No abstract

The translational roles of the Shine-Dalgarno sequence, the initiation codon, the space between them, and the second codon have been studied. The Shine-Dalgarno sequence UAAGGAGG initiated translation roughly four times more efficiently than did the shorter AAGGA sequence. Each Shine-Dalgarno sequence required a minimum distance to the initiation codon in order to drive translation; spacing, however, could be rather long. Initiation at AUG was more efficient than at GUG or UUG at each spacing examined; initiation at GUG was only slightly better than UUG. Translation was also affected by residues 3' to the initiation codon. The second codon can influence the rate of initiation, with the magnitude depending on the initiation codon. The data are consistent with a simple kinetic model in which a variety of rate constants contribute to the process of translation initiation.

No abstract

Mutant ribosome binding sites of the bacteriophage T4 rIIB gene, resident on an 873 bp DNA fragment, were cloned into a plasmid vector as in-frame fusions to a reporter gene, beta-galactosidase. The collection of mutations included changes in the region 5' to the Shine/Dalgarno sequence, a mutation of the Shine/Dalgarno sequence, the alternate initiation codons GUG, AUA and ACG, and mutants in which several closely spaced initiation codons compete with each other on the same mRNA. The results show that the secondary structure variations we have installed 5' to the Shine/Dalgarno sequence have little effect on translation. GUG is essentially as good an initiator of translation as AUG when they are assayed on separate messages, but is outcompeted at least 50-fold in the sequence AUGUG. AUA and ACG are poor start codons, and are temperature sensitive. The initiation codon pair AUGAUA, in which the AUG is only two nucleotides from the Shine/Dalgarno sequence, displays a novel cold-sensitive phenotype.

The bacteriophage T4 rII genes and the lambda rex (r exclusion) genes interact; rll mutants are unable to productively infect rex+ lambda lysogens. The relationship between rex and rll has been found to be quantitative, and plasmid clones of rex have excluded not only rII mutants but T4 wild type and most other bacteriophages as well. Mutations in the T4 motA gene substantially reversed exclusion of T4 by rex.

No abstract

We cloned the intact T4 rHB gene by joining plasmids carrying gene fragments. rIlB was expressed at a low level under control of the lac promoter, and the clone complemented rIIB mutants. We suspect that earlier attempts to clone the intact gene were unsuccessful because of transcription from T4 middle-mode promoters. These promoters are silent early in infection but are recognized when resident on a plasmid in an uninfected cell.

Electron-dense particles 80-100 nm in diameter were observed in electron micrographs of mitochondria in transformed mouse fibroblasts and in a human amnion cell line infected with supernatants of a human mammary carcinoma cell culture. Intramitochondrial particles were more numerous in the human cell cultures early after subculture. These particles were morphologically similar to those reported in mitochondria of Rous sarcoma virus-infected cells.