Exploration of the cis -elements and trans -factors involved in post-transcriptional mechanisms of the gene atpI in chloroplast gene expression
Smithers, Emily Rose
MetadataShow full item record
Chloroplast gene expression is strongly influenced by posttranscriptional mechanisms, many involving protein-RNA complexes. The focus of my research is the identification and study of RNA-protein complexes that form in the 5’ untranslated regions (UTRs) of chloroplast-encoded mRNAs. To analyze cis -elements in vivo, variants of the atpI 5’UTR were placed upstream of a reporter gene and transformed into the tobacco chloroplast genome. I showed that the ribosome binding site is necessary, but not sufficient, for efficient translation in vivo (Chapter 2). Two conserved regions in the atpI 5’UTR, named Con1 and Con2, have positive roles in translation efficiency in vivo. In addition, one conserved region, Con1, assists in stabilizing translation efficiency in dark-adapted plants (Chapter 3). Through in vitro analyses with a fractionated chloroplast extract, I determined that at least two of the five tobacco chloroplast ribonucleoproteins (cpRNPs), a class of nuclear-encoded chloroplast RNA binding proteins, are part of the RNA-protein binding complexes that form with chloroplast extracts. Further investigation of the binding characteristics of the cpRNPs revealed their ability to preferentially bind to a chloroplast RNA over a non-chloroplast RNA (Chapter 4). This work is the first example of cpRNPs displaying specificity for a particular nucleic acid. Impact and future directions of this research are discussed.