JNK LEVELS AND APOTOSIS IN DEVELOPMENT OF GENITAL DISCS

NICOLÁS TOMASINI; NOELIA ROQUÉ; ANA MACÍAS

Los Cocos, Cordoba

Workshop; International Workshop “Latest Concepts in Developmental Biology”; 2006

The genital discs are originated by invagination and fusion of ventral cells from the eighth (A8), ninth (A9) and tenth (A10) abdominal segments. A particular sex regulation in combination with Hox genes determine complete different structures from each disc, constituting specific genetic contexts, this make them excellent models to analyse identical genetic condition in distinct genetic environments.Apoptosis have been demonstrated has a relevant role in the control of growth in the wing disc by the mechanism of cell competition. Also apoptosis is related with growth through the compensatory mechanism.JNK in Drosophila have been related as an executor of apoptosis. JNK is also involved in motility and adhesion properties. Also the JNK pathway has been implicated in the secretion of mitogen in the compensatory mechanism. How does JNK perform their functions, it has been proposed the levels and duration of JNK activation are critical. These two variables will depend in the balance among the activators and the negative regulators. There is only one negative regulator to the Drosophila JNK: the gene puckered (puc). puc levels depends in JNK as it is its transcriptional activator and might be one of its early genes.  We have previously reported the importance of JNK-dependent apoptosis in development of male genital discs, also the signal mechanism of PVF1/PVR (PDGF/PDGFR mammalian homologue) is genetically related as an activator of JNK.We analyzed the growth effects related with apoptosis in the genital disc, manipulating: • the apoptosis with apoptotic inhibitors (p35, DIAP1) and apoptotic genes (hid, rpr)• the apoptotic executor mechanism JNK with pathway activators (hepCA) and inhibitors (puc, bskDN)• both mechanism at the same time.To direct the expression of inhibitors and activators we used the Gal4–UAS method. We used two drivers: Abd-Bm-Gal4 (expressed in male in A8 cells and in some cells outside the A8; in the female preferentially expressed in A8 cells of external genitalia) and cad-Gal4 (specific to A10 cells).In the male A8 primordium, preventing apoptosis with one dose of p35 there is not growth of T8 and there is an incomplete terminalia rotation. Prevention of apoptosis with two doses of p35 produced a notable overgrowth indicating that compensatory mechanism exist and is relevant. These result also indicate it is necessary a control in growth of A8, because changing the conditions of JNK mechanism or apoptosis, we change the size of all disc.In the female A8 primordium, when we prevent the apoptosis with p35 we saw a growth which is notable incremented with two doses of p35. The same happen when we induced apoptosis and prevented it simultaneously (hid, p35) indicating that apoptosis normally occurs and a compensatory mechanism take place, in the control of size of external female genitalia. JNK pathway is not the apoptotic executor in this primordium due to when we block the pathway with puc there is no phenotype. But the JNK are able to execute apoptosis because hepCA produce the same phenotype of induction apoptosis with UAS hid. This indicates that high levels of JNK induce apoptosis.In the female A10 primordium, through driven expression of puc or p35, we observed that the JNK pathway and apoptosis are required for the development of anal plates, and prevent growth of some cells which origin we are going to determine. In males the same conditions are necessary for a complete terminalia rotation, and also as in females to prevent the growth of certain cells. We still do not know if the effects in the A10 are autonomous or non autonomous.In the males manipulating JNK with  puc we obtained the same phenotypic effects as the partial loss of PVF1/PVR suggesting that PVR is the principal activator of this pathway in A8. However in female A8 and A10 and male A10, PVR activates JNK but is not the main activator.