title: Localization of RNS2 ribonuclease to the vacuole is required for its role in cellular homeostasis.
abstract: Localization of the RNase RNS2 to the vacuole via a C-terminal targeting signal is essential for its function in rRNA degradation and homeostasis. RNase T2 ribonucleases are highly conserved enzymes present in the genomes of nearly all eukaryotes and many microorganisms. Their constitutive expression in different tissues and cell types of many organisms suggests a housekeeping role in RNA homeostasis. The Arabidopsis thaliana class II RNase T2, RNS2, is encoded by a single gene and functions in rRNA degradation. Loss of RNS2 results in RNA accumulation and constitutive activation of autophagy, possibly as a compensatory mechanism. While the majority of RNase T2 enzymes is secreted, RNS2 is located within the vacuole and in the endoplasmic reticulum (ER), possibly within ER bodies. As RNS2 has a neutral pH optimum, and the endomembrane organelles are connected by vesicle transport, the site within the endomembrane system at which RNS2 functions is unclear. Here we demonstrate that localization to the vacuole is essential for the physiological function of RNS2. A mutant allele of RNS2, rns2-1, results in production of an active RNS2 RNase but with a mutation that removes a putative C-terminal vacuolar targeting signal. The mutant protein is, therefore, secreted from the cell. This results in a constitutive autophagy phenotype similar to that observed in rns2 null mutants. These findings illustrate that the intracellular retention of RNS2 and localization within the vacuole are critical for its cellular function.
authors.affiliation: Department of Genetics, Development and Cell Biology, Iowa State University, Ames, IA, 50011, USA.
authors.forename: Brice E
authors.initials: BE
authors.lastname: Floyd
authors.affiliation: Department of Genetics, Development and Cell Biology, Iowa State University, Ames, IA, 50011, USA.
authors.forename: Yosia
authors.initials: Y
authors.lastname: Mugume
authors.affiliation: Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, IA, 50011, USA.
authors.forename: Stephanie C
authors.initials: SC
authors.lastname: Morriss
authors.affiliation: Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, IA, 50011, USA. gustavo@iastate.edu.
authors.forename: Gustavo C
authors.initials: GC
authors.lastname: MacIntosh
authors.affiliation: Department of Genetics, Development and Cell Biology, Iowa State University, Ames, IA, 50011, USA. bassham@iastate.edu.
authors.forename: Diane C
authors.initials: DC
authors.lastname: Bassham
mesh_terms: D017360:Arabidopsis / Q000201:enzymology / Q000502:physiology; D029681:Arabidopsis Proteins / Q000502:physiology*; D001343:Autophagy / Q000502:physiology; D003001:Cloning, Molecular; D006706:Homeostasis / Q000502:physiology; D011523:Protoplasts / Q000502:physiology; D012335:RNA, Ribosomal / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012260:Ribonucleases / Q000502:physiology*; D014617:Vacuoles / Q000201:enzymology* / Q000502:physiology
chemical_list: D029681:Arabidopsis Proteins; D012335:RNA, Ribosomal; C489907:RNS2 protein, Arabidopsis; D012260:Ribonucleases
keywords: Arabidopsis; Autophagy; RNA degradation; Ribosomal RNA; Vacuolar targeting