Home Economics American Trypanosomiasis Chagas Disease, Second Edition: One Hundred Years of Research
Replication model of kDNA
The structure of the kDNA network has an unusual replication mechanism.
Replication of kDNA minicircles has been first characterized in Crithidia fasci- culata (for review see Shapiro and Englund5). Nevertheless, the replication mechanism is similar to that of T. cruzi.6 The minicircles are not linked to the network when they replicate themselves.
Their replication is initiated at dodecamer 5'-GGGGTTGGTGTA-3' called the universal minicircle sequence (UMS). During the early stages of replication, this dodecamer binds specifically to a zing finger protein called UMS-binding protein. This UMS sequence is identical for trypanosomid minicircles and the UMS-binding protein sequence presents a high similarity between various Trypanosoma species.7
The minicircles may be freed by the enzyme topoisomerase II, enabling them to replicate freely. They are in a natural state closed covalently in their replication. The first step is replication in 0 like structures, then, in the second step, Okazaki fragments are synthesized and their descendants, which contain gaps, are subsequently reconnected to the periphery of the network.6 This mechanism is fundamentally different from that of all others cells. In either prokaryotes or eukaryotes, Okazaki fragments are ligated immediately after their synthesis.8
When the network is being replicated, the central region which is not involved diminishes in size, and the peripheral region containing the new replicates of the minicircles with gaps, grows bigger. Once the minicircles have replicated (minicircles containing the gaps), the number has doubled.
Whereas the connection sites are opposed, the new minicircles containing gaps are rapidly uniformly distributed around the periphery of the network in a sequential manner. This uniform distribution is considered to be a movement relative to the kinetoplast disk as well as to the protein complexes. It has been suggested that the kinetoplast disk rotates between the two protein complexes. Due to the distribution of recently replicated minicircles in the network, it has been called the “annular” replication mechanism.6 This kind of movement during replication of T. cruzi kDNA is similar in Crithidia fasciculata, and Leishmania species but is distinct in T. brucei where minicircles are during replication removed from the central region and reattached at the poles adjacent to theantipodal sites.9
In order for this mechanism to function, it is necessary for the gaps between the Okazaki fragments to be repaired, and then the network divides into two. The latter process is possibly mediated by the enzymes contained in the antipodal site such as endonuclease I (SSE-1), responsible for primer removing and which colocalizes with the kinetoplast topoisomerase II and DNA polymerase в during replication.10 These enzymes then untie neighboring minicircles along the cleavage line of the network.6
Following primer removal, the gaps between fragments are repaired by DNA polymerase в and DNA ligase ke as well as mitochondrial DNA helicase.4 The new replicates of the minicircles are connected, in two opposite positions, to the periphery of the network. Several authors have suggested that these positions are adjacent to two protein complexes, known to contain topoisomerase II and a DNA polymerase в.11 As the structure trypanosomids topoisomerases is specific, this class of enzyme represents a potential therapeutic drug target.12 They use berenil, which inhibits the minicircle decatenation of the network and thus produces significant changes in kDNA arrangement affecting T. cruzi proliferation, has been described.13
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