2-DIMENSIONAL PULSED-FIELD GEL-ELECTROPHORESIS OF YEAST CHROMOSOMES -EVIDENCE FOR TRIPLEX-MEDIATED DNA CONDENSATION

The mobility of yeast chromosomes was analysed by two-dimensional puls ed-field gel electrophoresis. The first dimension was run at pH 8.0 in a 1% agarose gel. In the second dimension the electrophoresis conditi ons were identical, except that the pH was lowered and ethidium, sperm ine, or ionic detergents were added. Any mobility changes between the two dimensions could be identified as a deviation from the diagonal. A t pH 6.0 the mobility of the chromosomes increases severalfold, wherea s at pH 4.5 none of the chromosomes move into the agarose gel. The pH- induced mobility changes were reversed by the addition of 2 mug/mL of ethidium or 1% lauryl sarcosine. Alternatively, spermine at 1 muM enha nced the pH-mediated mobility changes. Hysteresis was also evident, si nce upon lowering the pH to 4.5 and then running the gel at pH 7 the m obilities were decreased. These results are interpreted in terms of pH -mediated triplex formation which causes chromosome condensation and t hus mobility shifts. The effects of pH are reversed by ethidium which destabilizes triplexes, but enhanced by spermine which favours triplex formation. Therefore, chromosomes may be capable of spontaneous conde nsation which is mediated by tertiary interactions between appropriate duplex DNA sequences.