Doctoral Dissertations

Date of Award

5-2009

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Life Sciences

Major Professor

Albrecht von Arnim

Abstract

The objective of this study was to dissect the function of the h subunit of translation initiation factor 3, eIF3h, in Arabidopsis shoot apical meristem (SAM) maintenance and auxin response. Translational regulation has been proposed to play important roles in plant development and environmental stress responses. eIF3, with 12-13 protein subunits the largest of the initiation factor complexes, is involved in many essential steps in translation initiation, but little is known about the structural and functional roles of most eIF3 subunits in any organism. Using various techniques, such as scanning electron microscopy and confocal microscopy, I revealed defects in the eif3h mutant that included an enlarged SAM and a pin-formed shoot and a variety of defects in leaf initiation and morphogenesis.Many groups of genes, such as those encoding auxin response factors (ARFs) and stem cell regulators (CLAVATA1 and 3) were identified to have multiple upstream open reading frames (uORFs), which are generally inhibitory for the translation of downstream open reading frames and involved in translational control. Optimized translation assays indicated that ARFs and CLAVATA (CLV1, 3) genes were significantly less translated in the eif3h mutant, and removal of uORFs from CLV1 and CLV3 leaders significantly reduced their eIF3h dependence. Furthermore, the eif3h growth defects can be partially complemented by expressing the CLAVATA3 gene in its native domain. These results indicated that, eIF3h is involved in efficient translation of ARFs and CLV1, 3 genes by overcoming the translation repression by uORFs. The functional characterization of eIF3h provided the first evidence that translational control by eIF3h plays important roles in auxin signaling and shoot apical meristem maintenance.As part of this project I explored the utility of a viral sequence element described as an internal ribosome entry site. Even though no robust IRES activity could be confirmed under the experimental conditions used here, the sequence had substantial promoter activity. Because the promoter activity was de factor eIF3h independent, the sequence could be utilized a component of a new generation of translational reporter genes.

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