Lin Shao

Date of Award

Spring 2013

Project Type

Dissertation

Program or Major

Genetics

Degree Name

Doctor of Philosophy

First Advisor

Subhash C Minocha

Abstract

Polyamines are ubiquitously distributed cationic compounds, which play important roles in numerous cellular functions in plants. This study was aimed at elaborating the regulation of polyamine biosynthetic gene expression and polyamine metabolism. The organ/tissue specific expression patterns of two genes encoding the polyamine biosynthetic enzyme spermidine synthase ( AtSPDS1 and AtSPDS2) were studied in Arabidopsis at different developmental stages using promoter::reporter approach. The two homologues showed similar ubiquitous expression with subtle differences being observed in certain tissues (e.g. root, siliques, and embryos). Neither transgenic manipulation by over-expression of AtSPDSI alone nor its concomitant expression with genes encoding other biosynthetic enzymes (mouse ornithine decarboxylase or Plasmodium falciparum ornithine decarboxylase/S-adenosylmethionine decarboxylase) altered spermidine/spermine content much, which indicates a complex and tightly regulated machinery for higher polyamine homeostasis. The turn-over study on all three major polyamines showed a short half-life of 6-8 h for putrescine and longer ones for spermidine (>40 h) and spermine (>10 h). Catabolic rate of putrescine was proportionate to its endogenous level. Spermine was degraded mainly via back-conversion into spermidine while spermidine showed both back-conversion and terminal catabolism.

The other objective was to destablize 13-glucuronidase protein by attaching PEST signal sequence from mouse ornithine decarboxylase at N or/and C terminus. Modified proteins were shown to have variable reduction in their stability in Arabidopsis seedlings and cell cultures. This should open up a way to use GUS as a more sensitive reporter for transient or short-term gene expression studies.

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