Plasmid-dependent temperature-sensitive phase in crown gall tumorigenesis

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Abstract
Two methods were used to show that the temperature-sensitive (ts) genes for plasmid maintenance located on pTiC58 are responsible for the presence of a specific 37.degree. C ts period during crown gall tumorigenesis. Tumors [in Helianthus annus] apparently induced by Agrobacterium tumefaciens strain C58 become resistant to 32.degree. C by 96 h after infection, indicating that the inception phase of crown gall is complete at that time. C58 tumors remain sensitive to 37.degree. C until 168 h after infection. When pTiC58 is replaced in strain C58 by a plasmid that is temperature resistant for maintenance within the bacterium at 37.degree. C (B6806) the tumors induced by the new strain (A277) become resistant to 37.degree. C by 96 h after infection. When pTiC58 is selected for temperature resistance in strain C58 at 37.degree. C, the tumors induced by the new strain, C58TR, become resistant to 37.degree. C approximately 40 h earlier than do tumors induced by the parent strain C58. The 37.degree. C ts period (96-168 h after infection) is apparently coded for ts plasmid maintenance genes located on pTiC58. Incorporation of [3H]-thymidine into pTiC58 is specifically inhibited at 37.degree. C compared to chromosomal DNA and is not inhibited at 32.degree. C. The data presented support a mechanism in which the 37.degree. C, ts period between 96-168 h after infection is due to the requirement for the plasmid-coded maintenance genes involved in plasmid replication within incipient tumor cells. The fact that the 37.degree. C ts period ends after 168 h indicates a transient requirement for the ts gene product, perhaps due to the stable integration of plasmid DNA into plant genetic elements by that time.