Synergistic binding of the Vibrio fischeri LuxR transcriptional activator domain and RNA polymerase to the lux promoter region.

Abstract

LuxR, the Vibno fischeri luminescence gene (lux) activator, is the best-studied member ofa family of bacterial transcription factors required for cell density-dependent expression of specific genes involved in associations with eukaryotic hosts. Neither LuxR nor any other LuxR homolog has been shown to bind DNA directly. We have purified the LuxR C-terminal transcriptional activator domain from extracts of recombinant Escherichia coli in which this polypeptide was expressed. The purified polypeptide by itself binds to lux regulatory DNA upstream of the lux box, a 20-bp palindrome that is required for LuxR activity in vivo, but it does not bind to the lux box. However, the LuxR C-terminal domain together with RNA polymerase protects a region including the lux box and the lux operon promoter from DNase I cleavage. There is very little protection of the lux operon promoter region from DNase I digestion in the presence of RNA polymerase alone. Apparently, there is a synergistic binding of the LuxR C-terminal domain and RNA polymerase to the promoter region LuxR-facilitated autoinduction controls transcription of luminescence genes in Vibriofischeri. LuxR homologs occur in a number of different Gram-negative -bacteria, and these transcription factors are involved in a phenomenon termed quorum sensing and response (for recent reviews, see refs.

Bacterial Hash Function Using DNA-Based XOR Logic Reveals Unexpected Behavior of the LuxR Promoter

“…Perhaps the lower average and larger error bars is due to stochastic binding of 3OC6 and the resulting inconsistent forward vs. backward transcription from the same promoter. The literature indicates that LuxR binds Plux only after 3OC6 binds to LuxR 14,15,[24][25][26] . Our results show that the BioBrick part containing Plux supports backwards transcription when LuxR is present in the cell and 3OC6 is absent.…”

“…Our results indicated that the Plux promoter is bidirectional and is induced "backwards" by LuxR in the absence of 3OC6. The backwards transcription in the absence of 3OC6 had not been documented in the literature 14,15,[24][25][26] . In the V. fischeri genome, the Plux promoter points in the direction of the LuxI gene and away from the LuxR gene.…”

“…The purpose of the (promoter)+RBS+GFP constructs was to quantify the level of transcription in the direction the promoter is pointing, while the RFPrev+RBSrev+(promoter) constructs was used to quantify the amount of transcription caused by each promoter in the reverse direction. Note that the pre-existing part (BBa_K09100) containing Plux +RBS+GFP also contained Plac +RBS+LuxR+TT (TT is a transcription terminators) to the left of it because LuxR is needed for Plux to be activated 25 .…”

When being Alone is Enough: Noncanonical Functions of Canonical Bacterial Quorum-Sensing Systems

“…Briefly, LuxI is a synthase that catalyzes a reaction between S-adenosyl methionine and an acyl carrier protein to produce an acyl-homoserine lactone (AHL) that freely diffuses out of the bacterial cell [7]. At high concentrations, the AHL will diffuse back into the cell and bind to its cognate cytoplasmic receptor LuxR and induce the transcription of the luciferase operon, forming an autoregulatory feedback loop (Figure 1) [8][9][10][11][12]. Since this first description, numerous LuxI/LuxR homologous systems have been identified in more than a 100 Gram-negative bacteria and have been shown to regulate a variety of genes involved in a plethora of functions including virulence gene expression, motility, biosynthesis of bacteriocins and biofilm formation [1,3,13].…”

Determination of Extracellular Molecules Produced by Vibrio Harveyi Using MS/MS

“…As more AI is produced, their concentration starts to increase outside the cell, diffusing back into the cell and binding more frequently to LuxR, forming a stable complex. This complex will then activate the luxICDABE operon that codes for luciferase (Stevens et al, 1994). Without the AHL, LuxR is very unstable and gets degraded, and only with a high concentration of the autoinducer can the stable AHL-LuxR complex be formed.…”