance to Meloidogyne arenaria in Peanut Germplasm

Segregation of resistance to Meloidogyne arenaria in six BC₅F₂ peanut breeding populations was examined in greenhouse tests. Chi-square analysis indicated that segregation of resistance was consistent with resistance being conditioned by a single gene in three breeding populations (TP259-3, TP262-3, and TP271-2), whereas two resistance genes may be present in the breeding populations TP259-2, TP263-2, and TP268-3. Nematode development in clonally propagated lines of resistant individuals of TP262-3 and TP263-2 was compared to that of the susceptible cultivar Florunner. Juvenile nematodes readily penetrated roots of all peanut genotypes, but rate of development was slower (P = 0.05) in the resistant genotypes than in Florunner. Host cell necrosis indicative of a hypersensitive response was not consistently observed in resistant genotypes of either population. Three RFLP loci linked to resistance at distances of 4.2 to 11.0 centiMorgans were identified. Resistant and susceptible alleles for RFLP loci R2430E and R2545E were quite distinct and are useful for identifying individuals homozygous for resistance in segregating populations.