RESTING CYST FORMATION OF EUTREPTIELLA GYMNASTICA (EUGLENOPHYCEAE) IN THE NORTHERN COASTAL BALTIC SEA1

Resting cyst formation of Eutreptiella gymnastica Throndsen was observed during a mesocosm experiment, where nutrient enrichment had induced almost a unialgal bloom. Cells and resting cysts of E. gymnastica were examined in scanning (SEM) and transmission electron microscopy (TEM) and light microscopy. Mature cysts were spherical, with a smooth thick mucilaginous cover that appeared layered when observed with the TEM. Intermediate forms were spherical and lacking flagella and a mucilaginous cover; the euglenoid pellicular striation and canal opening were easily visible. The volume of these intermediate spherical cells and mature cysts was estimated to have increased threefold compared to flagellated cells and contained many paramylon grains. When the cells were grazed by zooplankton, the paramylon grains passed the gut intact and were packed into fecal pellets. Intact undigested paramylon grains were observed in SEM after the breaking up of the pellets.     

HIGH ENCYSTMENT SUCCESS OF THE DINOFLAGELLATE SCRIPPSIELLA CF. LACHRYMOSA IN CULTURE EXPERIMENTS 1

Close to 100% encystment efficiency and a yield above 10⁵ cysts·mL ^{? 1} were routinely achieved in full strength f/2 medium‐based batch cultures (883 μM NO₃ ^? and 36 μM PO₄ ^{? 3}) of the marine dinoflagellate Scrippsiella cf. lachrymosa Lewis. Increases in cell density led to nutrient depletion in this enriched medium, which was the most likely cause for initiation of cyst formation. Lowering the concentration of either nutrient to 1/10 the initial levels decreased the encystment efficiency, whereas use of ammonium as the N source resulted in both low cell yield and low encystment efficiency. The mandatory dormancy period was ca. 60 days and was not affected by cold dark storage of the cysts. Cysts produced in the initial phase of sexual reproduction were relatively large (length 47 μm, width 31 μm) with a heavy calcareous cover. Cysts produced thereafter lacked apparent calcareous cover and were smaller (length 29 μm, width 19 μm). The decrease of cyst volume (by a factor of 0.24–0.4) suggested strong resource limitation during the course of encystment. However, after the mandatory dormancy period, germination success of the smaller cysts was higher (80%), compared with the larger cysts that had been produced initially (50%). Germling survival (74%) was independent of cyst type but was enhanced by higher nutrient concentration during incubation. The ratio of initial nutrient concentration in the medium to the cyst yield was used as a proxy to estimate the cellular nutrient quota. The conservative estimates of 9 pmol N·cyst ^{? 1} and 0.4 pmol P·cyst ^{? 1} obtained in this manner are at the low end of the range of previous published estimates for other dinoflagellate cysts. Given the high encystment observed in laboratory experiments, we have no reason to assume an inherently lower encystment success in dinoflagellate field populations. Our results do not challenge the low nutrient paradigm for dinoflagellate sexuality. We believe that the high encystment success and cyst yield of this particular species is at least partly due to its ability to achieve very high cell densities in cultures, which evidently leads to nutrient depletion even in f/2 medium.     

Exercise-induced hormone responses in girls at different stages of sexual maturation

The dependence of exercise-induced hormone responses on sexual maturation was tested in a 3-year longitudinal experiment on 34 girls (aged 11–12 years at the beginning). Sexual maturation was evaluated by Tanners five-stage scale. Children cycled for 20-min at 60% maximal oxygen uptake once a year. Cortisol, insulin, growth hormone, β-oestradiol, progesterone and testosterone concentrations in venous blood were determined by radioimmunoassay procedures. Basal concentrations of growth hormone increased and of cortisol decreased when breast stage III was reached. Reaching breast stage IV was associated with an increase in basal concentrations of β-oestradiol, progesterone and testosterone. The exercise induced significant increases in concentrations of cortisol, growth hormone and β-oestradiol and a decrease in insulin concentration. At breast stage III the increase in cortisol concentration was to a lower level [467 (SEM 42) vs 567 (SEM 46)nmol · l⁻¹] and growth hormone concentration to a higher level [29.4 (SEM 0.5) vs 12.8 (SEM 0.4)ng · ml⁻¹], while the fall in insulin concentration was less pronounced [postexercise level 10.6 (SEM 0.9) vs 7.8 (SEM 0.8)mU · l⁻¹] than in stage II. The magnitude of the cortisol response was reduced in the last stage of breast development (+42.1% vs +55.5% at stage II, +66.2% at stage III, and +50.0% at stage IV). The magnitude of β-oestradiol response was the lowest in breast stage IV (+15.8%) and the highest at stage V (+41.1%). The progesterone response became significant at stage IV and testosterone response at stage V. In conclusion, we found that reaching breast stage III was associated with altered responses of cortisol, insulin and growth hormone concentrations while the responses of the sex hormone concentrations became pronounced in the last stages of sexual maturation. Accepted: 17 September 1997     

Modelling niche and neutral dynamics: on the ecological interpretation of variation partitioning results

Many studies have attempted to disentangle the effects of neutral and niche‐mediated processes on community composition using partial Mantel tests and variance partitioning on dissimilarity matrices. Recently, doubts about the reliability of these methods have emerged. Here we explore how the results are affected by three confounding factors that may affect ecological data to different degrees: spatial autocorrelation of the environmental variables, length of the compositional gradient, and sampling noise. We document that the statistical hypotheses tested in these methods may or may not correspond to the ecological hypotheses of interest. A major discrepancy emerges if a large proportion of sampling units in the analysed dataset share no species, in which case compositional dissimilarities saturate to a fixed maximum value although explanatory dissimilarities do not. With increasing dissimilarity saturation, the explanatory power of regression models decrease, which may lead to the erroneous conclusion that the ecological processes represented by the explanatory variables are not operating. A survey of recent literature suggests that there is a general lack of awareness of this problem, although it appears to affect > 10% of relevant studies. Our simulations show that if dissimilarity saturation is due to a long ecological gradient, extended dissimilarities essentially solve the problem for any degree of saturation. Using distances from a hybrid multidimensional scaling alleviates the saturation problem when the degree of saturation is < 60%. However, neither correction method can provide a solution to problems caused by insufficient sampling. How the presence of multiple explanatory gradients in combination with sampling noise affects overall analysis performance remains to be clarified.     

Human immunodeficiency virus type 1 Nef recruits the guanine exchange factor Vav1 via an unexpected interface into plasma membrane microdomains for association with p21-activated kinase 2 activity

Alterations of T-cell receptor signaling by human immunodeficiency virus type 1 (HIV-1) Nef involve its association with a highly active subpopulation of p21-activated kinase 2 (PAK2) within a dynamic signalosome assembled in detergent-insoluble membrane microdomains. Nef-PAK2 complexes contain the GTPases Rac and Cdc42 as well as a factor providing guanine nucleotide exchange factor (GEF) activity for Rac/Cdc42. However, the identity of this GEF has remained controversial. Previous studies suggested the association of Nef with at least three independent GEFs, Vav, DOCK2/ELMO1, and βPix. Here we used a broad panel of approaches to address which of these GEFs is involved in the functional interaction of Nef with PAK2 activity. Biochemical fractionation and confocal microscopy revealed that Nef recruits Vav1, but not DOCK2/ELMO1 or βPix, to membrane microdomains. Transient RNAi knockdown, analysis of cell lines defective for expression of Vav1 or DOCK2 as well as use of a βPix binding-deficient PAK2 variant confirmed a role for Vav1 but not DOCK2 or βPix in Nef's association with PAK2 activity. Nef-mediated microdomain recruitment of Vav1 occurred independently of the Src homology 3 domain binding PxxP motif, which is known to connect Nef to many cellular signaling processes. Instead, a recently described protein interaction surface surrounding Nef residue F195 was identified as critical for Nef-mediated raft recruitment of Vav1. These results identify Vav1 as a relevant component of the Nef-PAK2 signalosome and provide a molecular basis for the role of F195 in formation of a catalytically active Nef-PAK2 complex.     

Generation of a conditionally replicating adenovirus based on targeted destruction of E1A mRNA by a cell type-specific MicroRNA

MicroRNAs have emerged as important players in tissue-specific mammalian gene regulation and have also been exploited in experimental targeting of gene expression. We have constructed a recombinant adenovirus that contains sequences complementary to the liver-specific microRNA 122 (miR122) in the 3′ untranslated region of the E1A gene. In Huh7 cells, which resemble normal hepatocytes in expressing high levels of miR122, this feature resulted in strongly reduced levels of E1A mRNA and protein. This property allowed us to generate a novel recombinant adenovirus that was severely attenuated in cells of hepatic origin but replicated normally in other cells. This strategy may be useful in circumventing liver toxicity associated with the systemic delivery of oncolytic adenoviruses. These data provide the first example of exploiting differential microRNA expression patterns to alter the natural tropism of a DNA virus. In addition, these results suggest that other microRNAs expressed in a tissue- or transformation-specific manner may also be used for the targeting of adenoviral replication and that the same principle may be applied to other viruses that have shown promise as oncolytic or gene delivery platforms.     

Avian and 1918 Spanish influenza a virus NS1 proteins bind to Crk/CrkL Src homology 3 domains to activate host cell signaling

NS1 (nonstructural protein 1) is an important virulence factor of the influenza A virus. We observed that NS1 proteins of the 1918 pandemic virus (A/Brevig Mission/1/18) and many avian influenza A viruses contain a consensus Src homology 3 (SH3) domain-binding motif. Screening of a comprehensive human SH3 phage library revealed the N-terminal SH3 of Crk and CrkL as the preferred binding partners. Studies with recombinant proteins confirmed avid binding of NS1 proteins of the 1918 virus and a representative avian H7N3 strain to Crk/CrkL SH3 but not to other SH3 domains tested, including p85alpha and p85beta. Endogenous CrkL readily co-precipitated NS1 from cells infected with the H7N3 virus. In transfected cells association with CrkL was observed for NS1 of the 1918 and H7N3 viruses but not A/Udorn/72 or A/WSN/33 NS1 lacking this sequence motif. SH3 binding was dispensable for suppression of interferon-induced gene expression by NS1 but was associated with enhanced phosphatidylinositol 3-kinase signaling, as evidenced by increased Akt phosphorylation. Thus, the Spanish Flu virus resembles avian influenza A viruses in its ability to recruit Crk/CrkL to modulate host cell signaling.     

A test of the compensatory continuum: fertilization increases and below-ground competition decreases the grazing tolerance of tall wormseed mustard (Erysimum strictum)

Contrary to the general expectation, the compensatory continuum hypothesis proposes that grazing may not always affect plant performance adversely. Instead, the effects may vary from negative (undercompensation) to positive (overcompensation), depending on the local availability of resources and the intensity of competition experienced by individual plants. We tested this hypothesis in a common garden experiment by growing tall wormseed mustard, Erysimum strictum, under a factorial design involving simulated grazing (0, 10, or 50% of the main stem clipped), supplemental fertilization and below-ground competition. The results supported the hypothesis. On an average, fertilization increased and competition decreased plant performance. Overcompensation was only observed among the fertilized plants growing free of competition. Simulated grazing increased seed yield 1.6 (10% clipping) and 1.4 times (50% clipping) as compared to unclipped plants when the plants were grown with fertilization and without competition. In contrast, clipping did not significantly increase seed yield in the plants grown without fertilization and/or with competition. The breakage of apical dominance provides a proximate mechanism of these regrowth responses. This is consistent with the fact that most plants (85%) had an unbranched shoot architecture in our study population. However, it is not clear why E. strictum has a relatively unbranched architecture in natural populations. We briefly discuss the alternative ecological factors – competition for light, adaptation to herbivory and optimal timing of flowering as a bet-hedging strategy in monocarpic plants – which might maintain unbranched architecture in this species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Climatic responses in spring migration of boreal and arctic birds in relation to wintering area and taxonomy

Large‐scale climate fluctuations, such as the North Atlantic Oscillation (NAO), have a marked effect on the timing of spring migration of birds. It has however been suggested that long‐distance migrants wintering in Africa could respond less to NAO than short‐distance migrants wintering in Europe, making them more vulnerable to climatic changes. We studied whether migratory boreal and arctic bird species returning from different wintering areas show differences in responses to the NAO in the timing of their spring migration. We used data on 75 species from two bird observatories in northern Europe (60°N). By extending the examination to the whole distribution of spring migration and to a taxonomically diverse set of birds, we aimed at finding general patterns of the effects of climate fluctuation on the timing of avian migration. Most species arrived earlier after winters with high NAO index. The degree of NAO‐response diminished with the phase of migration: the early part of a species’ migratory population responded more strongly than the later part. Early phase waterfowl responded strongest to NAO, but in later phases their response faded to non‐significant. This pattern may be related to winter severity and/or ice conditions in the Baltic. In the two other groups, gulls and waders and passerines, all phases of migration responded to NAO and fading with phase was non‐significant. The difference between waterfowl and other groups may be related to differences between the phenological development of their respective macrohabitats. Wintering area affected the strength of NAO response in a complicated way. On average medium distance migrants responded most strongly, followed by short‐distance migrants and partial migrants. Our results concerning the response of long‐distance migrants were difficult to interpret: there is an overall weak yet statistically significant effect, but patterns with phase of migration need further study. Our results highlight the importance of examining the whole distribution of migration and warrant the use of data sets from several sampling sites when studying climatic effects on the timing of avian life‐history events.