The roles of inhibition of topoisomerases I and II in the formation of chromosomal alterations

Author

Jo Ann Thomas Croom

Date of Award

5-1992

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Biomedical Sciences

Major Professor

R. Julian Preston

Committee Members

Larry Waters, Gayle Littlefield, Raymond Popp

Abstract

Topoisomerases are enzymes which maintain the structural integrity of DNA by transiently breaking the backbone strands of the molecule and passing another DNA strand through the open gate. These enzymes are believed to play important roles in replication, transcription, repair, and packaging of DNA, but precise roles for the two topoisomerase classes are unclear. In an effort to investigate their specific actions in the cell, cytogenetic comparison was made of the effects of inhibiting topoisomerase I (topo I) with camptothecin (CPT) and topoisomerase II (topo II) with 4'-(9- acridinylamino)-methanesulfonic-m-anisidine (mAMSA) at different phases of the cell cycle in human lymphocyte culture. Topo I was inhibited in G₀ and S phases of both the first and second cell cycles after stimulation, while topo II was inhibited in G₀, S of the first and second cell cycles, and G₂. End-point measurements of sister chromatid exchange and chromosomal aberrations were used for analyses of the differential cell cycle responses. The proliferation index was calculated for each treatment.

Sensitivity to inhibition by CPT was found to be equivalent in G₂ and S of the first cell cycle after stimulation, but greatly increased in S phase of the second cell cycle when replication is occurring on a BrdU substituted template. Sensitivity to inhibition by mAMSA was highest in G₂, then 8, and then G₀, with no difference in sensitivity in S of the first and second cell cycles. These results (except for the increased sensitivity to camptothecin in S of the second cell cycle) parallel the known concentrations of the two enzymes at different phases of the cell cycle.

The chromosomal aberration patterns induced by treatment with mAMSA and CPT were very different. Treatment with mAMSA at each phase of the cell cycle produced aberrations consistent with stabilization of the topo ll-DNA cleavable complex leading to either failure to rejoin or reunion via subunit exchange of topo II. Treatment with CPT in both G₀ and S produced aberrations which were chromatid-type and probably a result of stalled replication due to the persistence of induced single-strand breaks. One unusual group of induced aberrations were homologous interchanges. Sister chromatid exchanges were greatly increased with CPT, but only slightly with mAMSA.

Chromosome banding was undertaken to identify the chromosomes involved in homologous interchanges. The pattern of chromosomal alteration induced by topo I inhibition is compared to that in Bloom syndrome, and an explanation is proposed for the similarity.

Recommended Citation

Croom, Jo Ann Thomas, "The roles of inhibition of topoisomerases I and II in the formation of chromosomal alterations. " PhD diss., University of Tennessee, 1992.
https://trace.tennessee.edu/utk_graddiss/10865