Cyclin D1 is an important regulator of G1 phase cell cycle progression and the target of multiple Signaling pathways. ERK and rac/cdc42 are both capable of inducing cyclin D1 gene expression, but we find that use of these pathways has distinct consequences for cells. First, ERK signaling leads to the induction of cyclin D1 in mid-G1 phase while Rac/Cdc42 signaling leads to cyclin D1 induction approximately 6-8 h earlier; S phase entry is proportionally affected. Second, ERK-dependent cyclin D1 expression requires the formation of stress fibers and leads to stress fiber-dependent proliferation while rac/cdc42-dependent cyclin D 1 expression and proliferation are stress fiber-independent. The Rho-Rho kinase pathway determines whether cyclin D1 expression occurs via ERK or rac/cdc42: Rho-Rho kinase signaling stimulates stress fiber formation and ERK-dependent cyclin D 1 expression while repressing rac/cdc42-dependent cyclin D1 expression. Our newest data identify LIM Kinase (LIMK) as the Rho kinase effector that determines whether ERK or Rac signaling is used to express cyclin D1. Not surprisingly, we find that LIMK is required for ERK-dependent cyclin D1 expression because it is required for actin polymerization and stress fiber formation. However, we also find that LIMK represses Rac-dependent induction of cyclin D1, and this effect is independent of stress fibers and actin polymerization. Moreover, this repressive effect requires localization of LIMK in the nucleus. These data reveal a completely new role for LIMK, and the major goal of this application is to understand this new biology.
Aim 1 will identify the Rac-activated cis-elements in the cyclin D1 promoter and the transcription factors that bind to them. Using this information, aim 2 will then determine how nuclear LIMK inhibits Rac-dependent activation of the cyclin D1 promoter.
Aim 3 will study the mechanisms regulating the shuttling of L1MK from cytoplasm to nucleus, characterizing potential regulatory phosphorylations, protein-protein interactions, and the responsible motifs on LIMK itself. Finally, aim 4 will use microarray technology to address the larger question of whether nuclear LIMK is a general repressor of Rac-dependent gene expression, whether it affects a subset of Rac-regulated genes, or whether its effect is specific to the cyclin D1 gene. Information from this aim will provide invaluable insight into common promoter motifs in genes that are reciprocally regulated by Rac and nuclear LIMK and set the stage for a broader analysis of LIMK effects on Rac-dependent gene expression.
| Yang, Chengfeng; Klein, Eric A; Assoian, Richard K et al. (2008) Heregulin beta1 promotes breast cancer cell proliferation through Rac/ERK-dependent induction of cyclin D1 and p21Cip1. Biochem J 410:167-75 |
| Klein, Eric A; Campbell, Latoya E; Kothapalli, Devashish et al. (2008) Joint requirement for Rac and ERK activities underlies the mid-G1 phase induction of cyclin D1 and S phase entry in both epithelial and mesenchymal cells. J Biol Chem 283:30911-8 |
| Klein, Eric A; Yang, Chengfeng; Kazanietz, Marcelo G et al. (2007) NFkappaB-independent signaling to the cyclin D1 gene by Rac. Cell Cycle 6:1115-21 |
