Arabidopsis chloroplast mini-ribonuclease III participates in rRNA maturation and intron recycling.
RNase III proteins recognize double-stranded RNA structures and catalyze endoribonucleolytic cleavages that often regulate gene expression. Here, we characterize the functions of RNC3 and RNC4, two Arabidopsis thaliana chloroplast Mini-RNase III-like enzymes sharing 75% amino acid sequence identity. Whereas rnc3 and rnc4 null mutants have no visible phenotype, rnc3/rnc4 (rnc3/4) double mutants are slightly smaller and chlorotic compared with the wild type. In Bacillus subtilis, the RNase Mini-III is integral to 23S rRNA maturation. In Arabidopsis, we observed imprecise maturation of 23S rRNA in the rnc3/4 double mutant, suggesting that exoribonucleases generated staggered ends in the absence of specific Mini-III-catalyzed cleavages. A similar phenotype was found at the 3' end of the 16S rRNA, and the primary 4.5S rRNA transcript contained 3' extensions, suggesting that Mini-III catalyzes several processing events of the polycistronic rRNA precursor. The rnc3/4 mutant showed overaccumulation of a noncoding RNA complementary to the 4.5S-5S rRNA intergenic region, and its presence correlated with that of the extended 4.5S rRNA precursor. Finally, we found rnc3/4-specific intron degradation intermediates that are probable substrates for Mini-III and show that B. subtilis Mini-III is also involved in intron regulation. Overall, this study extends our knowledge of the key role of Mini-III in intron and noncoding RNA regulation and provides important insight into plastid rRNA maturation.
Boyce Thompson Institute for Plant Research, Ithaca, New York 14853.
Amber M
AM
Hotto
Boyce Thompson Institute for Plant Research, Ithaca, New York 14853.
Benoît
B
Castandet
Centre National de la Recherche Scientifique FRE3630, Université de Paris Diderot, Sorbonne Paris Cité, Institut de Biologie Physico-Chimique, 75005 Paris, France.
Laetitia
L
Gilet
Boyce Thompson Institute for Plant Research, Ithaca, New York 14853.
Andrea
A
Higdon
Centre National de la Recherche Scientifique FRE3630, Université de Paris Diderot, Sorbonne Paris Cité, Institut de Biologie Physico-Chimique, 75005 Paris, France.
Ciarán
C
Condon
Boyce Thompson Institute for Plant Research, Ithaca, New York 14853 ds28@cornell.edu.
David B
http://orcid.org/0000-0002-0653-6602
DB
Stern
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001412:Bacillus subtilis / Q000378:metabolism; D001483:Base Sequence; D002736:Chloroplasts / Q000378:metabolism*; D019143:Evolution, Molecular; D005091:Exons / Q000235:genetics; D005816:Genetic Complementation Test; D007438:Introns / Q000235:genetics*; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011132:Polyribosomes / Q000378:metabolism; D017434:Protein Structure, Tertiary; D020871:RNA Stability; D012335:RNA, Ribosomal / Q000235:genetics* / Q000378:metabolism; D012338:RNA, Ribosomal, 23S / Q000235:genetics; D022661:RNA, Untranslated / Q000235:genetics; D043244:Ribonuclease III / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism; D017423:Sequence Analysis, RNA; D017386:Sequence Homology, Amino Acid; D019076:Transgenes
D029681:Arabidopsis Proteins; D012335:RNA, Ribosomal; D012338:RNA, Ribosomal, 23S; D022661:RNA, Untranslated; C000606165:RNC3 protein, Arabidopsis; C000606166:RNC4 protein, Arabidopsis; D043244:Ribonuclease III