Please use this identifier to cite or link to this item: http://scholarbank.nus.edu.sg/handle/10635/22943
Title: Study of Interaction between deleted in Liver Cancer 1 and its Novel interacting partner adenine nucleotide translocase 2
Authors: WONG MING ZHI DENISE
Keywords: deleted in liver cancer 1, adenine nucleotide translocase 2, proteomics pull down, mitochondrial membrane potential
Issue Date: 21-Jul-2010
Source: WONG MING ZHI DENISE (2010-07-21). Study of Interaction between deleted in Liver Cancer 1 and its Novel interacting partner adenine nucleotide translocase 2. ScholarBank@NUS Repository.
Abstract: Deleted in liver cancer 1 (DLC1) is a ubiquitously expressed Rho GTPase-activating protein (RhoGAP) that has been shown to enhance the intrinsic GTPase activity of RhoA and to a much lesser extent, Cdc42. Through mechanisms such as loss of heterozygosity and promoter hypermethlyation, the inactivation of DLC1 and subsequent enhancement of RhoA activity has contributed to rapid tumor expansion and metastasis. The re-introduction of DLC1 into various cancer cell lines has been shown to result in drastic cytoskeletal remodeling and inhibition of tumor cell growth via inhibiting cell proliferation, inducing apoptosis and reducing cell migration. This tumor suppressor activity of DLC1 has been attributed to the ability of the GAP domain to negatively regulate Rho GTPases as well as through other GAP-independent mechanisms. The multi-modular architecture of DLC1 suggest that other domains such as the sterile alpha motif (SAM) and STAR-related lipid transfer (START) domains may be important for interaction with proteins other than Rho that may contribute to its function. To address this possibility, pulldown experiments using DLC1 as a bait protein was conducted to identify potential interacting partners in 293T cells. Adenine nucleotide translocase 2 (ANT2), a resident of the inner mitochondrial membrane, was identified by mass spectrometry and validated by co-immunoprecipitation studies to associate with DLC1. The regions on both DLC1 and ANT2 that are required for their interaction have been mapped to unique sites not known to bind other known DLC1 interacting partners. While the co-expression of DLC1 with ANT2 did not reduce levels of apoptotic markers pro-caspase 3 or 9 in 293T cells, confocal microscopic studies in HeLa cells indicate that wild-type or a GAP-inactive DLC1 mutant could reduce mitochondria membrane potential via interaction with ANT2, as measured by a reduction in the fluorescence of MitoTracker Red. This study has established a GAP-independent function of DLC1 to bring about a reduction in mitochondrial membrane potential through its interaction with ANT2. The significance of the findings presented here is also discussed.
URI: http://scholarbank.nus.edu.sg/handle/10635/22943
Appears in Collections:Master's Theses (Open)

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