Synthesis and processing of RNA by isolated human placental nuclei

Placenta
A S LauH N Munro

Abstract

Conditions for isolating intact and active nuclei from human term pacenta and for studying their transcription products are described. The isolated nuclei can synthesize cell-free RNA for a prolonged period at 29 degree C in a medium containing 100 mM KCl and 5 mM MgCl2. Actinomycin D inhibited 92 per cent of RNA synthesis, whereas approximately 60 per cent of RNA synthesis was sensitive to alpha-amanitin. When nuclei were incubated at 29 degrees C for 1 h, about 27 per cent of the newly synthesized RNA was released into the medium outside the nucleus. Analysis of this released material by affinity chromatography on an oligo(dT)-cellulose column revealed that 2.4 per cent of the total released RNA was adsorbed at high salt concentration. Most of this fraction was eluted with a low-salt buffer at 45 degrees C and the remainder by 50 per cent formamide, conditions that are necessary for elution of poly(A)-containing mRNP particles from oligo(dT)-cellulose. These results show that placental nuclei incubated in vitro synthesize poly(A)-containing RNA, which is released as a protein-bound complex. This procedure allows exploration of changes in mRNA release during placental development.

References

Jun 1, 1977·Nucleic Acids Research·Y Y Mory, M L Gefter
Jul 27, 1976·Biochemistry·D K BiswasA H Tashjian
Feb 24, 1976·Biochemistry·M J ErnestP Feigelson
Oct 1, 1969·Proceedings of the National Academy of Sciences of the United States of America·U Maitra, F Barash
Sep 5, 1972·Biochemical and Biophysical Research Communications·D E Schumm, T E Webb
Jun 1, 1971·Proceedings of the National Academy of Sciences of the United States of America·S Y LeeG Brawerman
Aug 20, 1963·Biochimica Et Biophysica Acta·R M FRANKLIN

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Citations

Apr 15, 1983·Biochimica Et Biophysica Acta·A D BronsteinH N Munro
Nov 1, 1984·Placenta·D Resendez-PerezF J Sanchez-Anzaldo
Jun 1, 1986·Journal of Cellular Physiology·E Freund, P M McGuire

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