The GTP-binding protein Giα translocates to kinetochores and regulates the M-G1 cell cycle transition of Swiss 3T3 cells

The receptor-generated signals that are responsible for driving the cell cycle are incompletely characterised in mammalian cells. It is clear, however, that the cellular messenger systems that stimulate DNA synthesis and mitosis are separable. These are interwoven with biochemical checkpoints that ensure that processes, such as chromosomal replication and microtubule attachment to duplicated chromosomes, are complete before the following phase of the cell cycle is initiated. In some cells, activation of DNA synthesis by factors such as LPA and serum has been shown to require the GTP-binding protein G(i). We have found that G(i) plays an additional role in mitosis activated by both 7-transmembrane receptors and tyrosine kinase receptors, and that this involves the translocation of the alpha-subunit of G(i) (G(i alpha)) to the nucleus. Here we show by confocal microscopy that G(i alpha) migrates to the nucleus near the onset of mitosis in serum-activated Swiss 3T3 cells and binds to the kinetochore region of replicated chromosomes. Inhibition of G(i) function with pertussis toxin had no effect on the induction of DNA synthesis by serum, but cell proliferation was inhibited. Flow cytometric analysis showed that this resulted from retardation of the transition through mitosis and into G(1). Additionally, pertussis toxin impaired the activity of p34(cdc2), a cyclin-dependent kinase involved in the transition from M-phase to G(1), but not the S-phase cyclin, cyclin E. These data show that the G-protein G(i) has a key role in the regulation of mitosis in fibroblasts. (C) 2000 Elsevier Science Inc. All rights reserved.