Influence of 5'-flanking sequence on 4.5SI RNA gene transcription by RNA polymerase III

Genome. 2018 May;61(5):367-370. doi: 10.1139/gen-2017-0223. Epub 2018 Feb 2.

Abstract

Short nuclear 4.5SI RNA can be found in three related rodent families. Its function remains unknown. The genes of 4.5SI RNA contain an internal promoter of RNA polymerase III composed of the boxes A and B. Here, the effect of the sequence immediately upstream of the mouse 4.5SI RNA gene on its transcription was studied. The gene with deletions and substitutions in the 5'-flanking sequence was used to transfect HeLa cells and its transcriptional activity was evaluated from the cellular level of 4.5SI RNA. Single-nucleotide substitutions in the region adjacent to the transcription start site (positions -2 to -8) decreased the expression activity of the gene down to 40%-60% of the control. The substitution of the conserved pentanucleotide AGAAT (positions -14 to -18) could either decrease (43%-56%) or increase (134%) the gene expression. A TATA-like box (TACATGA) was found at positions -24 to -30 of the 4.5SI RNA gene. Its replacement with a polylinker fragment of the vector did not decrease the transcription level, while its replacement with a GC-rich sequence almost completely (down to 2%-5%) suppressed the transcription of the 4.5SI RNA gene. The effect of plasmid sequences bordering the gene on its transcription by RNA polymerase III is discussed.

Keywords: ARN non-codant; ARN polymérase III; RNA polymerase III; TATA box; boîte TATA; non-coding RNA; promoter; promoteur; transcription.

MeSH terms

  • 5' Flanking Region*
  • Animals
  • Base Sequence
  • HeLa Cells
  • Humans
  • Mice
  • Plasmids / chemistry
  • Plasmids / metabolism
  • Point Mutation
  • Promoter Regions, Genetic
  • RNA Polymerase III / genetics*
  • RNA Polymerase III / metabolism
  • RNA, Small Nuclear / genetics*
  • RNA, Small Nuclear / metabolism
  • Sequence Alignment
  • Sequence Deletion*
  • Sequence Homology, Nucleic Acid
  • Transcription Initiation Site
  • Transcription, Genetic*
  • Transfection

Substances

  • RNA, Small Nuclear
  • RNA Polymerase III