Interaction between tumour necrosis factor alpha ribozyme and cellular proteins. Involvement in ribozyme stability and activity

Journal of Molecular Biology
M Sioud

Abstract

Ribozymes are RNA molecules that cleave other RNA molecules. Thus, ribozymes offer a new way of inhibiting expression of specific genes whose nucleotide sequences are known. Intracellular stability of ribozymes is an important factor for their efficacy. We previously showed that hammerhead ribozyme directed against mRNA of tumour necrosis factor alpha (TNF alpha) slowly acquires resistance to degradation in cultured human cells. In order to explain this resistance, we now report on endogenous cellular protein(s) that bind to TNF alpha-ribozyme (TNF alpha-Rz) in solution to form stable complexes during native gel electrophoresis. Suppression of the effects of ribonucleases in the cytoplasmic extracts allowed approximately 80% of the input ribozyme RNA to be recovered in the form of complexes, indicating that complex formation protected the ribozyme from degradation. Treatment of the ribozyme-protein complexes with proteinase K prior to electrophoresis led to the recovery of full-length ribozyme. Interestingly, ribozyme-protein complexes retained cleavage activity, suggesting that the binding is in reversible equilibrium. Analysis of protein cytoplasmic extracts for binding to sub-fragments of TNF alpha-Rz demonstrated that prote...Continue Reading

Citations

Jun 13, 1998·Nature Biotechnology·M Sioud, D R Sørensen
Jun 25, 1996·Proceedings of the National Academy of Sciences of the United States of America·P J L'HuillierJ L Vilotte
Dec 25, 1994·Nucleic Acids Research·M SioudK Drlica
Jun 25, 1995·Nucleic Acids Research·O HeidenreichM Nerenberg
Jul 9, 2002·Biochemical and Biophysical Research Communications·Marianne Leirdal, Mouldy Sioud
Jul 15, 1999·Biochemical and Biophysical Research Communications·S L MacKayL L Moldawer
Apr 17, 2003·FEMS Microbiology Reviews·Elena Puerta-FernándezAlfredo Berzal-Herranz
Feb 13, 1997·Biochemical and Biophysical Research Communications·M SioudM J McCall
Jan 1, 1996·Antisense & Nucleic Acid Drug Development·O HeidenreichM Nerenberg
Jul 17, 1999·British Journal of Cancer·M Leirdal, M Sioud
Jun 22, 2000·The Journal of Immunology : Official Journal of the American Association of Immunologists·T C JarvisC M Flory
Oct 23, 2001·American Journal of Physiology. Heart and Circulatory Physiology·P O IversenM Sioud
Apr 21, 1999·Biochimica Et Biophysica Acta·D J Gaughan, A S Whitehead

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