| Summary: | Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, February 2002. === Includes bibliographical references. === The retinoblastoma tumor suppressor and the closely related p107 and p130 proteins play important roles in the regulation of both tumorigenesis and tissue-specific differentiation. To determine the role of Rb in development more precisely, we analyzed chimeric embryos and adults made with marked Rb-/- cells. We demonstrated that although brains of chimeric embryos exhibited extensive ectopic S-phase entry, unlike in germline Rb-/- littermates, in these chimeras the majority of Rb-deficient cells survived and differentiated into neuronal fates. Therefore, the role of Rb during development can be divided into a cell-autonomous function in exit from the cell cycle and a non-cell autonomous role in the suppression of apoptosis and induction of differentiation. Rb-/- ES cells were also able to efficiently differentiate into neurons in vitro. However, similarly to the situation observed in the chimeric embryos, Rb-/- neuronal cells failed to properly exit the cell cycle and continued to progress through the S-phase, while displaying fully differentiated neuronal morphology and expressing mature neuronal markers. Therefore, Rb function is not required for the commitment to the neuronal lineage or progression of neuronal differentiation but necessary for the cell cycle exit of differentiating neurons. In agreement, in wild-type differentiating ES cells, activity of the retinoblastoma protein (pRB) was increased specifically at the time of neuronal precursor cell cycle exit. === (cont.) In differentiating Rb-/- ES cells, levels of the pRB-related p107 protein were specifically elevated, suggesting that it might be substituting for pRB to allow efficient neuronal differentiation. In order to determine if p107 and p130 proteins might play a role in tissue-specificity of tumorigenesis upon loss of Rb, we investigated the function of these proteins in the mouse pituitary tumors. Both p107 and p130 were present and active in primary pituitary tumors but levels of p107 were very low while protein levels if pocket-family targets, E2F 1 and E2F 4 were elevated. Although normally bound exclusively to E2F 4, in pituitary tumors p107 and p130 were able to form complexes also with E2F 1. Therefore, in the absence of pRB, p107 and p130 are able to bind and may regulate the function of the activating E2F species. Despite this partial substitution, p107 and p130 are unable to prevent pituitary tumorigenesis in Rb+'- mice, likely due to the elevated E2F expression. The levels of expression of the pRB- versus the E2F-family proteins in different tissues might be contributing to the sensitivity of these tissues to tumor development upon loss of Rb. === Marta M. Lipinski. === Ph.D.
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