Date of Award
Spring 2015
Document Type
Thesis
Degree Name
Master of Science (MS)
Department
Biological Sciences
First Advisor
Anderson, James T.
Second Advisor
Bozdag, Serdar
Third Advisor
Abbott, Allison
Abstract
RNA surveillance and degradation play an important role in the development and growth of organisms by eliminating RNA that contains errors, or that is no longer needed by the cell. In some processes, RNAs designated to be degraded are first labeled and then specifically recognized by the exosome, which performs the final degradation. One of the key labeling factors in yeast is the TRAMP complex, a three-subunit complex composed of Air2, Trf4 and Mtr4. Air2 facilitates TRAMP binding of RNA, Trf4 appends a 3′ end polyA tail and Mtr4 regulates the rate of adenylation and modifies RNA structures for ease of degradation through its RNA helicase activity. Though TRAMP has been studied extensively in yeast and its biochemistry and RNA recognition functions well delineated, the recent identification of TRAMP in mammals has made it possible for work to characterize mammalian TRAMP function in tissue culture cells. The mammalian transcriptome is much more complex and diverse compared to yeast in that a large portion is consisted of non-coding RNAs such as lncRNA, snoRNA, miRNA and so on. To understand the role of TRAMP complex in gene expression regulation, we knockdown the SKIV2L2 (mouse Mtr4) subunit and performed a polyA sequencing. With bioinformatics tools such as Bowtie, F-Seqq, MEDIPS, miRCompare, we constructed a data pipeline and identified several categories of targets including snoRNA, rRNA, miRNA and long-noncoding RNA in mouse cells. These data suggests that the targets of TRAMP are widely spread along the genome, and these targets involve a myriad of regulatory pathways. Understanding the relationship between the targets will help reveal the function and effect of this complex. Also, a more accurate and comprehensive target identification method remains to be developed.