The tL2 cluster of transcription termination sites between genes bet and ral of coliphage lambda

Abstract

The major leftward transcription of bacteriophage λ is controlled by several terminators ( t), including t L1, t L2, t L3, and others. The t L2 termination site, which was placed by Salstrom and Szybalski ( Virology 88, 252–260, 1978) between λ genes bet and ral, was found to consist of a cluster of four leftward terminators. As not to change the numbering of other leftward terminators, these were designated as t L2a, t L2b, t L2c and t L2d As determined by S1 nuclease mapping of the t L2-terminated in vitro transcripts, the normally p L- initiated major leftward λ transcription should encounter termination points at 1653 bp ( t L2a; between genes Ea10 and ral), 2089 by ( t L2b; between genes cIII and Ea10), 2441–2442 bp, and 2483 by ( t L2c and t L2d; both within gene gam) from the S L startpoint (= +1). All terminators were cloned in a pBR322-derived plasmid between the p′ R promoter and the galK gene, and their in vivo termination efficiencies are 69% ( t L2a), 53% ( t L2b), and 38% ( t L2c + t L2a), measured as reduction of galK expression in rho + galK − hosts at 30° The t L2a and t L2b terminators depend little on the rho factor, whereas the efficiency of t L2c + t L2d decreases from 38 to only 14% in the rho − host. When shifted to 42°, the termination efficiency of t L2b decreases from 53 to only 36%, while the other t L2 terminators are much less affected by increasing the temperature. The calculated joint efficiency of the entire t L2 cluster is 90%, which is in perfect agreement with the 90% termination efficiency reported by Salstrom and Szybalski (1978) for t L2. However, the natural location of t L2c and t L2d within the actively translated gam gene may interfere with their termination function. Under in vitro conditions, t L2a and t L2b are active only in the presence of rho factor, whereas t L2a and t L2b do not require rho. The structure of the t L2 terminators resembles that of some other known termination sites: a perfect (8 by for t L2a and t L2a) or imperfect (8–9 bp) dyad symmetry and a T 6 ( t L2a), T 5 ( t L2b), T 4 ( t L2c), or TTATT sequence ( t L2d) toward the 3′ end of the mRNA-like DNA strand.