Older than New Caledonia emergence? A molecular phylogenetic study of the eneopterine crickets (Orthoptera: Grylloidea)

Aim A New Caledonian insect group was studied in a world‐wide phylogenetic context to test: (1) whether local or regional island clades are older than 37 Ma, the postulated re‐emergence time of New Caledonia; (2) whether these clades show evidence for local radiations or multiple colonizations; and (3) whether there is evidence for relict taxa with long branches in phylogenetic trees that relate New Caledonian species to geographically distant taxa. Location New Caledonia, south‐west Pacific. Methods We sampled 43 cricket species representing all tribes of the subfamily Eneopterinae and 15 of the 17 described genera, focusing on taxa distributed in the South Pacific and around New Caledonia. One nuclear and three mitochondrial genes were analysed using Bayesian and parsimony methods. Phylogenetic divergence times were estimated using a relaxed clock method and several calibration criteria. Results The analyses indicate that, under the most conservative dating scenario, New Caledonian eneopterines are 5–16 million years old. The largest group in the Pacific region dates to 18–29 Ma. New Caledonia has been colonized in two phases: the first around 10.6 Ma, with the subsequent diversification of the endemic genus Agnotecous, and the second with more recent events around 1–4 Ma. The distribution of the sister group of Agnotecous and the lack of phylogenetic long branches in the genus refute an assumption of major extinction events in this clade and the hypothesis of local relicts. Main conclusions Our phylogenetic studies invalidate a simple scenario of local persistence of this group in New Caledonia since 80 Ma, either by survival on the New Caledonian island since its rift from Australia, or, if one accepts the submergence of New Caledonia, by local island‐hopping among other subaerial islands, now drowned, in the region during periods of New Caledonian submergence.     

Distinguishing between convergence and parallelism is central to comparative biology: a reply to Williams and Ebach

The definitions of character similarity, homology, homoplasy, heterology, parallelism and convergence are clarified in the framework of current phylogenetic methodology. They are all associated with definite patterns of character change and can consequently be tested by phylogeny building. Their crucial significance in comparative biology is illustrated using demonstrative examples. © The Willi Hennig Society 2006.     

Urinary levels of 2,3-dinor-6-oxo-PGF1 alpha: a reliable index of the production of PGI2 in the spontaneously hypertensive rat

The urinary levels of 2,3-dinor-6-oxo-PGF1 alpha (PGI2-M), a major metabolite of PGI2, are determined by the balance between the amount of PGI2 synthesized and the extent of its further metabolic oxidation. The purpose of the present study was to determine if the urinary excretion of PGI2-M can be used as a reliable index of the in vivo production of PGI2 in both normal Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). This involved the exclusion of differences in metabolism between these two strains of rats. In order to do so, we monitored the urinary excretion of PGI2-M during paired intravenous infusions of 6-oxo-PGF1 alpha (the stable product of the spontaneous hydrolysis of PGI2) in conscious, unrestrained SHR and WKY rats aged 12-15 weeks, in doses ranging from 250 to 700 ng. In one experiment, PGI2 was infused instead of 6-oxo-PGF1 alpha. The results of these experiments indicate that SHR and WKY rats are equal with regard to the transformation of 6-oxo-PGF1 alpha and PGI2 into PGI2-M. For both groups, there is a good correlation between the amount of 6-oxo-PGF1 alpha infused and the amount of PGI2-M excreted in urine. These observations confirm the validity of using the urinary levels of 2,3-dinor-6-oxo-PGF1 alpha as an index of PGI2 production in both WKY and SHR. In addition, they support the conclusions drawn from our previous studies, namely that SHR do not produce more PGI2 than WKY rats in vivo, contrary to the situation prevailing in vitro.     

The problem of characters susceptible to parallel evolution in phylogenetic analysis: a reply to Marquès and Gnaspini (2001) with emphasis on cave life phenotypic evolution

Marquès and Gnaspini [Cladistics 17 (2001) 371–381] analyzed the problem of the phylogenetic treatment of characters submitted to parallel evolution. Their proposal aimed at preserving the potential phylogenetic significance of supposedly homoplastic characters and considering them for phylogeny reconstruction. As an example, they used the troglobiomorphic features of animals restricted to caves; according to their preliminary hypothesis of homoplasy, troglobitic animals would exhibit a particular phenotype, referred to as troglobiomorphic, which they would acquire under similar selective pressures from the subterranean environment. We examined Marquès and Gnaspini's approach not from the point of view of its technical flaws, but from the point of view of the authors’ basic assertions on the treatment of troglobiomorphic characters and more generally the inclusion/exclusion/transformation of characters prior to phylogenetic analysis. In the present paper, we argue that this approach is invalidated by the repeated use of ad hoc hypotheses, which are supported neither by an existing phylogenetic pattern nor by available data. We consequently contest the adequateness of this approach with a cladistic analysis of historical questions.     

The C. elegans RSA complex localizes protein phosphatase 2A to centrosomes and regulates mitotic spindle assembly

Microtubule behavior changes during the cell cycle and during spindle assembly. However, it remains unclear how these changes are regulated and coordinated. We describe a complex that targets the Protein Phosphatase 2A holoenzyme (PP2A) to centrosomes in C. elegans embryos. This complex includes Regulator of Spindle Assembly 1 (RSA-1), a targeting subunit for PP2A, and RSA-2, a protein that binds and recruits RSA-1 to centrosomes. In contrast to the multiple functions of the PP2A catalytic subunit, RSA-1 and RSA-2 are specifically required for microtubule outgrowth from centrosomes and for spindle assembly. The centrosomally localized RSA-PP2A complex mediates these functions in part by regulating two critical mitotic effectors: the microtubule destabilizer KLP-7 and the C. elegans regulator of spindle assembly TPXL-1. By regulating a subset of PP2A functions at the centrosome, the RSA complex could therefore provide a means of coordinating microtubule outgrowth from centrosomes and kinetochore microtubule stability during mitotic spindle assembly.     

Differential regulation of mouse equilibrative nucleoside transporter 1 (mENT1) splice variants by protein kinase CK2

Nucleosides are accumulated by cells via a family of equilibrative transport proteins (ENTs). An alternative splice variant of the most common subtype of mouse ENT (ENT1) has been identified which is missing a protein kinase CK2 (casein kinase 2) consensus site (Ser(254)) in the central intracellular loop of the protein. We hypothesized that this variant (mENT1a) would be less susceptible to modulation by CK2-mediated phosphorylation compared to the variant containing the serine at position 254 (mENT1b). Each splice variant was transfected into nucleoside transporter deficient PK15 cells, and stable transfectants assessed for their ability to bind the ENT1-selective probe [(3)H]nitrobenzylthioinosine (NBMPR) and to mediate the cellular uptake of [(3)H]2-chloroadenosine, with or without treatment with the CK2 selective inhibitor, 4,5,6,7-tetrabromobenzotriazole (TBB). mENT1a had a higher affinity for NBMPR relative to mENT1b - measured both directly by the binding of [(3)H]NBMPR, and indirectly via inhibition of [(3)H]2-chloroadenosine influx by NBMPR. Furthermore, incubation of mENT1b-expressing cells with 10 microM TBB for 48 h decreased both the K(D) and B(max) of [(3)H]NBMPR binding, as well as the V(max) of 2-chloroadenosine uptake, whereas similar treatment of mENT1a-expressing cells with TBB had no effect. PK15 cells transfected with hENT1, which has Ser(254), was similar to mENT1b in its response to TBB. In conclusion, inhibition of CK2 activity, or deletion of Ser(254) from mENT1, enhances transporter affinity for the inhibitor, NBMPR, and reduces the number of ENT1 proteins functioning at the level of the plasma membrane.     

Mucosal vaccine delivery of antigens tightly bound to an adjuvant particle made from food-grade bacteria

Mucosal immunization with subunit vaccines requires new types of antigen delivery vehicles and adjuvants for optimal immune responses. We have developed a non-living and non-genetically modified gram-positive bacterial delivery particle (GEM) that has built-in adjuvant activity and a high loading capacity for externally added heterologous antigens that are fused to a high affinity binding domain. This binding domain, the protein anchor (PA), is derived from the Lactococcus lactis AcmA cell-wall hydrolase, and contains three repeats of a LysM-type cell-wall binding motif. Antigens are produced as antigen-PA fusions by recombinant expression systems that secrete the hybrid proteins into the culture growth medium. GEM particles are then used as affinity beads to isolate the antigen-PA fusions from the complex growth media in a one step procedure after removal of the recombinant producer cells. This procedure is also highly suitable for making multivalent vaccines. The resulting vaccines are stable at room temperature, lack recombinant DNA, and mimic pathogens by their bacterial size, surface display of antigens and adjuvant activity of the bacterial components in the GEM particles. The GEM-based vaccines do not require additional adjuvant for eliciting high levels of specific antibodies in mucosal and systemic compartments.