CHANGES IN THE STRUCTURE OF NUCLEOIDS ISOLATED FROM HEAT-SHOCKED HELA-CELLS

Abstract

Using a technique to detect changes in DNA supercoiling which allows one to visualize both DNA unwinding and rewinding in presence of the intercalating dye, propidium iodide (PI), we showed that hyperthermic treatment (30 min at 45 degrees C) of HeLa S3 cells alters the response to the intercalating dye. Depending on the treatment conditions, we observed a reduction in the maximum size of the DNA loop that can be measured at the relaxation point (PI concentration 5-7.5 micrograms/ml). Cellular heating also affected all degrees of DNA rewinding (measured as a function of PI concentrations between 10 and 50 micrograms/ml). By 6 h after cellular heating these heat effects had disappeared. This time interval correlated with the time necessary for recovery from a heat-induced increase to normal nuclear and nucleoid protein content. Using gel electrophoresis we showed that the nucleoids (DNA plus nuclear matrix proteins) after heat exposure are enriched in several polypeptides and that there is a specific increase in HSP 72/73. We hypothesize that the altered response to the intercalating dye after cellular heat shock is due to an increase in polypeptides associated with the nuclear matrix thereby altering the DNA-nuclear protein matrix anchor point