The long term objective of this project is to understand the processes that control mammalian cell growth, and how these processes are affected in the development of tumors. This includes the mechanisms by which growth factors can bind to the cell surface, transmit signals to the intracellular compartment and ultimately cause the cell to progress through mitosis. Mitogenic activation of quiescent cells involves multiple interacting signalling pathways that result in a number of immediate or early responses. In addition, other responses occur many hours after activation, after many of the early responses have subsided. Some of these late responses are obligatory for the cell to enter into the DNA synthetic phase of the cell cycle and an important, but little understood, question is the sequence of events that lead to their activation. This is a proposal to study the regulation of a vital late response to growth factor action: the elevated expression of transferrin receptors. Mitogenic stimulation of cells leads to a dramatic increase in the level of this receptor on the cell surface which occurs just before the onset of S phase. If the receptors are blocked at the cell surface, cell growth is prevented and the cells accumulate near the G1/S phase boundary. The increased expression of transferrin receptors that occurs during cell proliferation is associated with the transcriptional activation of its gene. Thus by examining the mechanisms that lead to this activation, some of the processes and pathways involved in regulating late events can be determined. First, the mitogen responsive elements in the regulatory region of the gene will be precisely mapped using a series of genetic constructs containing deletions an site- specific mutations. The transacting factors that act at these sites will be identified and then purified. The purified factors will be used to prepare immunological and genetic probes for the factors. Finally, using these probes and a number of specific growth factors and pharmacological reagents we will determine the effects of various signalling pathways on expression of the receptor gene and on the synthesis, transport, and post- translational modification of the transacting factors involved.
| Hirsch, S; Miskimins, W K (1995) Mitogen induction of nuclear factors that interact with a delayed responsive region of the transferrin receptor gene promoter. Cell Growth Differ 6:719-26 |
| Ouyang, Q; Bommakanti, M; Miskimins, W K (1993) A mitogen-responsive promoter region that is synergistically activated through multiple signalling pathways. Mol Cell Biol 13:1796-804 |
| Ouyang, Q; Miskimins, W K (1992) A supercoil-dependent structural alteration within the regulatory region of the human transferrin receptor gene. Biochem Biophys Res Commun 183:62-9 |
| Miskimins, W K (1992) Interaction of multiple factors with a GC-rich element within the mitogen responsive region of the human transferrin receptor gene. J Cell Biochem 49:349-56 |
