PERIODIC EXPRESSION OF NUCLEAR AND MITOCHONDRIAL-DNA REPLICATION GENES DURING THE TRYPANOSOMATID CELL-CYCLE

In trypanosomatids, DNA replication in the nucleus and in the single m itochondrion (or kinetoplast) initiates nearly simultaneously, suggest ing that the DNA synthesis (S) phases of the nucleus and the mitochond rion are coordinately regulated. To investigate the basis for the temp oral link between nuclear and mitochondrial DNA synthesis phases the e xpression of the genes encoding DNA ligase I, the 51 and 28 kDa subuni ts of replication protein A, dihydrofolate reductase and the mitochond rial type II topoisomerase were analyzed during the cell cycle progres sion of synchronous cultures of Crithidia fasciculata. These DNA repli cation genes were all expressed periodically, with peak mRNA levels oc curring just prior to or at the peak of DNA synthesis in the synchroni zed cultures. A plasmid clone (pdN-1) in which TOP2, the gene encoding the mitochondrial topoisomerase, was disrupted by the insertion of a NEO drug-resistance cassette was found to express both a truncated TOP 2 mRNA and a truncated topoisomerase polypeptide. The truncated mRNA w as also expressed periodically coordinate with the expression of the e ndogenous TOP2 mRNA indicating that cis elements necessary for periodi c expression are contained within cloned sequences. The expression of both TOP2 and nuclear DNA replication genes at the G(1)/S boundary sug gests that regulated expression of these genes may play a role in coor dinating nuclear and mitochondrial S phases in trypanosomatids.