Abstract
In 2006, Yamanaka and co-workers announced a milestone finding that somatic cells can be reprogrammed into a pluripotent, embryonic-like state. These “artificial” pluripotent cells, called “induced pluripotent stem cells” (iPSCs), have a tremendous impact on the stem cell biology field, revolutionizing our view about cellular plasticity and regenerative medicine practice.
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Human iPSCs hold the potential of solving most of the issues concerning hESCs, especially because embryos are no longer needed and autologous transplantation could overcome the immune barrier. Therefore in the last few years an intensive research activity focused on the characterization of iPSCs in comparison to their natural ESCs counterparts at the functional, morphological and molecular levels. The most significant findings on the molecular characterization of iPSCs are reviewed in chapter 2. A detailed overview is provided for iPSCs analyses at the genomic, epigenomic, transcriptomic and, with particular emphasis, proteomic level. In the context of MS-based proteomics technology, its potential spectrum of applications to further characterize iPSCs and to decipher the reprogramming process are also considered. Indeed, although all these efforts have allowed to reach a general agreement that iPSCs are very similar to ESCs, the reprogramming process is still rather enigmatic. A better understanding of the mechanisms governing reprogramming would ultimately improve its efficiency and clarify the molecular basis of cellular plasticity. An in-depth molecular investigation of the reprogramming process of mouse embryonic fibroblasts (mEFs) into miPSCs is the focus of chapter 3, 4 and 5. In chapter 3, a multi-omics approach to characterize reprogramming at different molecular level is presented. This includes the profiling of mRNA, miRNA, lincRNA, histone marks, 5-methyl cytosine and protein levels. The integration of the proteomics data with all the other molecular layers, such as mRNA and lincRNA is discussed in detail. A comprehensive description of proteome adaptation during reprogramming is provided in chapter 4, where state of the art quantitative MS-based analysis is used to perform an in-depth time course protein profiling of such process. The main results deriving from all these analyses indicate that multiple routes can lead the reprogramming cell to a pluripotent state exist. The main features of these reprogramming cell populations are a marked divergence at the proteome level and the persistence of fibroblast epigenetic memory. One of the best known epigenetic mechanism that cells use to control gene expression and chromatin status is represented by the ensemble of post-translational modification that decorate histones. Further investigations concerning the MS-based study of the epigenetic state in reprogramming cells are provided in chapter 5, which is focused on the quantitative time course profiling of histone H4 PTM states.
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