Structural congruence among membrane-active host defense polypeptides of diverse phylogeny

A requisite for efficacious host defense against pathogens and predators has prioritized evolution of effector molecules thereof. A recent multidimensional analysis of physicochemical properties revealed a novel, unifying structural signature among virtually all classes of cysteine-containing antimicrobial peptides. This motif, termed the gamma-core, is seen in host defense peptides from organisms spanning more than 2.6 billion years of evolution. Interestingly, many toxins possess the gamma-core signature, consistent with discoveries of their direct antimicrobial activity. Many microbicidal chemokines (kinocidins) likewise contain iterations of the gamma-core motif, reconciling their antimicrobial efficacy. Importantly, these polypeptide classes have evolved to target and modulate biomembranes in protecting respective hosts against unfavorable interactions with potential pathogens or predators. Extending on this concept, the current report addresses the hypothesis that antimicrobial peptides, kinocidins, and polypeptide toxins are structurally congruent and share a remarkably close phylogenetic relationship, paralleling their roles in host-pathogen relationships. Analyses of their mature amino acid sequences demonstrated that cysteine-stabilized antimicrobial peptides, kinocidins, and toxins share ancient evolutionary relatedness stemming from early precursors of the gamma-core signature. Moreover, comparative 3-D structure analysis revealed recurring iterations of antimicrobial peptide gamma-core motifs within kinocidins and toxins. However, despite such congruence in gamma-core motifs, the kinocidins diverged in overall homology from microbicidal peptides or toxins. These findings are consistent with observations that chemokines are not toxic to mammalian cells, in contrast to many antimicrobial peptides and toxins. Thus, specific functions of these molecular effectors may be governed by specific configurations of structural modules associated with a common gamma-core motif. These concepts are consistent with the hypothesis that the gamma-core is an archetype determinant in polypeptides that target or regulate with biological membranes, with specific iterations optimized to unique or cognate host defense contexts. Quantitative and qualitative data suggest these protein families emerged through both parallel and divergent processes of modular evolution. Taken together, the current and prior findings imply that the gamma-core motif contributes to conserved structures and functions of host defense polypeptides. The presence of this unifying molecular signature in otherwise diverse categories of membrane-active host defense peptides implies an ancient and essential role for such a motif in effector molecules governing host-pathogen relationships.     

Selective cytotoxic T-lymphocyte targeting of tumor immune escape variants

Defects in major histocompatibility complex (MHC) class I-restricted antigen presentation are frequently observed in human cancers and result in escape of tumors from cytotoxic T lymphocyte (CTL) immune surveillance in mice. Here, we show the existence of a unique category of CTLs that can prevent this escape. The CTLs target an alternative repertoire of peptide epitopes that emerge in MHC class I at the surface of cells with impaired function of transporter associated with antigen processing (TAP), tapasin or the proteasome. These peptides, although derived from self antigens such as the commonly expressed Lass5 protein (also known as Trh4), are not presented by normal cells. This explains why they act as immunogenic neoantigens. The newly discovered epitopes can be exploited for immune intervention against processing-deficient tumors through adoptive T-cell transfer or peptide vaccination.     

Phylogenetic relationships among the species of the genus Testudo (Testudines: Testudinidae) inferred from mitochondrial 12S rRNA gene sequences

To test phylogenetic relationships within the genus Testudo (Testudines: Testudinidae), we have sequenced a fragment of the mitochondrial (mt) 12S rRNA gene of 98 tortoise specimens belonging to the genera Testudo, Indotestudo, and Geochelone. Maximum likelihood and neighbor-joining methods identify two main clades of Mediterranean tortoises, one composed of the species Testudo graeca, Testudo marginata, and Testudo kleinmanni and a second of Testudo hermanni, Testudo horsfieldii, and Indotestudo elongata. The first clade, but not the second, was also supported by maximum parsimony analysis. Together with the genus Geochelone, a star-like radiation of these clades was suggested, as a sister-group relationship between the two Testudo clades could not be confirmed. The intraspecies genetic variation was examined by sequencing the mt 12S rRNA fragment from 28 specimens of T. graeca and 49 specimens of T. hermanni from various geographic locations. Haplotype diversity was found to be significantly larger in T. graeca compared with T. hermanni, suggestive of reduced genetic diversity in the latter species, perhaps due to Pleistocene glaciations affecting northern and middle Europe or other sources of lineage reduction. No ancient mt 12S rRNA gene haplotypes were identified in T. graeca and/or T. hermanni originating from islands in the Mediterranean Sea, suggesting that these islands harbor tortoise populations introduced from the European and African mainland.     

Population structure of island‐associated dolphins: Evidence from mitochondrial and microsatellite markers for common bottlenose dolphins (Tursiops truncatus) around the main Hawaiian Islands

We used mitochondrial and nuclear genetic markers to investigate population structure of common bottlenose dolphins, Tursiops truncatus, around the main Hawaiian Islands. Though broadly distributed throughout the world's oceans, bottlenose dolphins are known to form small populations in coastal waters. Recent photo‐identification data suggest the same is true in Hawaiian waters. We found genetic differentiation among (mtDNA Φ_ST= 0.014–0.141, microsatellite F’_ST= 0.019–0.050) and low dispersal rates between (0.17–5.77 dispersers per generation) the main Hawaiian Island groups. Our results are consistent with movement rates estimated from photo‐identification data and suggest that each island group supports a demographically independent population. Inclusion in our analyses of samples collected near Palmyra Atoll provided evidence that the Hawaiian Islands are also occasionally visited by members of a genetically distinct, pelagic population. Two of our samples exhibited evidence of partial ancestry from Indo‐Pacific bottlenose dolphins (T. aduncus), a species not known to inhabit the Hawaiian Archipelago. Our findings have important implications for the management of Hawaiian bottlenose dolphins and raise concerns about the vulnerability to human impacts of pelagic species in island ecosystems.     

Ralstonia solanacearum needs Flp pili for virulence on potato

Type IV pili are virulence factors in various bacteria. Several subclasses of type IV pili have been described according to the characteristics of the structural prepilin subunit. Although type IVa pili have been implicated in the virulence of Ralstonia solanacearum, type IVb pili have not previously been described in this plant pathogen. Here, we report the characterization of two distinct tad loci in the R. solanacearum genome. The tad genes encode functions necessary for biogenesis of the Flp subfamily of type IVb pili initially described for the periodontal pathogen Aggregatibacter actinomycetemcomitans. To determine the role of the tad loci in R. solanacearum virulence, we mutated the tadA2 gene located in the megaplasmid that encodes a predicted NTPase previously reported to function as the energizer for Flp pilus biogenesis. Characterization of the tadA2 mutant revealed that it was not growth impaired in vitro or in planta, produced wild-type levels of exopolysaccharide galactosamine, and exhibited swimming and twitching motility comparable with the wild-type strain. However, the tadA2 mutant was impaired in its ability to cause wilting of potato plants. This is the first report where type IVb pili in a phytopathogenic bacterium contribute significantly to plant pathogenesis.     

Amber mutants of bacteriophage T4D: their isolation and genetic characterization

We have isolated a large number of mutants of bacteriophage T4D that are unable to form plaques on strain B of Escherichia coli, but are able to grow (nearly) normally on some other strains of E. coli, in particular strain CR63. These mutants, designated amber (am), have been characterized by complementation tests, by genetic crosses, and by their response to chemical mutagens. It is concluded that a particular subclass of base substitution mutations may give rise to amber mutants and that such mutants occur in many genes, which are widely distributed over the T4 genome.     

Inotropic effects of prokinetic agents with 5-HT(4) receptor agonist actions on human isolated myocardial trabeculae

AimsBesides acting as gastrointestinal prokinetic agents, 5-hydroxytryptamine₄ (5-HT₄) receptor agonists can induce positive inotropism in human isolated atrium, but not in ventricles. We pharmacologically evaluated the gastroprokinetic 5-HT₄ receptor agonists tegaserod, prucalopride, R199715, cisapride, the cisapride metabolite norcisapride, and the 5-HT₃ receptor agonist MKC773 on human isolated myocardial trabeculae, and compared their effects with those induced by 5-HT and 5-methoxytryptamine (5-MeOT).Main methodsAtrial and ventricular trabeculae were paced and changes in contractile force were studied in the absence or presence of the 5-HT₄ receptor antagonist GR113808. Partial agonism was assessed using 5-HT₄ receptor agonists as antagonists against 5-HT. To test the contribution of L-type calcium channels, the inotropic responses to 5-HT and 5-MeOT were studied in the absence or presence of verapamil.Key findingsLike 5-HT and 5-MeOT, cisapride and tegaserod, but not prucalopride, R19971 and MKC­733, induced concentration-dependent positive inotropic responses on atrial trabeculae, which were abolished by GR113808. The L-type calcium channel blocker verapamil attenuated inotropic responses to 5-HT and 5-MeOT. None of the agonists affected the contraction of left ventricular trabeculae. Concentration response curves to 5-HT were shifted to the right in the presence of prucalopride, cisapride, tegaserod and R199715, but not MKC-773.SignificanceWe conclude that (i) inotropic responses to 5-HT and 5-MeOT seem to depend on L-type calcium channels, (ii) tegaserod and cisapride behave as partial 5-HT₄ receptor agonists, while prucalopride, norcisapride and MKC-733 cause no significant effects on human atrial trabeculae, (iii) R199715 seems to behave as a 5-HT₄ receptor antagonist.     

Establishment of new mutations under divergence and genome hitchhiking

Theoretical models addressing genome-wide patterns of divergence during speciation are needed to help us understand the evolutionary processes generating empirical patterns. Here, we examine a critical issue concerning speciation-with-gene flow: to what degree does physical linkage (r < 0.5) of new mutations to already diverged genes aid the build-up of genomic islands of differentiation? We used simulation and analytical approaches to partition the probability of establishment for a new divergently selected mutation when the mutation (i) is the first to arise in an undifferentiated genome (the direct effect of selection), (ii) arises unlinked to any selected loci (r = 0.5), but within a genome that has some already diverged genes (the effect of genome-wide reductions in gene flow for facilitating divergence, which we term 'genome hitchhiking'), and (iii) arises in physical linkage to a diverged locus (divergence hitchhiking). We find that the strength of selection acting directly on a new mutation is generally the most important predictor for establishment, with divergence and genomic hitchhiking having smaller effects. We outline the specific conditions under which divergence and genome hitchhiking can aid mutation establishment. The results generate predictions about genome divergence at different points in the speciation process and avenues for further work.