Genetic analysis of activation and silencing of the cryptic bgl operon of Escherichia coli

Abstract

Activation and Silencing of the bgl Operon in Escherichia coli: Genetic and Physiological Perspectives

Abstract and Synopsis Introduction The bgl operon of E. coli is one of four genetic systems in Enterobacteriaceae for the utilization of -glucosidic sugars. In wild-type E. coli under laboratory conditions, the operon is silent (Bgl phenotype). Silencing is mediated by negative elements flanking the promoter, including DNA structural motifs and sequences interacting with the nucleoid-associated protein H-NS.

Activated mutants (Bgl ) arise spontaneously, most often due to IS insertions in the regulatory region, which disrupt negative elements. Point mutations in the CRP-binding site and unlinked mutations in loci such as hns, gyr, bglJ, and leuO also activate the operon. Once activated, the operon is inducible by -glucosides (salicin, arbutin) and regulated by transcriptional antitermination.

The evolutionary puzzle lies in the maintenance of a silent operon despite its sophisticated regulation. This thesis investigates the genetic and physiological relevance of bgl activation and silencing.

Key Findings

IS-Mediated Activation and Reversion

Spontaneous revertants of IS-activated strains were analyzed.

Most revertants retained the IS element but showed large chromosomal deletions (Type II deletions), permanently inactivating the operon.

Precise excision of IS elements was not detected under laboratory conditions, suggesting IS-mediated oscillations between silent and active states are unlikely in rich media.

Role of rpoS

rpoS ( ^S), the central regulator of stress response, was tested for involvement in reversion.

No difference in mutation type between rpoS and rpoS strains, but stationary-phase rpoS cultures produced more revertants, indicating ^S suppresses IS-mediated deletions in stationary phase.

Spectrum of Activating Mutations

In rpoS strains, activation was predominantly due to IS insertions in bglR.

In rpoS strains, activation was more often due to mutations in bglJ and hns.

hns mutations relieve repression of stress-responsive genes, suggesting bgl may be part of the ^S regulon.

Physiological Activation under Nutrient Stress

Expression of the wild-type bgl promoter was measured under rich and poor nutrient conditions.

In rpoS strains, expression increased progressively from exponential to stationary phase, especially under nutrient limitation.

In rpoS strains, this increase was marginal.

Starvation-induced expression suggests stress-related regulation, though insufficient for salicin utilization under tested conditions.

Conclusions

IS-mediated oscillations between silent and active states are unlikely under laboratory conditions but may occur in natural environments.

rpoS modulates the spectrum of activating mutations, shifting from local (bglR) to global (hns, bglJ) regulators.

Stress and nutrient limitation transiently enhance bgl expression, supporting its role as a stress-responsive operon.

The ability to utilize aryl -glucosides like salicin may provide E. coli with a competitive advantage under nutrient scarcity.