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CD9, a member of the tetraspanin family, has been implicated in hematopoietic and leukemic stem cell homing. We investigated the role of CD9 in the dissemination of B acute lymphoblastic leukemia (B-ALL) cells, by stably downregulating CD9 in REH and NALM6 cells. CD9 expression was associated with higher levels of REH cell adhesion to fibronectin and C-X-C motif chemokine receptor 4 (CXCR4)-mediated migration. Death occurred later in NOD/SCID mice receiving REH cells depleted of CD9 for transplantation than in mice receiving control cells. After C-X-C motif chemokine ligand 12 (CXCL12) stimulation, CD9 promoted the formation of long cytoplasmic actin-rich protrusions. We demonstrated that CD9 enhanced RAC1 activation, in both REH cells and blasts from patients. Conversely, the overexpression of a competing CD9 C-terminal tail peptide in REH cytoplasm decreased RAC1 activation and cytoplasmic extension formation in response to CXCL12. Finally, the inhibition of RAC1 activation decreased migration in vitro, and the depletion of RAC1 protein from transplanted REH cells increased mouse survival. Furthermore, a testis-conditioned medium induced the migration of REH and NALM6 cells, and this migration was impeded by an anti-CD9 antibody. The level of CD9 expression also influenced the homing of these cells in mouse testes. These findings demonstrate, for the first time, that CD9 plays a key role in the CXCR4-mediated migration and engraftment of B-ALL cells in the bone marrow or testis, through RAC1 activation.

CD9, a member of the tetraspanin family, has been implicated in hematopoietic and leukemic stem cell homing. We investigated the role of CD9 in the dissemination of B acute lymphoblastic leukemia (B-ALL) cells, by stably downregulating CD9 in REH and NALM6 cells. CD9 expression was associated with higher levels of REH cell adhesion to fibronectin and C-X-C motif chemokine receptor 4 (CXCR4)-mediated migration. Death occurred later in NOD/SCID mice receiving REH cells depleted of CD9 for transplantation than in mice receiving control cells. After C-X-C motif chemokine ligand 12 (CXCL12) stimulation, CD9 promoted the formation of long cytoplasmic actin-rich protrusions. We demonstrated that CD9 enhanced RAC1 activation, in both REH cells and blasts from patients. Conversely, the overexpression of a competing CD9 C-terminal tail peptide in REH cytoplasm decreased RAC1 activation and cytoplasmic extension formation in response to CXCL12. Finally, the inhibition of RAC1 activation decreased migration in vitro, and the depletion of RAC1 protein from transplanted REH cells increased mouse survival. Furthermore, a testis-conditioned medium induced the migration of REH and NALM6 cells, and this migration was impeded by an anti-CD9 antibody. The level of CD9 expression also influenced the homing of these cells in mouse testes. These findings demonstrate, for the first time, that CD9 plays a key role in the CXCR4-mediated migration and engraftment of B-ALL cells in the bone marrow or testis, through RAC1 activation.

This study examined the function of H3 and H4 tail domains in replication-dependent chromatin assembly. Results show distinct functions of H3 and H4 tails in nuclear import and chromatin assembly. Further investigations show that H4 diacetylation is essential but not sufficient for nuclear import, as preventing Hat1 binding impedes histone transport in nuclei.

While specific posttranslational modification patterns within the H3 and H4 tail domains are associated with the S-phase, their actual functions in replication-dependent chromatin assembly have not yet been defined. Here we used incorporation of trace amounts of recombinant proteins into naturally synchronous macroplasmodia of Physarum polycephalum to examine the function of H3 and H4 tail domains in replication-coupled chromatin assembly. We found that the H3/H4 complex lacking the H4 tail domain was not efficiently recovered in nuclei, whereas depletion of the H3 tail domain did not impede nuclear import but chromatin assembly failed. Furthermore, our results revealed that the proper pattern of acetylation on the H4 tail domain is required for nuclear import and chromatin assembly. This is most likely due to binding of Hat1, as coimmunoprecipitation experiments showed Hat1 associated with predeposition histones in the cytoplasm and with replicating chromatin. These results suggest that the type B histone acetyltransferase assists in shuttling the H3/H4 complex from cytoplasm to the replication forks.

Duchenne muscular dystrophy is the most common and severe form of muscular dystrophy, and although the genetic basis of this disease is well defined, the overall mechanisms that define its pathogenesis remain obscure. Alterations in individual signaling pathways have been described, but little information is available regarding their putative implications in Duchenne muscular dystrophy pathogenesis. Here, we studied the status of various major signaling pathways in the Golden Retriever muscular dystrophy dog that specifically reproduces the full spectrum of human pathology. Using antibody arrays, we found that Akt1, glycogen synthase kinase-3β (GSK3β), 70-kDa ribosomal protein S6 kinase (p70S6K), extracellular signal-regulated kinases 1/2, and p38δ and p38γ kinases all exhibited decreased phosphorylation in muscle from a 4-month-old animal with Golden Retriever muscular dystrophy, revealing a deep alteration of the phosphatidylinositol 3-kinase (PI3K)/Akt and mitogen-activated protein kinase pathways. Immunohistochemistry analysis revealed the presence of muscle fibers exhibiting a cytosolic accumulation of Akt1, GSK3β, and phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase (PTEN), an enzyme counteracting PI3K-mediated Akt activation. Enzymatic assays established that these alterations in phosphorylation and expression levels were associated with decreased Akt and increased GSK3β and PTEN activities. PTEN/GSK3β-positive fibers were also observed in muscle sections from 3- and 36-month-old animals, indicating long-term PI3K/Akt pathway alteration. Collectively, our data suggest that increased PTEN expression and activity play a central role in PI3K/Akt/GSK3β and p70S6K pathway modulation, which could exacerbate the consequences of dystrophin deficiency.

Nous montrons que C1-génériquement les difféomorphismes symplectiques d'une variété compacte sont transitifs. Sur les variétés symplectiques non-compactes, nous montrons que les orbites génériques d'un difféomorphisme générique ne sont pas bornées. L'outil principal est un lemme de connexion pour les pseudo-orbites des difféomorphismes symplectiques. Pour cela nous donnons des conditions générales sur un espace de difféomorphismes, en particulier au voisinage des orbites périodiques, qui permettent d'obtenir le lemme de connexion pour les pseudo-orbites.

Orobanche ramosais a parasitic Angiosperm responsible for severe yield losses in several economical crops. It is a serious threat in oilseed rape in France and Morocco and is appearing in carrot crops in Morocco. In this study, several varieties of oilseed rape and carrot were screened in order to identify resistant cultivars and to characterize the resistance mechanisms involved. All the 15 oilseed rape varieties tested were susceptible. In carrot, the varieties 'Colmar à coeur rouge' and 'Nantaise demi-longue' were susceptible, whereas 'Palaiseau' and 'Buror' were resistant. In the susceptible 'Colmar à coeur rouge' carrot no defence reactions were found and the development of the parasite inhibited carrot tap root formation. In the resistant carrot varieties, the parasite germinated, became attached to the host root but became necrotic before emergence. In 'Buror' carrot, formation of a mechanical barrier was associated with the restriction to the cortex of the parasite. In maize cv. 'Vigni', a non-host of O. ramosa, thickening of xylem vessels, cell divisions in the central cylinder and formation of an encapsulation layer were observed in association with restricted development of Orobanchehaustoria.

We prove that if (M,\omega) is a connected and compact four-dimensional symplectic manifold, there exist three open sets U_1, U_2, U_3 of {\rm Diff}^1_{\omega}(M) (for the C^1 topology) such that: U_1\cup U_2\cup U_3 is dense in {\rm Diff}^1_{\omega}(M);f\in U_1 if and only if f is Anosov and transitive;f\in U_2 if and only if f is partially hyperbolic; andf\in U_3 if and only if f has a stable completely elliptic periodic point.

Dans cet article nous allons commencer par démontrer un résultat perturbatif, nous en déduirons différentes manières de créer des connexions à l'aide de perturbations de difféomorphismes petites en topologie C^1. Ensuite, nous nous intéresserons plus spécifiquement à la place des variétés stables des points périodiques hyperboliques ainsi qu'à la fermeture d'orbites récurrentes en un sens faible; puis, nous ferons une étude plus fine des difféomorphismes de classe C^1 ‘génériques’, montrant en particulier l'existence d'attracteurs, au sens de Liapounov, transitif en un certain sens faible ; enfin, nous expliquerons ce qui se passe pour les difféomorphismes qui préservent une forme volume finie.

Using certain quadratic forms associated to symplectic endomorphisms which we compare with the Clarke-Ekeland dual action functional, we prove: THEOREM. — Let H be a C2-Hamiltonian defined on R2n, strictly convex, proper and invariant under a certain symplectic rational positive and non-degenerate rotation (this is defined in the introduction); then, every hypersurface of H contains a completely elliptic periodic orbit. This generalizes the result of G. Dell'Antonio, B. D'Onofrio and I. Ekeland contained in [1].

No relevant conflicts of interest to declare.

No relevant conflicts of interest to declare.

Introduction and hypothesis :B-acute lymphoblastic leukemia (B-ALL) expressing the fusion transcript ETV6/RUNX1 is one of the most frequent subtype of childhood leukemia. This disease is associated with a good survival rate but 5 to 10% of patients relapse more than five years after treatment most notably in extra-hematopoietic sanctuary sites like testis.

Introduction and objectives:

CD9 is a membrane protein, member of the tetraspanin family. Recent publications have reported the role of CD9 on engraftment of hematopoietic stem cells, and on cancer stem cell potential. The expression of CD9 has been correlated to the risk of metastases and to a poor clinical outcome in various types of cancer. Surprisingly, CD9 protein is downregulated in ETV6/RUNX1 pre-B acute lymphoblastic leukemia. The purpose of our study is to investigate the effect of CD9 expression on migration and engraftment abilities of pre-B lymphocytes.The CD9-positive Nalm6 and REH (ETV6/RUNX1) pre-B cells were used. By lentiviral transduction of shRNA targeting mRNA, we generated Nalm6 and REH cell lines depleted in CD9 protein. Engraftment tests were performed in vivo using Nod Scid Gamma immunodeficient mice. REH and Nalm6 cells were detected in bone marrow by CD10 and respectively CD19 or HLA-DR labelling. Ability of the different cell lines to adhere on fibronectin and to migrate through double chambers system in response to CXCL12 were measured in vitro. We also investigated the presence of membrane villosities on REH and REH shCD9 cell surface by scanning electron microscopy. Finally, F-actin polymerization after CXCL12 stimulation was measured by rhodamin-phalloidin labelling.In vivo engrafments tests showed that the human cells detected in bone marrow is strongly enriched in CD9 positive cells compared to the initially injected population. This result suggests that CD9 facilitates pre-B lymphobasts homing. An in vitro analysis of adhesion on fibronectin demonstrated that cellular adhesion is dependent on membrane expression of CD9. As well, the more CD9 is expressed, the higher the migration rate in response to CXCL12 chemokine is. The analysis of membrane villosities on REH cell surface revealed that cells over expressing CD9 had longer villosity than shCD9 transducted cell lines. Moreover, F-actin labelling after CXCL12 stimulation showed an increased F-actin polymerization in CD9-positive cells and the formation of actin extensions.We provide novel evidence that CD9 is a key player of pre-B lymphoblasts engraftment, adhesion and CXCL12 dependant migration. CD9 expression is related to actin remodelling. We are now investigating a potential link between CD9 and RAC1 activation in response to CXC12. Therefore, the expression level of CD9 could impact leukemic blasts abilities to spread and be responsible of relapses.This work is supported by CNRS, University of Rennes 1, University Hospital of Rennes, la Ligue Régionale contre le Cancer (committee 22, 35 and 56) (MPA, VG, MBT), SFR Biosit UMS 3480 (VG, MBT), Association Laurette Fugain (VG) and Europe Career Integration Grant (MBT).No relevant conflicts of interest to declare.

CD9 is a membrane protein, member of the tetraspanin family. Recent publications have reported the role of CD9 on engraftment of hematopoietic stem cells, and on cancer stem cell potential. The expression of CD9 has been correlated to the risk of metastases and to a poor clinical outcome in various types of cancer. Surprisingly, CD9 protein is downregulated in ETV6/RUNX1 pre-B acute lymphoblastic leukemia. The purpose of our study is to investigate the effect of CD9 expression on migration and engraftment abilities of pre-B lymphocytes.

Germination, attachment to host root and growth of Striga hermonthica (Del.) Benth. seeds and seedlings were studied in in vitro co-culture w ith Sorghum bicolor (L.) Moench and in pot experiments. Two varieties, the resistant Framida and the susceptible CK-60B, were used. Histological, morphological and physiological studies revealed the key stages of resistance mechanisms involved. Resistance of Framida to Striga does not occur at the germination or the attachment stages, since its roots do not support fewer Striga than does CK-60B. As Framida roots support the lowest number of young Striga stems with scale leaves, its resistance appears to occur during the establishment of a functional haustorium. Metabolite uptake by the haustorium and growth rate of the young parasite were lower on Framida roots than on CK-60B roots, even when similarly developed haustoria were compared. Furthermore, at a later stage of infestation, significant accumulation of a coloured material likely to be rich in phenolic compounds was observed in and around Framida conductive tissues, but not CK-60B tissues. These features indicate the involvement of at least three steps in development of resistance in Framida roots: the first is linked to a partial inhibition of development of the young haustorium; the second could play a role in the physiological events that decrease nutrient translocation towards the haustoria; and the last seems to be associated with the accumulation of a coloured phenolic-like material. Keywords: Striga hermonthica, Sorghum bicolor (L.) Moench, haustorium, host–parasite interaction, resistance.