Inflammation js the body's natural protective response to infection or injury, but abnormal or uncontrolled inflammatory responses can do more harm than good. A breakdown in the appropriate regulation of inflammation underlies a wide range of common diseases, such as rheumatoid arthritis, inflammatory bowel disease, multiple sclerosis, and septic shock. The expression of pro-inflammatory genes by cells in the immune system is the most crucial step in the development of inflammation. Although significant progress has been made, the mechanism of pro-inflammatory gene expression is still not fully understood. The long- term objective of this proposal is to gain an understanding of the regulation and function of microRNA (miRNA), a newly identified group of short non-coding single-stranded RNA, in the expression of pro- inflammatory genes. It is knownthat pro-inflammatory genes such as tumor necrosis factor, interleukin-1, arid cyclooxygenase 2 are controlled at multiple levels of gene expression, one of the most important of which is at the regulation of then- mRNAstability. Quick mRNAdegradation of these pro-inflammatorygenes serves as a mechanism to control the level of pro-inflammatory molecules. AU-rich elements (AREs) located in the 3' untranslated region of these short-lived mRNAs dictate their degradation. Ourrecent study revealed that Dicer and Argonaute (Ago/eif2C) family members, components involved in microRNA(miRNA) processing and function, are required for the rapid decay of mRNA that contain AREs (ARE-mRNA). Furthermore, we found miR16, a human miRNAwith a sequence that is partially complementaryto the ARE sequence, to be required for ARE-mRNAturnover. The requirement of miR16 in ARE-mRNA decay in resting cells suggests a physiological 'housekeeping' function of miR16 in which it prevents high basal levels of pro-inflammatory gene expression. This proposal outlines a study into the mechanism of miRNA-regulated cytokine gene expression, with an emphasis on how inflammatory stimuli induce stabilization of ARE-mRNA by blocking miRNA-mediated mRNA degradation. In this research plan, we will also evaluate the overall function of miR16 by genome-wide identifying and classifying of miR16-targeted genes. To achieve our aims, we will utilize a combination of genetic, biochemical, and molecular biological approaches. The work proposed in this application will contribute to a better understanding of pro-inflammatory gene expression. Understanding the function of miRNA in mRNA stability of pro-inflammatory genes should provide new avenues for developing novel anti-inflammatoryagents.
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