Population genetic structure and male-biased dispersal in the queenless ant Diacamma cyaneiventre

In this study we investigated the population genetic structure of the queenless ant Diacamma cyaneiventre. This species, lacking winged queens, is likely to have a restricted female dispersal. We used both mitochondrial and microsatellite markers to assess the consequence of such restricted female dispersal at three geographical scales: within a given locality (< 1 km), between localities within a given region (< 10 km) and between regions (> 36 km). Within a locality, a strong population structure was observed for mitochondrial DNA (mtDNA) whereas weak or nonexistent population genetic structure was observed for the microsatellites (around 5% of the value for mtDNA). Male gene flow was estimated to be about 20-30 times higher than female gene flow at this scale. At a larger spatial scale, very strong genetic differentiation for both markers was observed between localities - even within a single region. Female dispersal is nonexistent at these scales and male dispersal is very restricted, especially between regions. The phylogeographical structure of the mtDNA haplotypes as well as the very low genetic diversity of mtDNA within localities indicate that new sites are colonized by a single migration event from adjacent localities, followed by successive colony fissions. These patterns of genetic variability and differentiation agree with what is theoretically expected when colonization events are kin-structured and when, following colonization, dispersion is mainly performed by males.     

A clonal analysis of the roles of somatic cells and germ line during oogenesis in Drosophila

In order to test whether particular female sterile mutations block functions which normally occur in somatic or germ line derivatives, clones homozygous for each mutation were X-ray induced in heterozygous females. Using the germ line-dependent egg marker, fs(1)K10, it was possible to identify the eggs derived from clones which had been induced in the germ line. Mutations were classified as germ line dependent when these eggs also showed the phenotype associated with the female sterile mutation. Two mutations which caused early abnormalities in oogenesis (fs(1)116, fs(1)1304) were shown to affect germ cells, whereas two mutations which caused egg retention (fs(1)462, fs(1)1001) were somatically dependent. A mutation altering egg dimensions without affecting egg volume (short egg) was also shown to depend on somatic cells in the ovary. With one exception (fs(1)K4), mutations which caused production of fragile, collapsed eggs (fs(1)180, fs(1)473, fs(1)384, and fs(1)1163) were somatically dependent. Patches of mutant fs(1)384 morphology were found in the chorions of the eggs not derived from germ line clones. These patches are interpreted as being caused by homozygous clones in the somatically derived follicle cell epithelium and suggest that fs(1)384 affects processes occurring in these cells during the synthesis of the egg coverings.     

Interaction of the Bordetella pertussis filamentous hemagglutinin with heparin

Heparin, a glycosaminoglycan synthesized in connective tissue-mast cells, appeared to inhibit the hemagglutination of rabbit erythrocytes induced by the filamentous hemagglutinin (FHA), a major adhesin of Bordetella pertussis. This inhibition suggested an interaction of heparin with the FHA region responsible for the hemagglutination activity. FHA-heparin interactions may play a role in bacterial attachment and persistence in the lungs during human pertussis. To confirm a direct FHA-heparin interaction, heparin was used as ligand in an affinity chromatography procedure. This technique allowed to purify FHA directly from the bacterial culture medium in a single-step using heparin-Sepharose CL-6B or Zetaffinity heparin 60 disks. The purified FHA was highly immunoreactive with anti-FHA monoclonal antibodies and showed no signs of degradation after 15 successive cycles of freezing-thawing. The described purification method is simple, and suitable for the rapid preparation of FHA.     

Induction of apoptosis by protein kinase C pseudosubstrate lipopeptides in several human cells

Summary Intracellular enzymes or receptors are interesting targets for the pharmacomodulation of cellular metabolism. We have previously shown that modification of relatively long peptides by a palmitoyl-lysine residue could facilitate their delivery into the cytoplasm of living cells. Several peptides containing pseudosubstrate sequences of protein kinase C (PKC) have been evaluated for their ability to modulate phosphorylation of model substrate, neuronal morphology or tumor necrosis factor secretion. In this work we have evaluated the effect of palmitoyl-modified PKC-pseudosubstrate peptides on induction of apoptosis. We have established that these peptides are able to induce apoptosis in different human cell types (primary fibroblasts, T- and B-lymphocyte cell lines) as assessed by (terminal deoxynucleotidyl transferase dUTP nick-end labelling) and DNA fragmentation. In contrast, control peptides (non-lipidic PKC-pseudosubstrate peptides and irrelevant lipopeptides) had no or little effect on programmed cell death. This work highlights the pharmacological interest of lipopeptides and argues in favor of the potential role of PKC(s) in the cell death machinery. K. Thiam and E. Loing have contributed equally to this work.     

Kallikrein activates bradykinin B2 receptors in absence of kininogen

Kallikreins cleave plasma kininogens to release the bioactive peptides bradykinin (BK) or kallidin (Lys-BK). These peptides then activate widely disseminated B2 receptors with consequences that may be either noxious or beneficial. We used cultured cells to show that kallikrein can bypass kinin release to activate BK B2 receptors directly. To exclude intermediate kinin release or kininogen uptake from the cultured medium, we cultured and maintained cells in medium entirely free of animal proteins. We compared the responses of stably transfected Chinese hamster ovary (CHO) cells that express human B2 receptors (CHO B2) and cells that coexpress angiotensin I-converting enzyme (ACE) as well (CHO AB). We found that BK (1 nM or more) and tissue kallikrein (1-10 nM) both significantly increased release of arachidonic acid beyond unstimulated baseline level. An enzyme-linked immunoassay for kinin established that kallikrein did not release a kinin from CHO cells. We confirmed the absence of kininogen mRNA with RT-PCR to rule out kininogen synthesis by CHO cells. We next tested an ACE inhibitor for enhanced BK receptor activation in the absence of kinin release and synthesized an ACE-resistant BK analog as a control for these experiments. Enalaprilat (1 microM) potentiated kallikrein (100 nM) in CHO AB cells but was ineffective in CHO B2 cells that do not bear ACE. We concluded that kallikrein activated B2 receptors without releasing a kinin. Furthermore, inhibition of ACE enhanced the receptor activation by kallikrein, an action that may contribute to the manifold therapeutic effects of ACE inhibitors.     

Cadmium induces caspase-independent apoptosis in liver Hep3B cells: role for calcium in signaling oxidative stress-related impairment of mitochondria and relocation of endonuclease G and apoptosis-inducing factor

Cadmium-induced cellular toxicity has been related to necrosis and/or caspase-dependent apoptosis. In the present study, we show that, on cadmium exposure, the human hepatocarcinoma Hep3B cells undergo caspase-independent apoptosis associated with nuclear translocation of endonuclease G and apoptosis-inducing factor, two mitochondrial apoptogenic proteins. Release of these proteins is likely related to calcium-induced alteration of mitochondrial homeostasis. Indeed, it was first preceded by a rapid and sustained increase in cytoplasmic calcium and then by a coincident loss in mitochondrial membrane potential and production of reactive oxygen species. Bapta-AM (acetoxymethyl ester of 5, 5′-dimethyl-bis (o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid), a calcium chelator, blocked all these events and prevented cadmium-induced apoptosis. Production of reactive oxygen species was inhibited by ruthenium red and rotenone, two mitochondrial inhibitors, and by diphenyleneiodonium, a flavoprotein inhibitor, which also prevented both loss in mitochondrial membrane potential and apoptosis. In addition, Bapta-AM and diphenyleneiodonium were found to almost totally block decreased expression of the mitochondrial anti-apoptotic nuclear factor-κB-regulated bcl-x_L protein in cadmium-treated cells. Taken together, our results show that cadmium induces Hep3B cells apoptosis mainly by calcium- and oxidative stress-related impairment of mitochondria, which probably favors release of apoptosis-inducing factor and endonuclease G.     

Modulation of the Suppressor of fused protein regulates the Hedgehog signaling pathway in Drosophila embryo and imaginal discs

The Suppressor of fused (Su(fu)) protein is known to be a negative regulator of Hedgehog (Hh) signal transduction in Drosophila imaginal discs and embryonic development. It is antagonized by the kinase Fused (Fu) since Su(fu) null mutations fully suppress the lack of Fu kinase activity. In this study, we overexpressed the Su(fu) gene in imaginal discs and observed opposing effects depending on the position of the cells, namely a repression of Hh target genes in cells receiving Hh and their ectopic expression in cells not receiving Hh. These effects were all enhanced in a fu mutant context and were suppressed by cubitus interruptus (Ci) overexpression. We also show that the Su(fu) protein is poly-phosphorylated during embryonic development and these phosphorylation events are altered in fu mutants. This study thus reveals an unexpected role for Su(fu) as an activator of Hh target gene expression in absence of Hh signal. Both negative and positive roles of Su(fu) are antagonized by Fused. Based on these results, we propose a model in which Su(fu) protein levels and isoforms are crucial for the modulation of the different Ci states that control Hh target gene expression.