Proteomic Analysis of Plasma Membrane from Hypoxia-Adapted Malignant Melanoma

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• 作者：Luke H. Stockwin ; Josip Blonder ; Maja A. Bumke ; David A. Lucas ; King C. Chan ; Thomas P. Conrads ; Haleem J. Issaq ; Timothy D. Veenstra ; Dianne L. Newton ; Susanna M. Rybak
• 刊名：Journal of Proteome Research
• 出版年：2006
• 出版时间：November 2006
• 年：2006
• 卷：5
• 期：11
• 页码：2996 - 3007
• 全文大小：591K
• 年卷期：v.5,no.11(November 2006)
• ISSN：1535-3907

文摘

Hypoxic conditions often persist within poorly vascularized tumors. At the cellular level constitutiveactivation of transcriptional regulators of the hypoxic response leads to the emergence of clones withaggressive phenotypes. The primary interface between the cell and the hypoxic environment is theplasma membrane. A detailed investigation of this organelle is expected to yield further targets fortherapeutic perturbation of the response to hypoxia. In the present study, quantitative proteomic analysisof plasma membrane from hypoxia-adapted murine B16F10 melanoma was performed using differential¹⁶O/¹⁸O stable isotopic labeling and multidimensional liquid chromatography-tandem mass spectrometry. The analysis resulted in the identification of 24,853 tryptic peptides, providing quantitativeinformation for 2,433 proteins. For a subset of plasma membrane and secreted proteins, quantitativeRT-PCR was used to gain further insight into the genomic regulatory events underlying the responseto hypoxia. Consistent increases at the proteomic and transcriptomic levels were observed foraminopeptidase N (CD13), carbonic anhydrase IX, potassium-transporting ATPase, matrix metalloproteinase 9, and stromal cell derived factor I (SDF-1). Antibody-based analysis of a panel of humanmelanoma cell lines confirmed that CD13 and SDF-1 were consistently upregulated during hypoxia.This study provides the basis for the discovery of novel hypoxia-induced membrane proteins.