Studies of κ2- and κ3-tripyridylamine complexes of ruthenium and π-stacking by pyridyls.

The reaction of tris(pyridin-2-yl)amine with [CyRuCl2]2 (Cy = p-isopropyltoluene or cymene) in refluxing diglyme led to the formation of cis-[RuCl2{κ2-(2-py)3N}2]·CHCl3 (1a) after recrystallization from chloroform/pentane, or cis-dichloridobis[tris(pyridin-2-yl)amine-κ2N,N']ruthenium(II) dichloromethane disolvate, [RuCl2(C15H12N4)2]·2CH2Cl2 or cis-[RuCl2{κ2-(2-py)3N}2]·2CH2Cl2 (1b). Treatment of 1a with one equivalent of silver(I) hexafluoridoantimonate in dichloromethane gave [RuCl{κ2-(2-py)3N}{κ3-(2-py)3N}][SbF6]·CH2Cl2 (2a). Crystallization of 2a from chloroform provided chlorido[tris(pyridin-2-yl)amine-κ2N,N'][tris(pyridin-2-yl)amine-κ3N,N',N'']ruthenium(II) hexafluoridoantimonate chloroform monosolvate, [RuCl(C15H12N4)2][SbF6]·CHCl3 or [RuCl{κ2-(2-py)3N}{κ3-(2-py)3N}][SbF6]·CHCl3 (2b). Complex 2a reacted with a further equivalent of silver(I) hexafluoridoantimonate to give [Ru{κ3-(2-py)3N}2][SbF6]2 (3). The reaction of (2-py)3N with [CyRuCl2]2 in dichloromethane followed by treatment with excess sodium hexafluoridoantimonate gave the known complex [CyRuCl{κ2-(2-py)3N}][SbF6] (4). Complex 2 is a rare example of a complex containing both κ2- and κ3-(2-py)3N. Intramolecular π-stacking interactions determine the orientation of the free pyridyl in the κ2 complexes. An interesting encapsulation of methylene chloride hydrogen-bonded tetramers was noted in one case.

Reconstructing the Origins of the Somatostatin and Allatostatin-C Signaling Systems Using the Accelerated Evolution of Biodiverse Cone Snail Toxins.

Somatostatin and its related peptides (SSRPs) form an important family of hormones with diverse physiological roles. The ubiquitous presence of SSRPs in vertebrates and several invertebrate deuterostomes suggests an ancient origin of the SSRP signaling system. However, the existence of SSRP genes outside of deuterostomes has not been established, and the evolutionary history of this signaling system remains poorly understood. Our recent discovery of SSRP-like toxins (consomatins) in venomous marine cone snails (Conus) suggested the presence of a related signaling system in mollusks and potentially other protostomes. Here, we identify the molluscan SSRP-like signaling gene that gave rise to the consomatin family. Following recruitment into venom, consomatin genes experienced strong positive selection and repeated gene duplications resulting in the formation of a hyperdiverse family of venom peptides. Intriguingly, the largest number of consomatins was found in worm-hunting species (>400 sequences), indicating a homologous system in annelids, another large protostome phylum. Consistent with this, comprehensive sequence mining enabled the identification of SSRP-like sequences (and their corresponding orphan receptor) in annelids and several other protostome phyla. These results established the existence of SSRP-like peptides in many major branches of bilaterians and challenge the prevailing hypothesis that deuterostome SSRPs and protostome allatostatin-C are orthologous peptide families. Finally, having a large set of predator-prey SSRP sequences available, we show that although the cone snail's signaling SSRP-like genes are under purifying selection, the venom consomatin genes experience rapid directional selection to target receptors in a changing mix of prey.

Integrating Venom Peptide Libraries Into a Phylogenetic and Broader Biological Framework.

The venomous marine snails are conventionally divided into three groups, the cone snails (family Conidae), the auger snails (family Terebridae) and the turrids (formerly all assigned to a single family, Turridae). In this study, a library of venom peptides from species conventionally assigned to the genus Turris was correlated to a phylogenetic analysis. Nucleotide sequences of multiple genes from transcriptomes were used to assess the phylogenetic relationships across a diverse set of species. The resulting tree shows that as conventionally defined, the conoidean genus Turris, is polyphyletic. We describe a new genus, Purpuraturris gen. nov., that comprises the outlier species. In addition to morphological distinctions, molecular data reveal that this group is divergent from Turris sensu stricto. The correlation between phylogenetic information and a family of peptide sequences was used to highlight those peptides mostly likely to be unique and intimately associated with biological diversity. The plethora of peptide sequences available requires two prioritization decisions: which subset of peptides to initially characterize, and after these are characterized, which to comprehensively investigate for potential biomedical applications such as drug developments. Life Science Identifiers: urn:lsid:zoobank.org; pub: 60D46561-28F0-4C39-BAC4-66DC8B4EAEA4.

An integrative approach to the facile functional classification of dorsal root ganglion neuronal subclasses.

Somatosensory neurons have historically been classified by a variety of approaches, including structural, anatomical, and genetic markers; electrophysiological properties; pharmacological sensitivities; and more recently, transcriptional profile differentiation. These methodologies, used separately, have yielded inconsistent classification schemes. Here, we describe phenotypic differences in response to pharmacological agents as measured by changes in cytosolic calcium concentration for the rapid classification of neurons in vitro; further analysis with genetic markers, whole-cell recordings, and single-cell transcriptomics validated these findings in a functional context. Using this general approach, which we refer to as tripartite constellation analysis (TCA), we focused on large-diameter dorsal-root ganglion (L-DRG) neurons with myelinated axons. Divergent responses to the K-channel antagonist, κM-conopeptide RIIIJ (RIIIJ), reliably identified six discrete functional cell classes. In two neuronal subclasses (L1 and L2), block with RIIIJ led to an increase in [Ca] i Simultaneous electrophysiology and calcium imaging showed that the RIIIJ-elicited increase in [Ca] i corresponded to different patterns of action potentials (APs), a train of APs in L1 neurons, and sporadic firing in L2 neurons. Genetically labeled mice established that L1 neurons are proprioceptors. The single-cell transcriptomes of L1 and L2 neurons showed that L2 neurons are Aδ-low-threshold mechanoreceptors. RIIIJ effects were replicated by application of the Kv1.1 selective antagonist, Dendrotoxin-K, in several L-DRG subclasses (L1, L2, L3, and L5), suggesting the presence of functional Kv1.1/Kv1.2 heteromeric channels. Using this approach on other neuronal subclasses should ultimately accelerate the comprehensive classification and characterization of individual somatosensory neuronal subclasses within a mixed population.

Age relationships of postmortem observations in Portuguese Water Dogs.

A dog model has been used to evaluate histological changes arising from senescence. Autopsies of 145 Portuguese Water Dogs have been used to evaluate the individual and group "state of health" at time of death. For each dog, weights or dimensions of organs or tissues were obtained, together with histological evaluation of tissues. Twenty-three morphological metrics correlated significantly to age at death. Many of these involved muscles; others were associated with derivatives of embryonic foregut. The latter included lengths of the small intestine and trachea as well as weights of the stomach and some lung lobes. Nearly all of the dogs examined had histological changes in multiple tissues, ranging from two to 12 per dog. Associations among pathologies included inflammatory bowel disease with osteoporosis and dental calculus/periodontitis with atherosclerosis and amyloidosis. In addition, two clusters of histological changes were correlated to aging: hyperplasia, frequency of adenomas, and hemosiderosis constituted one group; inflammation, plasmacytic and lymphocytic infiltration, fibrosis, and atrophy, another. Heritability analysis indicated that many of the changes in tissue/organ morphology or histology could be heritable and possibly associated with IGF1, but more autopsies will be required to substantiate these genetic relationships.

Genetic mapping of fixed phenotypes: disease frequency as a breed characteristic.

Traits that have been stringently selected to conform to specific criteria in a closed population are phenotypic stereotypes. In dogs, Canis familiaris, such stereotypes have been produced by breeding for conformation, performance (behaviors), etc. We measured phenotypes on a representative sample to establish breed stereotypes. DNA samples from 147 dog breeds were used to characterize single nucleotide polymorphism allele frequencies for association mapping of breed stereotypes. We identified significant size loci (quantitative trait loci [QTLs]), implicating candidate genes appropriate to regulation of size (e.g., IGF1, IGF2BP2 SMAD2, etc.). Analysis of other morphological stereotypes, also under extreme selection, identified many additional significant loci. Behavioral loci for herding, pointing, and boldness implicated candidate genes appropriate to behavior (e.g., MC2R, DRD1, and PCDH9). Significant loci for longevity, a breed characteristic inversely correlated with breed size, were identified. The power of this approach to identify loci regulating the incidence of specific polygenic diseases is demonstrated by the association of a specific IGF1 haplotype with hip dysplasia, patella luxation, and pancreatitis.

The aging feline kidney: a model mortality antagonist?

Traditional thinking views apparently non-programmed disruptions of aging, which medical science calls geriatric diseases, as separate from 'less harmful' morphological and physiological aging phenotypes that are more universally expected with passage of time (loss of skin elasticity, graying of hair coat, weight gain, increased sleep time, behavioral changes, etc). Late-life disease phenotypes, especially those involving chronic processes, frequently are complex and very energy-expensive. A non-programmed process of homeostatic disruption leading into a death trajectory seems inconsistent with energy intensive processes. That is, evolutionary mechanisms do not favor complex and prolonged energy investment in death. Taking a different view, the naturally occurring feline (Felis silvestris catus) renal model suggests that at least some diseases of late life represent only the point of failure in essentially survival-driven adaptive processes. In the feline renal model, individuals that succumbed to failure most frequently displayed progressive tubular deletion and peritubular interstitial fibrosis, but had longer mean life span than cats that died from other causes. Additionally, among cats that died from non-renal causes, those that had degrees of renal tubular deletion and peritubular interstitial fibrosis also had longer mean life span than those cats with no changes, even though causes of death differed minimally between these latter two groups. The data indicate that selective tubular deletion very frequently begins early in adult life, without a clear initiating phase or event. The observations support a hypothesis that this prolonged process may be intrinsic and protective prior to an ultimate point of failure. Moreover, given the genetic complexity and the interplay with associated risk factors, existing data also do not support the ideas that these changes are simple compensatory responses and that breed- or strain-based 'default' diseases are inevitable results of increasing individual longevity. Emerging molecular technology offers the future potential to further evaluate and refine these observations. At present, the existence of plastic and adaptive aging programming is suggested by these findings.

Bilaterally asymmetric effects of quantitative trait loci (QTLs): QTLs that affect laxity in the right versus left coxofemoral (hip) joints of the dog (Canis familiaris).

In dogs hip joint laxity that can lead to degenerative joint disease (DJD) is frequent and heritable, providing a genetic model for some aspects of the human disease. We have used Portuguese water dogs (PWDs) to identify Quantitative trait loci (QTLs) that regulate laxity in the hip joint. A population of 286 PWDs, each characterized by ca. 500 molecular genetic markers, was analyzed for subluxation of the hip joint as measured by the Norberg angle, a quantitative radiographic measure of laxity. A significant directed asymmetry was observed, such that greater laxity was observed in the left than the right hip. This asymmetry was not heritable. However, the average Norberg angle was highly heritable as were the Norberg angles of either the right or left hips. After correction for pedigree effects, two QTLs were identified using the metrics of the left and right hips as separate data sets. Both are on canine chromosome 1 (CFA1), separated by about 95 Mb. One QTL, associated with the SSR marker FH2524 was significant for the left, but not the right hip. The other, associated with FH2598, was significant for the right but not the left hip. For both QTLs, some extreme phenotypes were best explained by specific interactions between haplotypes.