Giulia Costa

Functional validation of the anaplastic lymphoma kinase signature identifies CEBPB and Bcl2A1 as critical target genes

Anaplastic large cell lymphomas (ALCLs) represent a subset of lymphomas in which the anaplastic lymphoma kinase (ALK) gene is frequently fused to the nucleophosmin (NPM) gene. We previously demonstrated that the constitutive phosphorylation of ALK chimeric proteins is sufficient to induce cellular transformation in vitro and in vivo and that ALK activity is strictly required for the survival of ALK-positive ALCL cells. To elucidate the signaling pathways required for ALK-mediated transformation and tumor maintenance, we analyzed the transcriptomes of multiple ALK-positive ALCL cell lines, abrogating their ALK-mediated signaling by inducible ALK RNA interference (RNAi) or with potent and cell-permeable ALK inhibitors. Transcripts derived from the gene expression profiling (GEP) analysis uncovered a reproducible signature, which included a novel group of ALK-regulated genes. Functional RNAi screening on a set of these ALK transcriptional targets revealed that the transcription factor C/EBPbeta and the antiapoptotic protein BCL2A1 are absolutely necessary to induce cell transformation and/or to sustain the growth and survival of ALK-positive ALCL cells. Thus, we proved that an experimentally controlled and functionally validated GEP analysis represents a powerful tool to identify novel pathogenetic networks and validate biologically suitable target genes for therapeutic interventions.

Control of Plasmodium falciparum erythrocytic cycle : gamma-delta T cells target the red blood cell-invasive merozoites

The control of Plasmodium falciparum erythrocytic parasite density is essential for protection against malaria, because it prevents pathogenesis and progression toward severe disease. P falciparum blood-stage parasite cultures are inhibited by human Vγ9Vδ2 γδ T cells, but the underlying mechanism remains poorly understood. Here, we show that both intraerythrocytic parasites and the extracellular red blood cell-invasive merozoites specifically activate Vγ9Vδ2 T cells in a γδ T cell receptor-dependent manner and trigger their degranulation. In contrast, the γδ T cell-mediated antiparasitic activity only targets the extracellular merozoites. Using perforin-deficient and granulysin-silenced T-cell lines, we demonstrate that granulysin is essential for the in vitro antiplasmodial process, whereas perforin is dispensable. Patients infected with P falciparum exhibited elevated granulysin plasma levels associated with high levels of granulysin-expressing Vδ2(+) T cells endowed with parasite-specific degranulation capacity. This indicates in vivo activation of Vγ9Vδ2 T cells along with granulysin triggering and discharge during primary acute falciparum malaria. Altogether, this work identifies Vγ9Vδ2 T cells as unconventional immune effectors targeting the red blood cell-invasive extracellular P falciparum merozoites and opens novel perspectives for immune interventions harnessing the antiparasitic activity of Vγ9Vδ2 T cells to control parasite density in malaria patients.

Phosphoantigen Burst upon Plasmodium falciparum Schizont Rupture Can Distantly Activate Vγ9Vδ2 T Cells

ABSTRACT Malaria induces potent activation and expansion of the Vγ9Vδ2 subpopulation of γδT cells, which inhibit the Plasmodium falciparum blood cycle through soluble cytotoxic mediators, abrogating merozoite invasion capacity. Intraerythrocytic stages efficiently trigger Vγ9Vδ2 T-cell activation and degranulation through poorly understood mechanisms. P. falciparum blood-stage extracts are known to contain phosphoantigens able to stimulate Vγ9Vδ2 T cells, but how these are presented by intact infected red blood cells (iRBCs) remains elusive. Here we show that, unlike activation by phosphoantigen-expressing cells, Vγ9Vδ2 T-cell activation by intact iRBCs is independent of butyrophilin expression by the iRBC, and contact with an intact iRBC is not required. Moreover, blood-stage culture supernatants proved to be as potent activators of Vγ9Vδ2 T cells as iRBCs. Bioactivity in the microenvironment is attributable to phosphoantigens, as it is dependent on the parasite DOXP pathway, on Vγ9Vδ2 TCR signaling, and on butyrophilin expression by Vγ9Vδ2 T cells. Kinetic studies showed that the phosphoantigens were released at the end of the intraerythrocytic cycle at the time of parasite egress. We document exquisite sensitivity of Vγ9Vδ2 T cells, which respond to a few thousand parasites. These data unravel a novel framework, whereby release of phosphoantigens into the extracellular milieu by sequestered parasites likely promotes activation of distant Vγ9Vδ2 T cells that in turn exert remote antiparasitic functions.

Analysis of the reaction π+p → ρ0Δ++ at 11.7 GeV/c incident momentum

SummaryA model-independent analysis is made to determine the relative contribution of natural and unnatural parity exchange in the reactions (1) π+p→ρ0Δ++ at 11.7 GeV/c and (2) π−p→ρ0n at 11 GeV/c. A simple evasive Regge-pole model with only π-trajectory exchange has been applied to the data of reaction (1) and then to the combined data of reactions (1) and (2). A satisfactory fit to the data is obtained. The parameters determined in fitting reaction (1) have been used to make predictions for the same reaction at 8, 13.1 and 18.5 GeV/c. A comparison with experimental data at these momenta has been made. Considering the severe nature of this test for the evasive model there is altogether satisfactory agreement between the predictions and the data.RiassuntoUn’analisi che prescinde da ogni modello specifico permette di separare il contributo relativo degli scambi di parità naturale e non naturale nelle reazioni (1) π+p → ρ0Δ++ a 11.7 GeV/c e (2) π−p → ρ0n a 11.2 GeV/c. In base ai risultati di tale analisi, dapprima si interpreta la reazione (1) mediante un semplice modello di Regge con scambio di un π di tipo evasivo; poi si interpretano i dati di entrambe le reazioni (1) e (2) congiuntamente, ottenendo risultati soddisfacenti. I parametri così determinati si usano quindi per ottenere le dovute previsioni sull’andamento della sezione d’urto differenziale dσ/d|t| per la reazione (1) da confrontare con dati esistenti a 8, 13.1 e 18.5 GeV/c. Considerato il carattere particolarmente severo della verifica cui il modello di Regge di tipo evasivo è qui sottoposto, l’accordo tra dati sperimentali e previsioni teoriche può ritenersi soddisfacente. Tuttavia il modello evasivo non è in grado di riprodurre il persistente picco in avanti presentato a tutte le energie dai dati sperimentali.РеэюмеПроводится аналиэ, не эависяший от модели, для определения относительного вклада обмена с естественной и неестественной четностью в реакциях (1) π+p → ρ0Δ++ при 11.7 ГзВ/с и (2) π−p → ρ0n при 11 ГзВ/с. Была применена простая модель « неуловимых » полюсов Редже с обменом единственной π-траекторией к данным реакции (1), а эатем к комбинированным данным реакций (1) и (2). Получается удовлетворительное соответствие. Параметры, полученные иэ подгонки первой реакции, были испольэованы для получения предскаэаний для той же реакции при начальных импульсах 8, 13.1 и 18.5 ГзВ/с. Было проведено сравнение с зкспериментальными данными при зтих импульсах. Учитывая строгий характер зтой проверки модели « неуловимых » полюсов, получается удовлетворительное согласие между предскаэаниями и полученными данными.