Lysyl oxidase (LO), a copper-(Cu) dependent enzyme, oxidizes peptidyl lysine residues in substrates, e.g., collagen, elastin and histone H1, essential for organization and stabilization of the extracellular matrix (ECM) and the cell nucleus. This enzyme has been identified as a tumor suppressor, for example, inhibiting transforming activity of ras, a proto oncogene. Thus, LO as an intra- and extracellular effector plays a critical role in human physiology and pathology. Chronic exposure of humans to cadmium (Cd), a heavy metal, either from occupational contamination or from cigarette smoke, induces emphysema and lung cancers. However, the mechanisms for Cd-elicited lung pathology remain poorly understood. LO as a metalloenzyme is susceptible to changes in cellular metal homeostasis. Previous studies by this lab investigating the phenotype change from Cd sensitive to Cd resistant of rat lung fibroblasts (RFL6) illustrated downregulation of LO by Cd at mRNA, protein and catalytic levels. Continuing studies further indicated that RFL6 cells in response to Cd displayed inhibition of LO transcription initiation and enhancement of LO mRNA decay both collectively contributing to decreased levels of steady-state LO mRNAs. These findings have led to a hypothesis that transcriptional control and regulation of the LO gene are critical targets for Cd insult and silencing of LO gene transcription by Cd is a key molecular basis for lung diseases. The overall goal of the proposed research is to test this hypothesis by achieving following specific aims: 1) to identify mechanisms for Cd silencing of the LO gene at the transcriptional level by examining Cd modulation of RNA polymerase II- directed LO pre-mRNA synthesis and processing, and of the LO promoter activation regulated by the core promoter;2) to identify mechanisms for Cd silencing of the LO gene at the transcriptional level by examining Cd modulation of the LO promoter activation regulated by metal and redox-sensitive transcription factors and their cognate cis-elements, and determining LO promoter methylation to provide evidence for Cd epigenetic damage to LO DNA;3) to identify mechanisms for Cd silencing of the LO gene at the posttranscriptional level by examining Cd effects on the 5'-capping and 3'-polyadenylation status of LO mRNA, and assessing Cd sensitive, LO mRNA stability-related cis-elements in the 3'-untranslation region and their corresponding binding proteins;and 4) to investigate biological consequences of Cd silencing of the LO gene in cell and animal models by examining effects of altered LO expression by Cd on substrate promoter activation and cell transformation in the cell model and assessing the active status of the major LO transcriptional and posttranscriptional machineries as well as aberrant methylation of the LO gene promoter in emphysematous and carcinogenic lungs of rats receiving Cd by chronic administration. The outcomes of the proposed research are expected to enhance our understanding of mechanisms of LO gene silence by Cd providing the basis for developing prophylaxis and treatment strategies for Cd-related lung diseases.

Public Health Relevance

Lysyl oxidase (LO) is a key copper (Cu)-dependent enzyme existing in the extracellular matrix (ECM) and the cell nucleus critical for organ morphogenesis, tissue repair and anti- tumorigenesis of the lung. Previous and preliminary studies by this lab have indicated that cadmium (Cd), a toxic and carcinogenic heavy metal, induced LO gene silencing in rat lung fibroblasts. The proposed research extends previous findings and aims at investigating molecular mechanisms for LO gene silence by Cd in cell and animal models providing the basis for developing protective and therapeutic strategies for Cd-related lung diseases such as emphysema and cancers.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES011340-09
Application #
8249076
Study Section
Lung Cellular, Molecular, and Immunobiology Study Section (LCMI)
Program Officer
Nadadur, Srikanth
Project Start
2001-12-01
Project End
2014-03-31
Budget Start
2012-04-01
Budget End
2013-03-31
Support Year
9
Fiscal Year
2012
Total Cost
$358,350
Indirect Cost
$137,827
Name
Boston University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
604483045
City
Boston
State
MA
Country
United States
Zip Code
02118
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Zhao, Yinzhi; Toselli, Paul; Li, Wande (2012) Microtubules as a critical target for arsenic toxicity in lung cells in vitro and in vivo. Int J Environ Res Public Health 9:474-95
Li, Wande; Zhou, Jing; Chen, Lijun et al. (2011) Lysyl oxidase, a critical intra- and extra-cellular target in the lung for cigarette smoke pathogenesis. Int J Environ Res Public Health 8:161-84
Zhao, Yinzhi; Chen, Lijun; Gao, Song et al. (2010) The critical role of the cellular thiol homeostasis in cadmium perturbation of the lung extracellular matrix. Toxicology 267:60-9
Chou, Denise K; Zhao, Yinzhi; Gao, Song et al. (2007) Perturbation of copper (Cu) homeostasis and expression of Cu-binding proteins in cadmium-resistant lung fibroblasts. Toxicol Sci 99:267-76
Gao, Song; Zhao, Yinzhi; Kong, Lingfa et al. (2007) Cloning and characterization of the rat lysyl oxidase gene promoter: identification of core promoter elements and functional nuclear factor I-binding sites. J Biol Chem 282:25322-37
Zhao, Yinzhi; Gao, Song; Chou, Iih-Nan et al. (2006) Inhibition of the expression of lysyl oxidase and its substrates in cadmium-resistant rat fetal lung fibroblasts. Toxicol Sci 90:478-89
Gao, Song; Chen, Keyang; Zhao, Yinzhi et al. (2005) Transcriptional and posttranscriptional inhibition of lysyl oxidase expression by cigarette smoke condensate in cultured rat fetal lung fibroblasts. Toxicol Sci 87:197-203
Chen, Li-Jun; Zhao, Yinzhi; Gao, Song et al. (2005) Downregulation of lysyl oxidase and upregulation of cellular thiols in rat fetal lung fibroblasts treated with cigarette smoke condensate. Toxicol Sci 83:372-9