Alternative pre-rRNA processing pathways in human cells and their alteration by cycloheximide inhibition of protein synthesis.
rRNA processing pathways in humans have been reinvestigated through systematic Northern-blot hybridizations of HeLa cell nuclear RNA with a collection of digoxigenin-labeled rDNA probes from different regions of the human rDNA transcriptional unit. In addition to the known 45S, 41S, 32S and 21S pre-rRNA, two major pre-rRNA fractions were identified; a '30S' (about 5800 nucleotides) precursor to 18S rRNA containing an external transcribed spacer at the 5' end (ETS) and internal transcribed spacer (ITS) 1 sequences and a '12S' (about 950 nucleotides) precursor to 5.8S rRNA containing ITS 2 sequences. These pre-rRNA species do not react with probes located near the 3'-terminal segments of ITS 1 or ITS 2, thus suggesting that processive endonuclease cuts occur within ITS spacer sequences. The simultaneous occurrence of at least two alternative 45S pre-rRNA processing pathways is deduced, which correspond to a different temporal order of endonuclease attack at the sites located near the 5' end of 18S rRNA and within ITS 1. In-vivo labeling experiments with [14C]uridine revealed that inhibition of protein synthesis with cycloheximide abolishes the endonuclease cut at the 5' end of 18S rRNA and the formation of 41S pre-rRNA, while the cut within ITS 1 and the processing to 32S and '30S' pre-rRNA remains relatively unaltered.
Hadjiolova
K V
KV
Laboratoire de Biologie Moléculaire Eucaryote, Université Paul-Sabatier, Toulouse, France.
Nicoloso
M
M
Mazan
S
S
Hadjiolov
A A
AA
Bachellerie
J P
JP
D001483:Base Sequence; D015152:Blotting, Northern; D003513:Cycloheximide / Q000494:pharmacology*; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D012322:RNA Precursors / Q000378:metabolism*; D012323:RNA Processing, Post-Transcriptional / Q000187:drug effects*; D012335:RNA, Ribosomal / Q000378:metabolism*
D012322:RNA Precursors; D012335:RNA, Ribosomal; D003513:Cycloheximide