The N-terminal domain that distinguishes yeast from bacterial RNase III contains a dimerization signal required for efficient double-stranded RNA cleavage.
Yeast Rnt1 is a member of the double-stranded RNA (dsRNA)-specific RNase III family identified by conserved dsRNA binding (dsRBD) and nuclease domains. Comparative sequence analyses have revealed an additional N-terminal domain unique to the eukaryotic homologues of RNase III. The deletion of this domain from Rnt1 slowed growth and led to mild accumulation of unprocessed 25S pre-rRNA. In vitro, deletion of the N-terminal domain reduced the rate of RNA cleavage under physiological salt concentration. Size exclusion chromatography and cross-linking assays indicated that the N-terminal domain and the dsRBD self-interact to stabilize the Rnt1 homodimer. In addition, an interaction between the N-terminal domain and the dsRBD was identified by a two-hybrid assay. The results suggest that the eukaryotic N-terminal domain of Rnt1 ensures efficient dsRNA cleavage by mediating the assembly of optimum Rnt1-RNA ribonucleoprotein complex.
Lamontagne
B
B
Département de Microbiologie et d'Infectiologie, Faculté de Médecine, Université de Sherbrooke, Sherbrooke, Québec, Canada J1H 5N4.
Tremblay
A
A
Abou Elela
S
S
D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D019281:Dimerization; D004722:Endoribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012330:RNA, Double-Stranded / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012331:RNA, Fungal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D043244:Ribonuclease III; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques
D017931:DNA Primers; D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D012330:RNA, Double-Stranded; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; D004722:Endoribonucleases; C468367:RNT1 protein, S cerevisiae; D043244:Ribonuclease III; C468366:ribonuclease III, E coli