SRC and ERK cooperatively phosphorylate DLC1 and attenuate its Rho-GAP and tumor suppressor functions

Tripathi, Brajendra K.; Anderman, Meghan F.; Qian, Xiaolan; Zhou, Ming; Wang, Dunrui; Papageorge, Alex G.; Lowy, Douglas R.

doi:10.1083/jcb.201810098

Cited by 14 publications

(14 citation statements)

References 42 publications

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“…DLC1 variant 2 cDNA was used in this study for the generation of the different expression plasmids. pCDNA DLC1, pEGFP-DLC1, pEGFP DLC1 1–600, pEGFP DLC1 500–1091, pEGFP DLC1 1–110, pEGFP DLC1 1–492, pEGFP DLC1 80–400, pEGFP DLC1 400–500, pEGFP DLC1 500–798, pEGFP DLC1 609–848, pEGFP-DLC2, and pEGFP-DLC3 have already been described [ 5 , 11 , 13 , 17 ]. To clone the DLC1-START domain containing region, the DLC1 sequence corresponding to amino acids 848–1091 was amplified by PCR using the primers listed in Supplementary Table 1 and the resulting PCR product was cloned into the pEGFP-C1 empty vector into EcoRI-SalI sites.…”

Section: Methodsmentioning

confidence: 99%

“…Together, these findings lead to the conclusion that DLC1 exerts some of its tumor suppresive effects via GAP-independent mechanisms. Moreover, the activity of DLC1 can also be affected post-translationally, as several kinases, including PKA, AKT, and SRC, phosphorylate and ultimately affect DLC1 tumor suppressor activities [ 10 , 11 ], and reviewed by [ 7 ].…”

Section: Introductionmentioning

confidence: 99%

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The tumor suppressor activity of DLC1 requires the interaction of its START domain with Phosphatidylserine, PLCD1, and Caveolin-1

et al. 2021

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Background DLC1, a tumor suppressor gene that is downregulated in many cancer types by genetic and nongenetic mechanisms, encodes a protein whose RhoGAP and scaffolding activities contribute to its tumor suppressor functions. The role of the DLC1 START (StAR-related lipid transfer; DLC1-START) domain, other than its binding to Caveolin-1, is poorly understood. In other START domains, a key function is that they bind lipids, but the putative lipid ligand for DLC1-START is unknown. Methods Lipid overlay assays and Phosphatidylserine (PS)-pull down assays confirmed the binding of DLC1-START to PS. Co-immunoprecipitation studies demonstrated the interaction between DLC1-START and Phospholipase C delta 1 (PLCD1) or Caveolin-1, and the contribution of PS to those interactions. Rho-GTP, cell proliferation, cell migration, and/or anchorage-independent growth assays were used to investigate the contribution of PS and PLCD1, or the implications of TCGA cancer-associated DLC1-START mutants, to DLC1 functions. Co-immunoprecipitations and PS-pull down assays were used to investigate the molecular mechanisms underlying the impaired functions of DLC1-START mutants. A structural model of DLC1-START was also built to better understand the structural implications of the cancer-associated mutations in DLC1-START. Results We identified PS as the lipid ligand for DLC1-START and determined that DLC1-START also binds PLCD1 protein in addition to Caveolin-1. PS binding contributes to the interaction of DLC1 with Caveolin-1 and with PLCD1. The importance of these activities for tumorigenesis is supported by our analysis of 7 cancer-associated DLC1-START mutants, each of which has reduced tumor suppressor function but retains wildtype RhoGAP activity. Our structural model of DLC1-START indicates the mutants perturb different elements within the structure, which is correlated with our experimental findings that the mutants are heterogenous with regard to the deficiency of their binding properties. Some have reduced PS binding, others reduced PLCD1 and Caveolin-1 binding, and others are deficient for all of these properties. Conclusion These observations highlight the importance of DLC1-START for the tumor suppressor function of DLC1 that is RhoGAP-independent. They also expand the versatility of START domains, as DLC1-START is the first found to bind PS, which promotes the binding to other proteins.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

The tumor suppressor activity of DLC1 requires the interaction of its START domain with Phosphatidylserine, PLCD1, and Caveolin-1

et al. 2021

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“…Moreover, DLC1 itself can be phosphorylated by a number of different kinases, including Src, ERK1/2, CDK5, and Akt, which may lead to changes in its Rho-GAP activity (45)(46)(47). By the same token, it is unlikely that VAV2 is the only GEF involved in regulating epithelial cell migration in the late phase.…”

Section: Discussionmentioning

confidence: 99%

Dynamic interplay of two molecular switches enabled by the MEK1/2–ERK1/2 and IL-6–STAT3 signaling axes controls epithelial cell migration in response to growth factors

Qin

Kaneko

Voss

et al. 2021

Journal of Biological Chemistry

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This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

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“…Activation of ERK promotes phosphorylation of the Rho-GAP DLC1 in Ser129, facilitating the binding of SRC to DLC1 and promoting further phosphorylation of DLC in Tyr451 and Tyr701. Thus, SRC and ERK1/2 signaling converge on DLC1 to cooperatively regulate it, attenuating the Rho-GAP and tensin-binding activities of DLC1 [124]. A more complex regulation of Rho-GEF and GEF-H1 (ARHGEF2) by ERK of 30 has been suggested, in which, by phosphorylation of different residues of GEF-H1, ERK controls RhoA activity.…”

Section: Ras-rho Crosstalk Through Raf/mek/erk Activationmentioning

confidence: 99%

The Crossroads between RAS and RHO Signaling Pathways in Cellular Transformation, Motility and Contraction

Soriano

Alcón-Pérez

Vicente‐Manzanares

et al. 2021

Genes

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Ras and Rho proteins are GTP-regulated molecular switches that control multiple signaling pathways in eukaryotic cells. Ras was among the first identified oncogenes, and it appears mutated in many forms of human cancer. It mainly promotes proliferation and survival through the MAPK pathway and the PI3K/AKT pathways, respectively. However, the myriad proteins close to the plasma membrane that activate or inhibit Ras make it a major regulator of many apparently unrelated pathways. On the other hand, Rho is weakly oncogenic by itself, but it critically regulates microfilament dynamics; that is, actin polymerization, disassembly and contraction. Polymerization is driven mainly by the Arp2/3 complex and formins, whereas contraction depends on myosin mini-filament assembly and activity. These two pathways intersect at numerous points: from Ras-dependent triggering of Rho activators, some of which act through PI3K, to mechanical feedback driven by actomyosin action. Here, we describe the main points of connection between the Ras and Rho pathways as they coordinately drive oncogenic transformation. We emphasize the biochemical crosstalk that drives actomyosin contraction driven by Ras in a Rho-dependent manner. We also describe possible routes of mechanical feedback through which myosin II activation may control Ras/Rho activation.

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SRC and ERK cooperatively phosphorylate DLC1 and attenuate its Rho-GAP and tumor suppressor functions

Cited by 14 publications

References 42 publications

The tumor suppressor activity of DLC1 requires the interaction of its START domain with Phosphatidylserine, PLCD1, and Caveolin-1

The tumor suppressor activity of DLC1 requires the interaction of its START domain with Phosphatidylserine, PLCD1, and Caveolin-1

Dynamic interplay of two molecular switches enabled by the MEK1/2–ERK1/2 and IL-6–STAT3 signaling axes controls epithelial cell migration in response to growth factors

The Crossroads between RAS and RHO Signaling Pathways in Cellular Transformation, Motility and Contraction

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