Protein transduction in human cells is enhanced by cell-penetrating peptides fused with an endosomolytic HA2 sequence

Endocytosis has been proposed as one of the primary mechanisms for cellular entry of cell-penetrating peptides (CPPs) and their cargoes. However, a major limitation of endocytic pathway is entrapment of the CPP-cargo in intracellular vesicles from which the cargo must escape into the cytoplasm to exert its biological activity. Here we demonstrate that a CPP tagged with an endosomolytic fusion peptide derived from the influenza virus hemagglutinin-2 (HA2) remarkably enhances the cytosolic delivery of proteins in human A549 cells. To determine the endosome-disruptive effects, recombinant DNA plasmids containing coding sequences of HA2, CPPs and red fluorescent proteins (RFPs) were constructed. The fusion proteins were purified from plasmid-transformed Escherichia coli, and their effects on protein transduction were examined using live cell imaging and flow cytometry. Our data indicate that endocytosis is the major route for cellular internalization of CPP-HA2-tagged RFP. Mechanistic studies revealed that the fusogenic HA2 peptide dramatically facilitates CPP-mediated protein entry through the release of endocytosed RFPs from endosomes into the cytoplasm. Furthermore, incorporating the HA2 fusion peptide of the CPP-HA2 fusion protein improved cytosolic uptake without causing cytotoxicity. These findings strongly suggest that the CPP-HA2 tag could be an efficient and safe carrier that overcomes endosomal entrapment of delivered therapeutic drugs.     

Arginine vasopressin 1a receptor gene and maternal behavior: evidence of association and moderation

This study examined associations among maternal sensitivity, mothers' early adversity and the Arginine Vasopressin 1a Receptor (AVPR1A) gene. Early adversity in mothers' background has been found to be associated with lower maternal sensitivity. Animal literature suggests that variation in the AVPR1A gene is associated with parenting quality. The goal of the study was to examine the role of the AVPR1A gene in maternal sensitivity, especially under conditions of high early adversity. Participants included 151 Caucasian women from a community sample. The women were videotaped in their home while interacting separately with two of their children (target child = 18 months, older sibling <6 years). Evidence was found for an association between the AVPR1A gene and maternal sensitivity. Mothers with two copies of the long RS3 alleles were less sensitive than mothers with one or zero copies of the long alleles. This association was strongest under conditions of high maternal early adversity.     

Temporal priority and intra-guild predation in temporary waters: an experimental study using Namibian desert dragonflies

Abstract. 1. Intra-guild predation between early larval instars of two co-existing dragonflies, Sympetrum fonscolombii and Trithemis kirbyi , was investigated with respect to temporal advantage and growth. Three situations were simulated experimentally: (1) Sympetrum fonscolombii began development 11 days before T. kirbyi . (2) Trithemis kirbyi began development 11 days before S. fonscolombii . (3) Both species began on the same day.
2. With a temporal advantage of 11 days to the second species, the resulting larval density of the respective first species was significantly higher than that of the second species.
3. Without a temporal advantage, the survival of S. fonscolombii was higher than that of T. kirbyi , and S. fonscolombii had a larger size due to faster growth than T. kirbyi . Hence, it is assumed that survival depended on early oviposition as well as on larval growth.
4. To test the relevance of the laboratory results, observations at artificial ponds in the Namibian semi-desert were conducted. Trithemis kirbyi was the first species colonising these ponds while S. fonscolombii arrived 15 days later. In field samples, many more Trithemis larvae than Sympetrum larvae were found, a pattern similar to the laboratory experiments in which T. kirbyi enjoyed a temporal advantage.     

Flood-mediated dispersal versus hatching: early recolonisation strategies of copepods in floodplain ponds

1. We explored the quantitative contribution of two alternative recolonisation strategies by a combination of experimental and field data from the Little Tallahatchie floodplain (Mississippi, U.S.A.). In this area, several floods of short duration occur in winter and spring. During floods, lentic and lotic waterbodies of different size are connected for a limited period. 2. Shortly before the onset of the flood season we collected sediment samples from six temporary ponds. The emergence of seven cyclopoid species was recorded from the experimentally flooded sediments and was quantified over a 4‐week period. Water samples were taken in the same ponds after flooding, from which eight further species were recorded and quantified. 3. The contribution of flood‐dispersed individuals was variable among flooded ponds (between 38 and 94%), but numbers of flood‐dispersed colonists did not differ significantly from those of hatching individuals in flooded ponds. Our results demonstrate the equal importance of both hydrological connections and hatching from dormant stages as pathways for recolonisation by copepods if ponds are hydrologically connected. 4. The significance of recolonisation by hatching individuals in disconnected waterbodies was apparent in an isolated temporary pond that was exclusively recolonised by cyclopoid copepods emerging from the sediment.     

Maternally expressed PGK-Cre transgene as a tool for early and uniform activation of the Cre site-specific recombinase

A transgenic mouse strain with early and uniform expres sion of the Cre site-specific recombinase is described. In this strain, PGK-Crem, Cre is driven by the early acting PGK-1 promoter, but, probably due to cis effects at the integration site, the recombinase is under dominant maternal control. When Cre is transmitted by PGK-Crem females mated to males that carry a reporter transgene flanked by loxP sites, even offspring that do not inherit PGK-Cre delete the target gene. It follows that in the PGK-Crem female Cre activity commences in the diploid phase of oogenesis. In PGK-Crem crosses complete recombination was observed in all organs, including testis and ovary. We prepared a mouse stock that is homozygous for PGK-Crem and at the albino (c) locus. This strain will be useful for the early and uniform induction of ectopic and dominant negative mutations, for the in vivo removal of selective elements from targeted mutations and in connection with the manipulation of targeted loci in 'knock in' and related technologies     

Fascicled Androecia in Dilleniidae and Some Remarks on the Garcinia Androecium

In Dilleniidae, stamen fascicles are interpreted either as phylogenetically secondary structures (derived from a single stamen primordium by dédoublement), or as the most archaic type of androecial organs in angiosperms. In context with an assumed high plasticity in the flowers at the beginning of angiosperm evolution (Endress, 1987 a, b), fascicled androecia can also be regarded as having coexisted with “magnolioid” spiral androecia since early in the evolution. On the basis of this assumption, it is easy to link the Dilleniidae, via their basal group, the Paeoniaceae, to the Magnoliidae. In Paeonia, the stamen clusters continue the spiral arrangement of the perianth members, with “limiting divergence” (Hiepko, 1964). Our investigations of Paeonia officinalis show that the fascicle primordia follow the spiral not only in their position, but also in their temporal sequence. In some Theaceae (Stewartia) and Clusiaceae (species of Garcinia) the divergence angle changes to 2/5, resulting in an epipetalous position of the stamen fascicles. The transition from fascicled (complex) androecia to simple ones (one- or two-whorled) has occurred repeatedly within the Dilleniidae. A great diversity of androecial structures, based on a fascicled androecium, can be found in the palaeotropical genus Garcinia (Clusiaceae), which comprises about 200 species. Some of these forms, including exceptional ones, are presented in this article. The diversity in the androecium in Garcinia can be interpreted phylogenetically as a secondarily increased plasticity, resulting in morphological curiosities.     

Experimental trial of Australian arid zone reptiles as early warning indicators of overgrazing by cattle

The value of reptiles as bioindicators of the initial effects of heavy cattle grazing in a South Australian chenopod shrubland was assessed in an experimental trial. Reptiles were sampled in three different subhabitats within two replicate treatment paddocks and a control region. A total of 30 sites was sampled in two sessions before grazing and four sessions after the commencement of intensive pulses of grazing. Capture rates of most common reptile species, fecundity of abundant gecko species and reptile species composition within different subhabitats were largely resilient to the initial effects of overgrazing. However, agamids in general, and particularly Ctenophorus nuchalis, increased at grazed sites relative to controls following grazing. Most changes observed in the reptile assemblages were predictable based upon species response to alteration in vegetation cover. Because vegetation cover can be rapidly and efficiently monitored, the use of reptiles as early warning indicators of unsustainable pastoralism in the study region was not supported.     

Identification of development and tissue-specific gene expression in the fathead minnow Pimephales promelas, Rafinesque using computational and DNA microarray methods

The fathead minnow Pimephales promelas serves as a model organism for assessing the effects of environmental contaminants on early life stage growth and development. Yet, the utilization of genomic tools has been hindered by the lack of genome sequence and genomic information known from this model species. Utilizing published cDNA library sequences, the authors used sequence similarity to compare 4105 cDNAs isolated from fathead minnow fry (<14 days old) with over 250 000 adult cDNA sequences derived from whole body and various tissue types. The objectives of the computational subtraction were to (1) assess the extent of sequence similarity between developing and adult cDNA libraries and (2) predict which cDNA clones are expressed only in developing organisms. The results of the computational predictions were assessed through the construction of a development‐specific DNA microarray targeting all 4105 sequences in the fry cDNA library as well as 56 known mRNAs in P. promelas. Gene expression was determined by comparing total RNA isolated from fry with total RNA isolated from adult samples (whole animal, kidney, liver, brain, ovary and testes). The results showed that 1381 of the targeted fry cDNA sequences (34%) displayed expression across all sample comparisons, and of these, only 166 genes were found to harbour fry‐specific expression (i.e. no expression in adult samples). Of note, 69% of the genes computationally predicted to be fry specific were found across all experimental results; yet, only 27% of the computationally predicted fry‐specific sequences were experimentally confirmed to be fry specific. An important result was the identification of many novel mRNA sequences specific to the developing minnow, which lack homology with any other known sequence. In addition, the study results included tissue‐specific expression in adult samples. These results demonstrate the capabilities and limitations of inter‐library sequence comparisons as a predictor of gene activity in non‐sequenced organisms and tissues, as well as DNA microarray gene expression studies in non‐sequenced organisms.     

Offspring performance is well buffered against stress experienced by ancestors

Evolution should render individuals resistant to stress and particularly to stress experienced by ancestors. However, many studies report negative effects of stress experienced by one generation on the performance of subsequent generations. To assess the strength of such transgenerational effects we propose a strategy aimed at overcoming the problem of type I errors when testing multiple proxies of stress in multiple ancestors against multiple offspring performance traits, and we apply it to a large observational dataset on captive zebra finches (Taeniopygia guttata). We combine clear one-tailed hypotheses with steps of validation, meta-analytic summary of mean effect sizes, and independent confirmatory testing. We find that drastic differences in early growth conditions (nestling body mass 8 days after hatching varied sevenfold between 1.7 and 12.4 g) had only moderate direct effects on adult morphology (95% confidence interval [CI]: r = 0.19–0.27) and small direct effects on adult fitness traits (r = 0.02–0.12). In contrast, we found no indirect effects of parental or grandparental condition (r = −0.017 to 0.002; meta-analytic summary of 138 effect sizes), and mixed evidence for small benefits of matching environments between parents and offspring, as the latter was not robust to confirmatory testing in independent datasets. This study shows that evolution has led to a remarkable robustness of zebra finches against undernourishment. Our study suggests that transgenerational effects are absent in this species, because CIs exclude all biologically relevant effect sizes.