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Natriuretic peptide pharmacogenetics: membrane metallo-endopeptidase (MME): common gene sequence variation, functional characterization and degradation.

ABSTRACT: Membrane metallo-endopeptidase (MME), also known as neutral endopeptidase 24.11 (EC 3.4.24.11), is involved in the metabolism of natriuretic peptides that play a key role in modulating cardiac structure and function. Common genetic variation in MME has not been addressed by resequencing the gene using DNA from different ethnic populations. We set out to identify and functionally characterize common genetic variation in MME in three ethnic groups. DNA samples from 96 European-American, 96 African-American, and 96 Han Chinese-American healthy subjects were used to resequence MME. Ninety polymorphisms, 65 novel, were identified, including 8 nonsynonymous single nucleotide polymorphisms (nsSNPs). Expression constructs for the nsSNPs were created and COS-1 cells were transfected with constructs for wild type (WT) and variant allozymes. Recombinant proteins were analyzed by quantitative Western blot analysis and by a one-step fluorometric assay. A significant reduction in enzyme activity (21% of WT) and immunoreactive protein (29% of WT) for the Val73 variant allozyme was observed. Proteasome-mediated degradation and autophagy participated in the degradation of this variant allozyme. The chaperone proteins, BiP and GRP94, were upregulated after transfection with Val73 MME, suggesting protein misfolding, compatible with conclusions based on the MME X-ray crystal structure. Multiple novel polymorphisms of MME were identified in three ethnic groups. The Val73 variant allozyme displayed a significant decrease in MME protein quantity and activity, with degradation mediated by both proteasome and autophagy pathways. This polymorphism could have a significant effect on the metabolism of natriuretic peptides.

SUBMITTER: Pereira NL

PROVIDER: S-EPMC3092155 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature

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Natriuretic peptide pharmacogenetics: membrane metallo-endopeptidase (MME): common gene sequence variation, functional characterization and degradation.

Pereira Naveen L NL Aksoy Pinar P Moon Irene I Peng Yi Y Redfield Margaret M MM Burnett John C JC Wieben Eric D ED Yee Vivien C VC Weinshilboum Richard M RM

Journal of molecular and cellular cardiology 20100806 5

Membrane metallo-endopeptidase (MME), also known as neutral endopeptidase 24.11 (EC 3.4.24.11), is involved in the metabolism of natriuretic peptides that play a key role in modulating cardiac structure and function. Common genetic variation in MME has not been addressed by resequencing the gene using DNA from different ethnic populations. We set out to identify and functionally characterize common genetic variation in MME in three ethnic groups. DNA samples from 96 European-American, 96 African ...[more]

PMID: 20692264

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Project description:BACKGROUND: Hypoxia-induced genes are potential targets in cancer therapy. Responses to hypoxia have been extensively studied in vitro, however, they may differ in vivo due to the specific tumor microenvironment. In this study gene expression profiles were obtained from fresh human lung cancer tissue fragments cultured ex vivo under different oxygen concentrations in order to study responses to hypoxia in a model that mimics human lung cancer in vivo. METHODS: Non-small cell lung cancer (NSCLC) fragments from altogether 70 patients were maintained ex vivo in normoxia or hypoxia in short-term culture. Viability, apoptosis rates and tissue hypoxia were assessed. Gene expression profiles were studied using Affymetrix GeneChip 1.0 ST microarrays. RESULTS: Apoptosis rates were comparable in normoxia and hypoxia despite different oxygenation levels, suggesting adaptation of tumor cells to hypoxia. Gene expression profiles in hypoxic compared to normoxic fragments largely overlapped with published hypoxia-signatures. While most of these genes were up-regulated by hypoxia also in NSCLC cell lines, membrane metallo-endopeptidase (MME, neprilysin, CD10) expression was not increased in hypoxia in NSCLC cell lines, but in carcinoma-associated fibroblasts isolated from non-small cell lung cancers. High MME expression was significantly associated with poor overall survival in 342 NSCLC patients in a meta-analysis of published microarray datasets. CONCLUSIONS: The novel ex vivo model allowed for the first time to analyze hypoxia-regulated gene expression in preserved human lung cancer tissue. Gene expression profiles in human hypoxic lung cancer tissue overlapped with hypoxia-signatures from cancer cell lines, however, the elastase MME was identified as a novel hypoxia-induced gene in lung cancer. Due to the lack of hypoxia effects on MME expression in NSCLC cell lines in contrast to carcinoma-associated fibroblasts, a direct up-regulation of stroma fibroblast MME expression under hypoxia might contribute to enhanced aggressiveness of hypoxic cancers.

Dataset Information

Natriuretic peptide pharmacogenetics: membrane metallo-endopeptidase (MME): common gene sequence variation, functional characterization and degradation.

Publications

Natriuretic peptide pharmacogenetics: membrane metallo-endopeptidase (MME): common gene sequence variation, functional characterization and degradation.

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