1.5.1
GROWTH PHASE AND TEMPERATURE INFLUENCE PROMOTER ACTIVITY, TRANSCRIPT ABUNDANCE AND PROTEIN STABILITY

I BUDDE1, B ROHDE1, C BENDER2 and M ULLRICH1

1Max-Planck-Institut fr terrestrische Mikrobiologie, Marburg, Germany; 2Oklahoma State University, Stillwater, OK, USA

Background and objectives
The plant pathogenic bacterium Pseudomonas syringae pv. glycinea PG4180.N9 synthesizes high levels of the polyketide phytotoxin coronatine (COR) at 18C, whereas no detectable toxin is produced at 28C. Previously, we reported that the temperature-sensitive activation of three promoters inside the COR biosynthetic gene cluster might explain thermoregulation of COR biosynthesis [1]. The present study was aimed at furthering our understanding of the transcriptional as well as the post-translational effects of temperature on expression of cmaB, which encodes an enzyme involved in COR biosynthesis.

Results and conclusions
Transcriptional fusions using a promoterless glucuronidase gene and Northern blot analyses were used to monitor promoter activities and transcript abundance for the cmaABT operon during bacterial growth at 18 and 28C. Promoter activity and transcription rates were maximal when cells were incubated at 18C and sampled at mid-logarithmic phase. Transcription declined moderately during the transition to stationary phase but remained high at 18C as compared to 28C. Western blot analysis indicated that CmaB accumulated in the late stationary phase of P. syringae cultures grown at 18C but not in cultures incubated at 28C. Temperature-shift experiments indicated that CmaB stability was more pronounced at 18 than at 28C. Although temperature has a strong impact on transcription of COR biosynthetic genes, we propose that thermoregulation of protein stability might also control COR synthesis.

Reference
1. Ullrich M, Penaloza-Vazquez A, Bailey AM, Bender CL, 1995. Journal of Bacteriology 177, 6160-6169.