Two RpoT genes of Physcomitrella patens encode phage-type RNA polymerases with dual targeting to mitochondria and plastids.
Angiosperms possess a small family of phage-type RNA polymerase genes that arose by gene duplication from an ancestral gene encoding the mitochondrial RNA polymerase. We have isolated and sequenced the genes and cDNAs encoding two phage-type RNA polymerases, PpRpoT1 and PpRpoT2, from the moss Physcomitrella patens. PpRpoT1 comprises 19 exons and 18 introns, PpRpoT2 contains two additional introns. The N-terminal transit peptides of both polymerases are shown to confer dual-targeting of green fluorescent protein fusions to mitochondria and plastids. In vitro translation of the cDNAs revealed initiation of translation at two in-frame AUG start codons. Translation from the first methionine gives rise to a plastid-targeted polymerase, whereas initiation from the second methionine results in exclusively mitochondrial-targeted protein. Thus, dual-targeting of Physcomitrella RpoT is caused by and might be regulated by multiple translational starts. In phylogenetic analyses, the Physcomitrella RpoT polymerases form a sister group to all other phage-type polymerases of land plants. The two genes result from a gene duplication event that occurred independently from the one which led to the organellar polymerases with mitochondrial or plastid targeting properties in angiosperms. Yet, according to their conserved exon-intron structures they are representatives of the molecular evolutionary line leading to the RpoT genes of higher land plants.
Richter
Uwe
U
Institut für Biologie, Humboldt-Universität, Chausseestrasse 117, Berlin, Germany.
Kiessling
Justine
J
Hedtke
Boris
B
Decker
Eva
E
Reski
Ralf
R
Börner
Thomas
T
Weihe
Andreas
A
D000595:Amino Acid Sequence; D019068:Bryopsida / Q000201:enzymology / Q000235:genetics*; D003001:Cloning, Molecular; D018744:DNA, Plant / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism; D005091:Exons; D017343:Genes, Plant / Q000187:drug effects; D049452:Green Fluorescent Proteins; D007438:Introns; D007527:Isoenzymes / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D010802:Phylogeny; D010940:Plant Proteins / Q000235:genetics*; D018087:Plastids / Q000201:enzymology*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D018744:DNA, Plant; D007527:Isoenzymes; D008164:Luminescent Proteins; D010940:Plant Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D012321:DNA-Directed RNA Polymerases