Oceanographic barriers,divergence, and admixture: Phylogeography and taxonomy of two putative subspecies of short‐finned pilot whale

Genomic phylogeography plays an important role in describing evolutionary processes and their geographic, ecological, or cultural drivers. These drivers are often poorly understood in marine environments, which have fewer obvious barriers to mixing than terrestrial environments. Taxonomic uncertainty of some taxa (e.g., cetaceans), due to the difficulty in obtaining morphological data, can hamper our understanding of these processes. One such taxon, the short‐finned pilot whale, is recognized as a single global species but includes at least two distinct morphological forms described from stranding and drive hunting in Japan, the “Naisa” and “Shiho” forms. Using samples (n = 735) collected throughout their global range, we examine phylogeographic patterns of divergence by comparing mitogenomes and nuclear SNP loci. Our results suggest three types within the species: an Atlantic Ocean type, a western/central Pacific and Indian Ocean (Naisa) type, and an eastern Pacific Ocean and northern Japan (Shiho) type. mtDNA control region differentiation indicates these three types form two subspecies, separated by the East Pacific Barrier: Shiho short‐finned pilot whale, in the eastern Pacific Ocean and northern Japan, and Naisa short‐finned pilot whale, throughout the remainder of the species' distribution. Our data further indicate two diverging populations within the Naisa subspecies, in the Atlantic Ocean and western/central Pacific and Indian Oceans, separated by the Benguela Barrier off South Africa. This study reveals a process of divergence and speciation within a globally‐distributed, mobile marine predator, and indicates the importance of the East Pacific Barrier to this evolutionary process.     

Structure-based reassessment of the caveolin signaling model: do caveolae regulate signaling through caveolin-protein interactions?

Caveolin proteins drive formation of caveolae, specialized cell-surface microdomains that influence cell signaling. Signaling proteins are proposed to use conserved caveolin-binding motifs (CBMs) to associate with caveolae via the caveolin scaffolding domain (CSD). However, structural and bioinformatic analyses argue against such direct physical interactions: in the majority of signaling proteins, the CBM is buried and inaccessible. Putative CBMs do not form a common structure for caveolin recognition, are not enriched among caveolin-binding proteins, and are even more common in yeast, which lack caveolae. We propose that CBM/CSD-dependent interactions are unlikely to mediate caveolar signaling, and the basis for signaling effects should therefore be reassessed.     

Sequence variation in the cytochrome oxidase I, internal transcribed spacer 1, and Ts14 diagnostic antigen sequences of Taenia solium isolates from South and Central America, India, and Asia.

We examined the genetic variability in the pig–human tapeworm, Taenia solium, by sequencing the genes for cytochrome oxidase I, internal transcribed spacer 1, and a diagnostic antigen, Ts14, from individual cysts isolated from Peru, Colombia, Mexico, India, China, and the Philippines. For these genes, the rate of nucleotide variation was minimal. Isolates from these countries can be distinguished based on one to eight nucleotide differences in the 396 nucleotide cytochrome oxidase I (COI) sequence. However, all of the 15 isolates from within Peru had identical COI sequences. The Ts14 sequences from India and China were identical and differed from the Peru sequence by three nucleotides in 333. These data indicate that there is minimal genetic variability within the species T. solium. Minimal variability was also seen in the ITS1 sequence, but this variation was observed within the individual. Twenty-two cloned sequences from six isolates sorted into 13 unique sequences. The variability observed within the sequences from individual cysts was as great as the variability between the isolates.     

Recognition of Dictyostelium discoideum lysosomal enzymes is conferred by the amino-terminal carbohydrate binding site of the insulin-like growth factor II/mannose 6-phosphate receptor

The insulin-like growth factor-II/mannose 6-phosphate receptor (IGF-II/MPR) is a type I glycoprotein that mediates both the intracellular sorting of lysosomal enzymes bearing mannose 6-phosphate (Man-6-P) residues to the lysosome and the bioavailability of IGF-II. The extracytoplasmic region of the IGF-II/MPR contains 15 repeating domains; the two carbohydrate recognition domains (CRDs) have been localized to domains 1-3 and 7-9, and the high-affinity IGF-II binding site maps to domain 11. To characterize the carbohydrate binding properties of the IGF-II/MPR, regions of the receptor encompassing the individual CRDs were produced in a baculovirus expression system. Characterization of the recombinant proteins revealed that the pH optimum for carbohydrate binding is significantly more acidic for the carboxyl-terminal CRD than for the amino-terminal CRD (i.e., pH 6.4-6.5 vs 6.9). Equilibrium binding studies demonstrated that the two CRDs exhibit a similar affinity for Man-6-P. Furthermore, substitution of the conserved arginine residue in domain 3 (R435) or in domain 9 (R1334) with alanine resulted in a similar >1000-fold decrease in the affinity for the lysosomal enzyme, beta-glucuronidase. In contrast, the two CRDs differ dramatically in their ability to recognize the distinctive modifications (i.e., mannose 6-sulfate and Man-6-P methyl ester) found on Dictyostelium discoideum lysosomal enzymes: the amino-terminal CRD binds mannose 6-sulfate and Man-6-P methyl ester with a 14-55-fold higher affinity than the carboxyl-terminal CRD. Taken together, these results demonstrate that the IGF-II/MPR contains two functionally distinct CRDs.     

In planta conditions induce genomic changes in Pseudomonas syringae pv. phaseolicola

The co-evolution of bacterial plant pathogens and their hosts is a complex and dynamic process. Plant resistance can impose stress on invading pathogens that can lead to, and select for, beneficial changes in the bacterial genome. The Pseudomonas syringae pv. phaseolicola (Pph) genomic island PPHGI-1 carries an effector gene, avrPphB (hopAR1), which triggers the hypersensitive reaction in bean plants carrying the R3 resistance gene. Interaction between avrPphB and R3 generates an antimicrobial environment within the plant, resulting in the excision of PPHGI-1 and its loss from the genome. The loss of PPHGI-1 leads to the generation of a Pph strain able to cause disease in the plant. In this study, we observed that lower bacterial densities inoculated into resistant bean (Phaseolus vulgaris) plants resulted in quicker PPHGI-1 loss from the population, and that loss of the island was strongly influenced by the type of plant resistance encountered by the bacteria. In addition, we found that a number of changes occurred in the bacterial genome during growth in the plant, whether or not PPHGI-1 was lost. We also present evidence that the circular PPHGI-1 episome is able to replicate autonomously when excised from the genome. These results shed more light onto the plasticity of the bacterial genome as it is influenced by in planta conditions.     

In vitro SAR of pyrrolidine-containing histamine H3 receptor antagonists: trends across multiple chemical series

Structure-activity relationships (SAR) were analyzed within a library of diverse yet simple compounds prepared as histamine H3 antagonists. The libraries were constructed with a variety of low molecular weight pyrrolidines, selected from (R)-2-methylpyrrolidine, (S)-2-methylpyrrolidine, and pyrrolidine.     

Characterization and sequence analyses of antibody-selected antigenic variants of herpes simplex virus show a conformationally complex epitope on glycoprotein H.

Thirteen antigenic variants of herpes simplex virus which were resistant to neutralization by monoclonal antibody 52S or LP11 were isolated and characterized. The antibodies in the absence of complement potently neutralize infectivity of wild-type virus as well as inhibit the transfer of virus from infected to uninfected cells ("plaque inhibition") and decrease virus-induced cell fusion by syncytial strains. The first variant isolated arose in vivo. Of 66 type 1 isolates analyzed from typing studies of 100 clinical isolates, one was identified as resistant to neutralization by LP11 antibody. The glycoprotein H (gH) sequence was derived and compared with those of wild-type and syncytial laboratory strains SC16, strain 17, and HFEM. The sequences were highly conserved in contrast to the diversity observed between gH sequences from herpesviruses of different subgroups. Only four coding changes were present in any of the comparisons, and only one unique coding change was observed between the laboratory strains and the clinical isolate (Asp-168 to Gly). These sequences were compared with those of antigenic variants selected by antibody in tissue culture. Twelve variants were independently selected with antibody LP11 or 52S from parent strain SC16 or HFEM. For each variant, the gH nucleotide sequence was derived and a point mutation was identified giving rise to a single amino acid substitution. The LP11-resistant viruses encoded gH sequences with amino acid substitutions at sites distributed over one-half of the gH external domain, Glu-86, Asp-168, or Arg-329, while the 52S-resistant mutant viruses had substitutions at adjacent positions Ser-536 and Ala-537. One LP11 mutant virus had a point mutation in the gH gene that was identical to that of the clinical isolate, giving rise to a substitution of Asp-168 with Gly. Both LP11 and 52S appeared to recognize distinct gH epitopes as mutant virus resistant to neutralization and immunoprecipitation with LP11 remained sensitive to 52S and the converse was shown for the 52S-resistant mutant virus. This is consistent with previous studies which showed that while the 52S epitope could be formed in the absence of other virus products, virus gene expression was required for stable presentation of the LP11 epitope, and for transport of gH to the cell surface (Gompels and Minson, J. Virol. 63:4744-4755, 1989). All mutant viruses produced numbers of infectious particles that were similar to those produced by the wild-type virus, with the exception of one variant which produced lower yields.(ABSTRACT TRUNCATED AT 400 WORDS)     

Outer membrane protein P of Pseudomonas aeruginosa: regulation by phosphate deficiency and formation of small anion-specific channels in lipid bilayer membranes.

A new major outer membrane protein, P, was induced in Pseudomonas aeruginosa PAO1 upon growth in medium containing 0.2 mM or less inorganic phosphate. Studies with media containing different levels of phosphate and with mutants of PAO1 suggested that protein P was coregulated with alkaline phosphatase and phospholipase C. Protein P was substantially purified and shown to form sodium dodecyl sulfate-resistant oligomers on polyacrylamide gels. The incorporation of purified protein P into artificial lipid bilayers resulted in an increase of the membrane conductance by many orders of magnitude. Single-channel experiments demonstrated that protein P channels were substantially smaller than all previously studied porins from P. aeruginosa and enteric bacteria, with an average single-channel conductance in 1 M NaCl of 0.25 nS. The protein P channel was apparently not voltage induced or regulated. The results of single-channel conductance experiments, using a variety of different salts, allowed a minimum channel diameter estimate of 0.7 nm. Furthermore, from these results it was concluded that the protein P channel was highly specific for anions. Zero-current potential measurements confirmed that protein P was at least 30-fold more permeable for Cl- than for K+ ions. The possible biological role of the small, anion-specific protein P channels in phosphate uptake from the medium is discussed.     

Interleukin 2 receptors are expressed by alveolar macrophages during pulmonary sarcoidosis and are inducible by lymphokine treatment of normal human lung macrophages, blood monocytes, and monocyte cell lines

Expression of receptors for IL 2 was believed initially to be restricted to T cells after their activation by IL 1 and antigen. However, recently IL 2 receptors (IL 2R) were demonstrated on activated B cells by using an anti-IL 2R monoclonal antibody (anti-Tac). In this study, we examined the capacity of cultured human alveolar macrophages, blood monocytes, and myelomonocytic (HL-60) or monoblast (U937) cell lines to bind three different anti-IL 2R monoclonal antibodies before or after stimulation with the monocyte-activating agents IFN-gamma, LPS, phorbol ester, or lymphokine-containing conditioned medium. For each of the four cell populations examined, resting unstimulated cells bound little or no anti-IL 2R antibody, as shown independently by quantitative cell binding assay and by immunoperoxidase labeling. By contrast, incubation with recombinant IFN-gamma, conditioned medium, or to a lesser extent, native or recombinant IL 2 itself, resulted in a significant enhancement of anti-IL 2 receptor monoclonal antibody binding by all four populations, whereas LPS, PMA, or IL 1 had no effect. In addition, membrane binding of anti-Tac antibody, similar to that seen after stimulation of normal lung macrophages with IFN-gamma, was detected by using macrophages obtained by bronchoalveolar lavage of five patients with active pulmonary sarcoidosis. These findings are consistent with the expression of a functional IL 2R on activated cells of the monocyte lineage, since anti-Tac binding to IFN-gamma-treated HL-60 cells was inhibited by addition of excess IL-2; specific binding of anti-IL 2 monoclonal antibodies was detected in the presence of exogenous IL 2; and a 50 to 55 kD molecule was immunoprecipitated from both activated lung macrophages and T lymphoblasts by using anti-Tac antibody. We conclude that human mononuclear phagocytes can be induced by lymphokines to express IL 2R, and that such IL 2R+ macrophages can be detected in vivo during inflammation.