Expression of the oligomerization domain of the replication-associated protein (Rep) of Tomato leaf curl New Delhi virus interferes with DNA accumulation of heterologous geminiviruses.
The minimal DNA binding domain of the replication-associated protein (Rep) of Tomato leaf curl New Delhi virus was determined by electrophoretic mobility gel shift analysis and co-purification assays. DNA binding activity maps to amino acids 1-160 (Rep-(1-160)) of the Rep protein and overlaps with the protein oligomerization domain. Transient expression of Rep protein (Rep-(1-160)) was found to inhibit homologous viral DNA accumulation by 70-86% in tobacco protoplasts and in Nicotiana benthamiana plants. The results obtained showed that expression of N-terminal sequences of Rep protein could efficiently interfere with DNA binding and oligomerization activities during virus infection. Surprisingly, this protein reduced accumulation of the African cassava mosaic virus, Pepper huasteco yellow vein virus and Potato yellow mosaic virus by 22-48%. electrophoretic mobility shift assays and co-purification studies showed that Rep-(1-160) did not bind with high affinity in vitro to the corresponding common region sequences of heterologous geminiviruses. However, Rep-(1-160) formed oligomers with the Rep proteins of the other geminiviruses. These data suggest that the regulation of virus accumulation may involve binding of the Rep to target DNA sequences and to the other Rep molecules during virus replication.
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International Laboratory for Tropical Agricultural Biotechnology, Donald Danforth Plant Science Center, 8001 Natural Bridge Road, St. Louis, MO 63121, USA.
Beachy
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Fauquet
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D001665:Binding Sites; D002874:Chromosome Mapping; D004265:DNA Helicases / Q000235:genetics* / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism*; D004268:DNA-Binding Proteins; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D017791:Geminiviridae / Q000235:genetics* / Q000502:physiology; D010449:Peptide Mapping; D011108:Polymers; D011487:Protein Conformation; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014764:Viral Proteins / Q000378:metabolism*; D014779:Virus Replication
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D011108:Polymers; D015534:Trans-Activators; D014764:Viral Proteins; C045264:replication initiator protein; D004265:DNA Helicases