Date of Award
Doctor of Philosophy
Genetics, Development and Cell Biology
Diane C. Bassham
The degradation and turnover of cellular components is essential for biological systems to remove damaged components and maintain functional intracellular contents. Autophagy is a vesicle-mediated process by which cellular contents can be degraded through trafficking to the lysosome or vacuole where lytic processes occur. The majority of a cell’s RNA and large quantities of its protein can be found in ribosomes. Ribosomes represent a significant metabolic investment. However, little is known about the turnover of these highly abundant ribonuclear organelles in plants.
RNase T2 enzymes represent a widely conserved class of ribonucleases present in the genomes of nearly all eukaryotes tested. Here I report on a function of the Arabidopsis T2 RNase, RNS2, and autophagy in the degradation of RNA within the vacuole under normal growing conditions. When RNS2 is knocked out in Arabidopsis, autophagy is increased. Autophagic vesicles in mutants contained RNA and ribosomal subunits. Both autophagy and RNS2 are involved in the degradation of rRNA in Arabidopsis. rns2 mutants also accumulated ribosomal RNA within the vacuole that was eliminated in an atg5 background, indicating autophagy is used as a pathway for rRNA transport into the vacuole for degradation. The functional localization of the RNS2 protein in plants was also unknown. RNS2 was previously shown to be localized to endoplasmic reticulum bodies and the vacuole. Studies here, report that intracellular retention and vacuolar localization are required for RNS2’s physiological function.
As an alternative degradative RNA transport mechanism, my studies also show that RNA can be directly transported into the vacuole via a trans-membrane, macroautophagy-independent pathway. Exogenously added RNA accumulated within purified Arabidopsis vacuoles and required the hydrolysis of ATP to facilitate transport. A DEVH-box helicase mutant shows reduced RNA transport and an increased autophagy phenotype, indicating it may be involved in an RNautophagy-like mechanism in plants.
Taken together, results reported herein show that autophagy and RNS2 ribonuclease function in the degradation of rRNA in Arabidopsis under normal growing conditions, and that RNA degradation carried out by RNS2 occurs within the vacuole. This work also provides evidence for a novel mechanism of RNA transport into the vacuole via an ATP-dependent, trans-membrane transport pathway in Arabidopsis.
Brice Edward Floyd
Floyd, Brice Edward, "Analysis of the role of RNS2 and autophagy in the turnover of ribosomal RNA in Arabidopsis thaliana" (2015). Graduate Theses and Dissertations. 14826.