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Nucleic Acids Research, 1984, Vol. 12, No. 23 8971-8985
© 1984

MOLECULAR BIOLOGY

Zinc ions are differentially required for the transcription of ribosomal 5S RNA and tRNA in a HeLa-cell extract

Edgar Wingender, Dagmar Dilloo and Klaus H. Seifart

Institut für Physiologische Chemie I, Universitaät Marburg 3550 Marburg/Lahnberge, FRG

Received August 21, 1984. Revised October 30, 1984. Accepted November 13, 1984.

Chelation of divalent cations by 5 mM EDTA and subsequent removal by dialysis from a cytoplasmic HeLa cell extract leads to a complete loss of 5S rRNA transcription without affecting tRNA synthesis. Transcription complexes for 5S RNA can no longer be assembled in such a zinc-depleted extract and this ability can be fully restored only by the re-addition of 5 µM zinc. Reconstitution experiments with isolated protein fractions show that transcription factor A from HeLa-cells requires zinc to exert its specific function. Pre-formation of transcription complexes partially protects the metal ion against removal by chelation even in the presence of 1.8 M KC1. These results indicate that the zinc ions are bound to mammalian transcription factor IIIA which, in a transcription complex, binds very strongly to the 5S RNA gene. Cation depletion with 75 mM EDTA also suppresses tRNA transcription; an effect which is reversible by zinc addition. We conclude that beside for the binding of TF IIIA, zinc is also bound with a different affinity to a transcription component common to 5S and tRNA synthesis, most likely polymerase III itself.


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