Interactions between the Repressor and the Early Operator Region of Bacteriophage Mu

Abstract

The repressor of bacteriophage Mu, c, binds to three operator sites, O1, O2, and O3, overlapping two divergent promoters, which regulate the lytic and lysogenic pathways. Its binding to this operator region generates several complexes, which were analyzed by DNase I protection experiments. We demonstrate that c first binds to two 11-base pair partially repeated sequences in O2 that could represent "core" binding sites for the repressor. This initial interaction serves as an organizer of a more complex nucleoprotein structure in which O2, O1, and O3 become successively occupied. The quaternary structure of the repressor was also investigated. Size exclusion chromatography and protein-protein crosslinking experiments with chemicals that possess linking arms of various lengths indicate that the repressor oligomerizes in solution. A model is proposed describing the successive interactions of c with the operator sites O2, O1, and O3 leading to the elaboration of a higher order structure in which the early lytic functions are repressed.The repressor, c, of the temperate bacteriophage Mu controls the choice between lysogeny and lytic development. The decision is exercised at the level of transcription by binding of c to the early operator region located about 1 kilobase from the left end of the phage genome ( Fig. 1). This operator is contained within a 200-bp 1 region composed of two divergent promoters (Pe and Pc) and three operator sequences, O1, O2, and O3. These have been defined by DNase I protection of the region by the repressor (Krause et al., 1983(Krause et al., , 1986Alazard et al., 1992;Goosen et al., 1984). Binding of c to O1 and O2 appears to occur simultaneously at low repressor concentrations and prevents the initiation of transcription at the early promoter Pe. Binding to O3, which requires higher concentrations of c, inhibits transcription of the c gene from the Pc promoter (Goosen et al., 1984;Alazard et al., 1992). A binding site for integration host factor (IHF) is located between O1 and O2. This DNA-binding protein bends DNA and promotes the formation of looped structures (Craig and Nash, 1984;Gamas et al., 1987;Prentki et al., 1987;Stenzel et al., 1987;Robertson et al., 1988;Thompson et al., 1988). IHF plays a complex role in the regulation of the early functions of Mu. It has been demonstrated that it stimulates transcription from Pe, favoring lytic growth (Krause et al., 1986), possibly by direct interaction between bound IHF and RNA polymerase (Goosen and . On the other hand, IHF stabilizes the interaction among the repressor and O1 and O2 and allows the establishment and the maintenance of lysogeny (Vogel et al., 1991;Alazard et al., 1992;Gama et al., 1992;Bétermier et al., 1995).Previous results (Alazard et al., 1992) show that the interaction of c with a linear operator fragment leads to the formation of at least five high molecular weight DNA-protein complexes, which can be separated using gel retardation techniques. The complexes with lower apparent molecular weight can be chased successiv...

Oligomeric structure of the repressor of the bacteriophage Mu early operon

“…The results showed that, at low concentration, c first present the sequences at which stable binding of the repressor is binds to two of the 11-bp partially repeated sequences in O2 initiated [2].…”

“…These may represent core binding sites for Another protein, the integration host factor, plays a complex the repressor, which serve as an organiser of a more complex role in the regulation of the early functions of Mu. A specific nucleoprotein structure in which O2, O1 and O3 become succesbinding site for this host protein is located between O1 and O2.sively occupied as the concentration of repressor is increased Binding of the integration host factor to this site stimulates tran- [2]. A preliminary analysis of the properties of the repressor scription from Pe and, under certain conditions, from Pc [1, 3Ϫ…”

“…Transcription from Pe, located in O2, leads to the expression of early lytic functions including ner, the transposase by competing with the transposase N-terminal domain for its binding to IAS, thus impairing synapsis leading to the first pA, and an accessory protein pB which stimulates the transposition process. Transcription from Pc, which is located in O3, transposition intermediate [17, 14].In an attempt to determine the organisation of the complex leads to the synthesis of the repressor c. The operator sites O1, O2 and O3 contain 4, 3 and 2 imperfectly repeated, 11-bp se-nucleoprotein structure which is involved in repression of the early operator functions, we investigated the occupancy of the quences, respectively, which have been suggested to compose operator sites in several repressor-operator complexes isolated the minimal repressor-binding site [1] and demonstrated to rein vitro [2]. The results showed that, at low concentration, c first present the sequences at which stable binding of the repressor is binds to two of the 11-bp partially repeated sequences in O2 initiated [2].…”

“…In an attempt to determine the organisation of the complex leads to the synthesis of the repressor c. The operator sites O1, O2 and O3 contain 4, 3 and 2 imperfectly repeated, 11-bp se-nucleoprotein structure which is involved in repression of the early operator functions, we investigated the occupancy of the quences, respectively, which have been suggested to compose operator sites in several repressor-operator complexes isolated the minimal repressor-binding site [1] and demonstrated to rein vitro [2]. The results showed that, at low concentration, c first present the sequences at which stable binding of the repressor is binds to two of the 11-bp partially repeated sequences in O2 initiated [2].…”

“…sively occupied as the concentration of repressor is increased Binding of the integration host factor to this site stimulates tran- [2]. A preliminary analysis of the properties of the repressor scription from Pe and, under certain conditions, from Pc [1, 3Ϫ…”