Various RNAs transcribed from genes implicated in cardiovascular and pulmonary diseases are regulated at the post- transcriptional level. These include the beta-adrenergic receptor, the beta2- adrenergic receptor, the vascular endothelial growth factor (VEGF) and interleukin-3 (IL-3) mRNAs. These transcripts harbor Adenosine/Uridine-rich elements (AREs) in their 3' untranslated regions (3'-UTR) which play a role in regulating their stability and translation. AREs target mRNAs for rapid decay, usually via a poly (A) shortening-dependent decay pathway. The mRNA destabilizing, poly(A) shortening and translational functions of the AREs are mediated by ARE-binding proteins (ARE-BPs). Interestingly, inappropriate control of the abundance of these transcripts leads to disease. Pharmacological manipulation of these mRNAs requires an understanding of the molecular mechanisms regulating their gene expression. The greatest difficulty, however, in studying how the AREs interact with ARE-BPs to regulate gene expression. The greatest difficulty, however, in studying how the AREs interact with ARE-BPs to regulate gene expression is to be able to demonstrate that the identified ARE-BP is actually required for post- transcriptional regulation of a particular transcript. We have develop a system to investigate the ARE-mediated mRNA decay pathway in yeast. Insertion of the Tumor Necrosis Factor alpha(TNFalpha) ARE into the 3'UTR of the yeast MFA2 mRNA, causes rapid degradation of the chimeric mRNA. Expression of AUF1, an ARE-BP, in the yeast S.cerevisiae specifically affects the decay rate of this ARE-containing mRNA suggesting that the yeast system recapitulates the results obtained in mammalian cells. The goals of the experiments in this grant proposal are to utilize the genetic and molecular aspects of yeast to investigate the ARE-mediated mRNA decay pathway in yeast and mammalian cells. Specifically, we propose to: a) investigate the mechanism of how the beta- adrenergic receptors, IL-3 and VEGF AREs promote mRNA turnover using the yeast S.cerevisiae as a model system; b) investigate how these AREs regulate the stability of their mRNAs in mammalian cells and in cell-free system; and c) develop assays to characterize the effects of the beta-adrenergic receptors, IL-3 and VEGF AREs on translation. These studies will help us expand our knowledge on AREs and ARE-BPs, and how these interactions control the expression of mRNAs involved in cardiovascular and pulmonary diseases, as well as in cancers and immune disorders.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Scientist Development Award - Research & Training (K01)
Project #
5K01HL004355-04
Application #
6555852
Study Section
Special Emphasis Panel (ZHL1-CSR-C (F1))
Program Officer
Commarato, Michael
Project Start
2000-05-01
Project End
2005-04-30
Budget Start
2002-05-01
Budget End
2003-04-30
Support Year
4
Fiscal Year
2002
Total Cost
$112,547
Indirect Cost
Name
University of Puerto Rico Rio Piedras
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
City
San Juan
State
PR
Country
United States
Zip Code
00931
Cunci, Lisandro; Vargas, Marina Martinez; Cunci, Roman et al. (2014) Real-Time Detection of Telomerase Activity in Cancer Cells using a Label-Free Electrochemical Impedimetric Biosensing Microchip. RSC Adv 4:52357-52365
González-Feliciano, José A; Hernández-Pérez, Marimar; Estrella, Luis A et al. (2014) The role of HuR in the post-transcriptional regulation of interleukin-3 in T cells. PLoS One 9:e92457
Lasalde, Clarivel; Rivera, Andrea V; León, Alfredo J et al. (2014) Identification and functional analysis of novel phosphorylation sites in the RNA surveillance protein Upf1. Nucleic Acids Res 42:1916-29
Estrella, Luis A; Wilkinson, Miles F; González, Carlos I (2009) The shuttling protein Npl3 promotes translation termination accuracy in Saccharomyces cerevisiae. J Mol Biol 394:410-22
Wang, Weirong; Cajigas, Ivan J; Peltz, Stuart W et al. (2006) Role for Upf2p phosphorylation in Saccharomyces cerevisiae nonsense-mediated mRNA decay. Mol Cell Biol 26:3390-400
Windgassen, Merle; Sturm, Dorothee; Cajigas, Ivan J et al. (2004) Yeast shuttling SR proteins Npl3p, Gbp2p, and Hrb1p are part of the translating mRNPs, and Npl3p can function as a translational repressor. Mol Cell Biol 24:10479-91
Gonzalez, C I; Bhattacharya, A; Wang, W et al. (2001) Nonsense-mediated mRNA decay in Saccharomyces cerevisiae. Gene 274:15-25
Gonzalez, C I; Wang, W; Peltz, S W (2001) Nonsense-mediated mRNA decay in Saccharomyces cerevisiae: a quality control mechanism that degrades transcripts harboring premature termination codons. Cold Spring Harb Symp Quant Biol 66:321-8