Our growth regulation research has been concerned with oncogenes as positive regulators and tumor suppressor genes as negative regulators of normal and neoplastic growth. The main current project is focused on the molecular biology of the tumor suppressor gene DLC1, including the targets that regulate it and the targets that it regulates. DLC1 is inactivated frequently in a wide range of tumors, but many aspects of its mechanism of action remain poorly understood. It negatively regulates Rho, via its Rho-GAP activity, but must encode other activities, as other Rho-GAPs are not known to be inactivated in cancer. One of our main hypotheses is that DLC1 is frequently inactivated in cancer because it encodes a multifunctional protein. In support of this possibility, we have previously determined that DLC1 interacts with: 1) members of the tensin gene family, via an N-terminal region of DLC1 for which no function had been previously identified; 2) with focal adhesion kinase (FAK) and with talin, via a shared 8 amino acid motif with homology to LD motifs in paxillin near the region of the protein that binds tensin; and 3) with caveolin-1 (CAV-1), via the StAR-related lipid transfer (START) domain near the C-terminus of DLC1. Analysis of various DLC1 mutants indicated that the binding sites for each of these interactions contributed to the growth suppressor function of DLC1, but that this binding did not affect in vivo Rho-GAP activity of DLC1. These studies validate the hypothesis that DLC1 is a multifunctional protein whose biological activity depends both on its RhoGAP activity and its ability to bind a variety of signaling molecules. We have begun to characterize how post-translational modifications of DLC1, especially phosphorylations, may affect its activity. We first identified CDK5, which is a cytoplasmic kinase whose physiologic role promotes differentiation, as a major activator of DLC1. This activation occurs because DLC1 has 4 serines that are phosphorylated by CDK5. When these serines, which are located N-terminal to the Rho-GAP domain of DLC1, are not phosphorylated, the N-terminus binds to the Rho-GAP domain, which places DLC1 in a closed, inactive conformation. When the serines are phosphorylated, it decreases the interaction of the N-terminus with the Rho-GAP domain, which places DLC1 in an open, active conformation. In cancer, CDK5 behaves as a pro-oncogenic factor, presumably because it stimulates more pro-oncogenic targets than anti-oncogenic targets, such as DLC1. Consistent with that hypothesis, down-regulation of DLC1 greatly increases the pro-oncogenic activity of CDK5 in cancer cells. AKT is another kinase that directly phosphorylates DLC1, on 3 serines located N-terminal to the Rho-GAP domain. The effects of AKT antagonize those of CDK5, in that the AKT phosphorylations attenuate the Rho-GAP and tumor suppressor activities of DLC1 by changing DLC1 from an open, active configuration to a closed, inactive configuration. AKT lies downstream from receptor tyrosine kinases (RTKs), and our data indicate that increased Rho-GTP following stimulation of cells with RTK ligands is attributable to the activation of AKT and its attenuation of DLC1. These findings may have therapeutic potential, as AKT inhibitor treatment of tumors that express DLC1 protein and have high AKT activity have much greater antitumor activity than tumors that do not express DLC1. The high anti tumor activity from AKT inhibition is associated with loss of phosphorylation of the DLC1 by AKT and reactivation of the Rho-GAP and tumor suppressor activities of DLC1.
| Ashraf, Shazia; Kudo, Hiroki; Rao, Jia et al. (2018) Mutations in six nephrosis genes delineate a pathogenic pathway amenable to treatment. Nat Commun 9:1960 |
| Tripathi, Brajendra K; Lowy, Douglas R (2017) DLC1: a tumor suppressor that regulates Rho signaling. Oncotarget 8:27674-27675 |
| Lowy, Doug R; Singer, Dinah S (2017) Implementing the Cancer Moonshot and beyond. Lancet Oncol 18:e622-e623 |
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| Lowy, Douglas; Singer, Dinah; DePinho, Ron et al. (2016) Cancer moonshot countdown. Nat Biotechnol 34:596-9 |
| Durkin, Marian E; Qian, Xiaolan; Popescu, Nicholas C et al. (2013) Isolation of Mouse Embryo Fibroblasts. Bio Protoc 3: |
| Qian, Xiaolan; Durkin, Marian E; Wang, Dunrui et al. (2012) Inactivation of the Dlc1 gene cooperates with downregulation of p15INK4b and p16Ink4a, leading to neoplastic transformation and poor prognosis in human cancer. Cancer Res 72:5900-11 |
| Du, Xiaoli; Qian, Xiaolan; Papageorge, Alex et al. (2012) Functional interaction of tumor suppressor DLC1 and caveolin-1 in cancer cells. Cancer Res 72:4405-16 |
| Li, Guorong; Du, Xiaoli; Vass, William C et al. (2011) Full activity of the deleted in liver cancer 1 (DLC1) tumor suppressor depends on an LD-like motif that binds talin and focal adhesion kinase (FAK). Proc Natl Acad Sci U S A 108:17129-34 |
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