Generation of novel monoclonal antibodies and their application for detecting ARD1 expression in colorectal cancer

Department of Biochemistry and Molecular Biology, Peking University School of Oncology, Beijing Cancer Hospital & Institute, HaiDian District, Beijing, China.
Cancer Letters (Impact Factor: 5.62). 07/2008; 264(1):83-92. DOI: 10.1016/j.canlet.2008.01.028
Source: PubMed


Arrest defective 1 (ARD1) is an acetyltransferase involved in cell cycle control in yeast. ARD1 interacts with human N-acetyltransferase (NATH) to form a functional N-terminal acetyltransferase complex. Recently it had been linked with proliferation and apoptosis in mammalian cells, but its function in cancer development remains unclear. To evaluate significance of ARD1 expression in human colorectal cancer, we generated a panel of monoclonal antibodies (mAbs) with high specificity and sensitivity against ARD1. All of the 10 different clones could be used in ELISA and Western blot, and clone 10C12, 13G2, and 4D10 can interact with ARD1 in eukaryotic cells by immunoprecipitation (IP). Clones of 14D4 and 10C12 were strongly reacted to ARD1 in immunocytochemistry (ICH) and immunohistochemistry (IHC). ARD1 expression was evaluated in human colorectal cancer and colitis tissues by immunohistochemical analysis with mAb 14D4. Forty-one were ARD1-positive in 50 colorectal cancer tissues and only 12 were weak positive in the 50 matched normal tissues (P < 0.001). Moreover, ARD1 expression was not detectable in 20 cases of colitis tissue (P < 0.001). Furthermore, all of the six human colorectal cancer cell lines we examined were also ARD1-positive at mRNA and protein levels. Taken together, the novel mAbs against ARD1 we generated could be good tools for both basic and clinical studies, and ARD1 could be a potential biomarker in colorectal cancer.

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    • "Whether or not there is any functional association between these pathways and PA28-regulated chymotrypsin-like proteasome activity still needs more considerations. It was reported that overexpression of Naa10p may serve as a prognostic factor in different cancers [7] [10] [15] [16] [33]. Overexpression of PA28b was found in gastric adenocarcinomas, though its prognostic value still needs exploration [34] [35]. "
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    ABSTRACT: N-α-acetyltransferase 10 protein (Naa10p) regulates various pathways associated with cancer cell proliferation, metastasis, apoptosis and autophagy. However, its role in protein quality control is unknown. Here, we report that Naa10p is physically associated with proteasome activator 28β (PA28β). Naa10p also interacts with PA28α in a PA28β-dependent manner. Naa10p negatively regulates PA28-dependent chymotrypsin-like proteasome activity in cancer cells and in a cell-free system reconstituted with purified proteins, which was not related to 26S proteasome. Acetyltransferase activity of Naa10p isn't required for its effect on chymotrypsin-like proteasome activity. Therefore, our data reveal that Naa10p suppresses 28S proteasome activity through interaction with PA28β. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: Naa10pphysically interactswithPS28alphaandPS28 betabyanti bait coimmunoprecipitation(View Interaction:1,2).
    FEBS letters 04/2013; 587(11). DOI:10.1016/j.febslet.2013.04.016 · 3.17 Impact Factor
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    • "By using immunohistochemical staining, Arnesen et al. found that the level of ARD1 protein is downregulated in most thyroid neoplasm specimens compared to that in non-neoplastic tissues [50]. However, two research groups showed conflicting results with a higher expression of ARD1 in tumor tissues in colorectal and other types of cancer [26] [51]. This discrepancy may have come from the histological specificity of ARD1 in different tissues, making it complicated to interpret the functional consequences of ARD1 expression in mammalian cells. "
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    ABSTRACT: Arrest-defect-1 protein (ARD1), an acetyltransferase, catalyzes N-alpha-acetylation in yeast. In mammalian cells, both N-alpha-acetylation and epsilon-acetylation induced by ARD1 have been reported. Emerging evidence has revealed that ARD1 is involved in a variety of cellular functions, including proliferation, apoptosis, autophagy, and differentiation and that dysregulation of ARD1 is associated with tumorigenesis and neurodegenerative disorder. This review will discuss recent discoveries regarding variations among the different ARD1 isoforms, the associated biological functions of ARD1, and ARD1 localization in different cells. We will also discuss the potential upstream regulators and downstream targets of ARD1 to provide new avenues for resolving its controversial roles in cancer development.
    American Journal of Translational Research 01/2010; 2(1):56-64. · 3.40 Impact Factor
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    • "In the same study hNAA10 was identified among genes associated with dedifferentiation of hepatocellular carcinoma [42]. By immunohistochemistry, hNaa10p was shown to be overexpressed in colorectal cancers as compared to adjacent normal tissue [43]. An other study demonstrated that hNaa10p was overexpressed in colorectal cancers as well as in breast cancers [44]. "
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    ABSTRACT: Protein Nalpha-terminal acetylation is one of the most common protein modifications in eukaryotic cells, occurring on approximately 80% of soluble human proteins. An increasing number of studies links Nalpha-terminal acetylation to cell differentiation, cell cycle, cell survival, and cancer. Thus, Nalpha-terminal acetylation is an essential modification for normal cell function in humans. Still, little is known about the functional role of Nalpha-terminal acetylation. Recently, the three major human N-acetyltransferase complexes, hNatA, hNatB and hNatC, were identified and characterized. We here summarize the identified N-terminal acetyltransferase complexes in humans, and we review the biological studies on Nalpha-terminal acetylation in humans and other higher eukaryotes.
    BMC proceedings 02/2009; 3 Suppl 6(Suppl 6):S3. DOI:10.1186/1753-6561-3-S6-S3
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