Polarisation des ARNm maternels et des régulateurs de l'initiation de la traduction dans le cortex des oeufs et embryons précoces d'ascidie

Ascidians (urochodates: Ciona intestinalis and Phallusia mammillata ) are considered the closest invertebrate ancestors of vertebrates. They develop into simple tadpoles made of 2600 cells harboring a notochord and a dorsal nerve chord. At the turn of the 20th century, the ascidians model contributed much to the idea that oocytes contained localized determinants of development. This led to the concept of mosaic development. Today we know that some of these determinants are maternal RNAs localized in the egg cortex. Ascidian determinant RNAs are part of a large family of maternal RNAs called postplasmic/PEM RNAs. They include about 40 transcrips classified as Type I (localized before fertilization) and Type II (localized after fertilization). Type I postplasmic/PEM RNAs are relocalized after fertilization in a stereotyped fashion such that they concentrate in the posterior region of the zygote and form a compact cortical zone in the 2 posterior vegetal blastomeres at the 8 cell stage called the CAB (Centrosome Attracting Body). The large numbers of postplasmic/PEM RNAs identified, the synchronicity and abundance of embryos, and the ability to isolate cortical fragments retaining the RNAs make the ascidian model particularly attractive for studying localization and cortical anchorage of polarized RNAs. It is possible to operate a first classification of postplasmic/PEM RNAs based on their localization in cells in the tail of the tadpole. The determinant Macho1 and PEM1 segregate in 2 B8.11 cells with somatic destinies while RNAs such as the germ plasm marker Vasa is in addition localized in 2 B.12 cells precursors of the germ line. The 3 UTR regions of postplasmic/PEM RNA are considered to contain part of the code for localization and anchorage. In Ascidians as in the amphibian Xenopus these regions are characterized by frequent xCACx repeats. Using this criteria to screen 3 UTRs of all Ciona intestinalis genes, we confirmed that frequent xCACx repeats constituted a predictive signature for postplasmic/PEM RNAs and found 2 new members (MnK et PSD). We have also identified subcellular anchorage sites of postplasmic/PEM RNAs using high resolution immuno-in situ localization techniques in oocytes and embryos as well as in cortical fragments isolated from them. The RNA determinants Macho1 and PEM1 are associated with a sub-domain of cortical Endoplasmic Reticulum (cER) while Vasa, PEM3, POPK RNAs are localized in granules (putative germ granules). This demonstrates that although at low resolution these RNAs appear co-localized in the cortex they have in fact different structures of anchorage. This allows to propose that postplasmic/PEM RNAs belong to 2 categories: a category ( Vasa-Type ) associated with granules and segregating in both 8.11 and 8.12 cells and a category ( Macho1-Type ) associated to cER which segregates only into B8.11 cells. We also investigated whether, as in nerve terminals, the translation machinery was also polarized in oocytes and embryos. Indeed we found that some factors (PAPB) and regulators (phosporylated forms of MnK, 4EBP et S6Kinase ) of translation initiation were co-localized with postplasmic/PEM RNAs associated with the cER (Macho1 and PEM1 RNAs). The fact that regulators such as MnK and S6Kinase change phosphorylation status after fertilization suggests that they control translation initiation of determinants RNAs in the cortex following egg activation.NICE-BU Sciences (060882101) / SudocSudocFranceF