The extrachromosomal replication of Dictyostelium plasmid Ddp2 requires a cis-acting element and a plasmid-encoded trans-acting factor.
Dictyostelium discoideum plasmid Ddp2 from the wild strain WS380B is a 5.8-kilobase (kb) supercoiled circle with a copy number of 300 per haploid genome. We previously described the construction of an extrachromosomally replicating transformation vector pnDeI carrying 4.7 kb of Ddp2 sequences (B. Leiting, and A. Noegel, Plasmid 20:241-248, 1988). In order to reduce the sequences required for extrachromosomal maintenance in D. discoideum, we characterized Ddp2 by sequence analysis, by deletion experiments, by transcription mapping, by electrophoretic mobility shift assays, and by expression of its single open reading frame in Escherichia coli. Two elements were involved in replication of Ddp2: a cis-acting sequence located on a 592-base-pair (bp) fragment that consisted of 220 bp of essential and 372 bp of auxiliary sequences, and a 2.7-kb open reading frame which most likely encodes a trans-acting factor. The cis- and trans-acting elements did not overlap and were shown to act independently from the location of the sequences encoding the trans-acting factor.
Leiting
B
B
Abteilung Zellbiologie, Max-Planck-Institut fuer Biochemie, Martinsried bei Muenchen, Federal Republic of Germany.
Lindner
I J
IJ
Noegel
A A
AA
D000595:Amino Acid Sequence; D000956:Antigens, Viral / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics*; D004023:Dictyostelium / Q000235:genetics*; D019309:Epstein-Barr Virus Nuclear Antigens; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics*; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid
D000956:Antigens, Viral; D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D005656:Fungal Proteins; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C064562:plasmid Ddp2 protein, Dictyostelium