Nuclear reprogramming in Xenopus using non-aqueous oocyte nuclei. NCE of 9 months approved, orig end date 30/09/09. SA (360G-Wellcome-079948_Z_06_Z)

We aim to identify some of the molecules and mechanisms by which an oocyte can reprogram the nuclei of readily accessible adult somatic cells to express embryo- or stem-cell specific genes such as Oct4 and Nanog. This is a step towards our ultimate aim of deriving rejuvenated cells for therapeutic replacement, starting with adult cells of the same individual, and thereby avoiding the need for immunosuppression. We inject mammalian somatic cell nuclei into the germinal vesicle (large specializ ed nucleus) of an amphibian oocyte and monitor nuclear reprogramming by the transcriptional activation of genes. In this project, we plan: (i) to do protein knockdown experiments, to test the necessity of identified oocyte chromosomal proteins for nuclear reprogramming; (ii) to do real-time imaging of injected nuclei in live germinal vesicles to relate the loss and gain of known proteins from and by somatic nuclei and their necessity for reprogramming; (iii) to use oocyte and egg extracts to id entify, by fractionation, so far unknown components of oocytes that are required for nuclear reprogramming. Having identified candidate reprogramming components of Xenopus oocytes, we expect to recognize mouse (or human) homologues, and to test these for function in mouse (or human) embryonic stem cells.