|Oliver, Melvin - Mel|
Submitted to: Proceedings Annual Riverside Symposium in Plant Physiology
Publication Type: Proceedings
Publication Acceptance Date: 7/15/1996
Publication Date: N/A
Citation: Interpretive Summary: Interpretive summary not required.
Technical Abstract: In response to rehydration, desiccated Torula ruralis produces a set of peptides unique to the rehydrated state; viz. rehydrins. Present evidence concludes that T. ruralis pre-loads mRNAs into a messenger ribonucleotide particle (mRNP) during desiccation. Density gradient analysis and FPLC purification of RNPs containing poly-(A+) RNA have demonstrated that mRNPs exist both in desiccated and hydrated gametophytes. However, desiccation induces the appearance of several high molecular weight complexes that are absent in hydrated controls. Utilizing cDNAs to the abundant rehydrin Tr288 and quantitative RT-PCR, we have demonstrated the sequestration of rehydrin transcripts within these desiccation-induced mRNPs. The interaction of T. ruralis proteins with T. ruralis mRNA during desiccation has also been demonstrated using RNA gel retardation analysis. A [32P]-labeled, 3' fragment of Tr288 forms a stable mRNA-protein complex under native conditions and we are delineating the mRNA protein-binding sequence(s). With this information, we will be able to purify the RNA-binding protein(s) (using affinity chromatography) and clone the corresponding cDNA(s) (by direct screening of an expression library). We can also demonstrate protein-binding using 5' [32P]-labeled probe. However, our Tr288 cDNA is lacking 43 bp of the 5' UTR which, at this time, limits our ability to define these protein-binding region(s). To obtain a clearer view of the 5' UTR, we are using 5'-RACE to generate full-length clones. The RNA-binding characters of both the 5' and 3' UTRs will be vital to understanding mRNP formation, altered mRNA stability and post-transcriptional gene regulation. These studies will be presented and discussed in relation to proposed mechanisms of desiccation-tolerance.