Genomic organization and organ-specific expression of a nuclear gene encoding phage-type RNA polymerase in Nicotiana sylvestris

Gene. 2001 Nov 14;279(1):33-40. doi: 10.1016/s0378-1119(01)00729-6.

Abstract

We have isolated and sequenced a nuclear gene and cDNA encoding bacteriophage T7-type single subunit RNA polymerase, NsRpoT-A, from Nicotiana sylvestris. NsRpoT-A consists of 19 exons and 18 introns; the first intron is 17 kb, the longest yet identified in a plant gene. Genomic Southern analysis indicated that N. sylvestris contains a small family of NsRpoT genes. Quantitative RT-PCR revealed that steady-state mRNA levels are highest in the leaves and lowest in the cotyledons. Phylogenetic analysis of NsRpoT-A and the RpoT proteins of other plant species suggested that NsRpoT-A is a mitochondrial protein. The TargetP program predicted localization of the NsRpoT-A gene product to the mitochondria. Using a transient expression assay and protoplasts from N. sylvestris mesophyll cells, we clearly demonstrated that the N-terminal sequence of NsRpoT-A targets the protein to the mitochondria. We therefore named this protein NsRpoTm.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Bacteriophage T7 / enzymology
  • Biological Transport
  • Blotting, Northern
  • Cell Nucleus / genetics
  • DNA, Complementary / chemistry
  • DNA, Complementary / genetics
  • DNA, Complementary / isolation & purification
  • DNA, Plant / chemistry
  • DNA, Plant / genetics
  • DNA, Plant / isolation & purification
  • DNA-Directed RNA Polymerases / genetics*
  • DNA-Directed RNA Polymerases / metabolism
  • Exons
  • Gene Expression Regulation, Plant
  • Genes, Plant / genetics*
  • Green Fluorescent Proteins
  • Introns
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Mitochondria / metabolism
  • Molecular Sequence Data
  • Phylogeny
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Sequence Alignment
  • Sequence Analysis, DNA
  • Sequence Homology, Amino Acid
  • Tissue Distribution
  • Tobacco / enzymology
  • Tobacco / genetics*

Substances

  • DNA, Complementary
  • DNA, Plant
  • Luminescent Proteins
  • RNA, Messenger
  • Recombinant Fusion Proteins
  • Green Fluorescent Proteins
  • DNA-Directed RNA Polymerases

Associated data

  • GENBANK/AB058954
  • GENBANK/AB058955
  • GENBANK/AB058956