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. 1984 Aug 10;12(15):5979–5993. doi: 10.1093/nar/12.15.5979

The bacteriophage T4 regA gene: primary sequence of a translational repressor.

M Trojanowska, E S Miller, J Karam, G Stormo, L Gold
PMCID: PMC320051  PMID: 6473098

Abstract

The regA gene product of bacteriophage T4 is an autogenously controlled translational regulatory protein that plays a role in differential inhibition (translational repression) of a subpopulation of T4-encoded "early" mRNA species. The structural gene for this polypeptide maps within a cluster of phage DNA replication genes, (genes 45-44-62-regA-43-42), all but one of which (gene 43) are under regA-mediated translational control. We have cloned the T4 regA gene, determined its nucleotide sequence, and identified the amino-terminal residues of a plasmid-encoded, hyperproduced regA protein. The results suggest that the T4 regA gene product is a 122 amino acid polypeptide that is mildly basic and hydrophilic in character; these features are consistent with known properties of regA protein derived from T4-infected cells. Computer-assisted analyses of the nucleotide sequences of the regA gene and its three upstream neighbors (genes 45, 44, and 62) suggest the existence of three translational initiation units in this four-gene cluster; one for gene 45, one for genes 44, 62 and regA, and one that serves only the regA gene. The analyses also suggest that the gene 44-62 translational unit harbors a stable RNA structure that obligates translational coupling of these two genes.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aksoy S., Squires C. L., Squires C. Translational coupling of the trpB and trpA genes in the Escherichia coli tryptophan operon. J Bacteriol. 1984 Feb;157(2):363–367. doi: 10.1128/jb.157.2.363-367.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Alberts B. M., Frey L. T4 bacteriophage gene 32: a structural protein in the replication and recombination of DNA. Nature. 1970 Sep 26;227(5265):1313–1318. doi: 10.1038/2271313a0. [DOI] [PubMed] [Google Scholar]
  3. Baughman G., Nomura M. Localization of the target site for translational regulation of the L11 operon and direct evidence for translational coupling in Escherichia coli. Cell. 1983 Oct;34(3):979–988. doi: 10.1016/0092-8674(83)90555-x. [DOI] [PubMed] [Google Scholar]
  4. Cardillo T. S., Landry E. F., Wiberg J. S. regA protein of bacteriophage T4D: identification, schedule of synthesis, and autogenous regulation. J Virol. 1979 Dec;32(3):905–916. doi: 10.1128/jvi.32.3.905-916.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Carey J., Cameron V., de Haseth P. L., Uhlenbeck O. C. Sequence-specific interaction of R17 coat protein with its ribonucleic acid binding site. Biochemistry. 1983 May 24;22(11):2601–2610. doi: 10.1021/bi00280a002. [DOI] [PubMed] [Google Scholar]
  6. Doherty D. H., Gauss P., Gold L. On the role of the single-stranded DNA binding protein of bacteriophage T4 in DNA metabolism. I. Isolation and genetic characterization of new mutations in gene 32 of bacteriophage T4. Mol Gen Genet. 1982;188(1):77–90. doi: 10.1007/BF00332998. [DOI] [PubMed] [Google Scholar]
  7. Doolittle R. F. Similar amino acid sequences: chance or common ancestry? Science. 1981 Oct 9;214(4517):149–159. doi: 10.1126/science.7280687. [DOI] [PubMed] [Google Scholar]
  8. Edman P., Begg G. A protein sequenator. Eur J Biochem. 1967 Mar;1(1):80–91. doi: 10.1007/978-3-662-25813-2_14. [DOI] [PubMed] [Google Scholar]
  9. Frederick R. J., Snyder L. [Regulation of anti-late RNA synthesis in bacteriophage T4: a delayed early control]. J Mol Biol. 1977 Aug 25;114(4):461–476. doi: 10.1016/0022-2836(77)90172-3. [DOI] [PubMed] [Google Scholar]
  10. Gold L., O'Farrell P. Z., Russel M. Regulation of gene 32 expression during bacteriophage T4 infection of Escherichia coli. J Biol Chem. 1976 Nov 25;251(22):7251–7262. [PubMed] [Google Scholar]
  11. Huberman J. A., Kornberg A., Alberts B. M. Stimulation of T4 bacteriophage DNA polymerase by the protein product of T4 gene 32. J Mol Biol. 1971 Nov 28;62(1):39–52. doi: 10.1016/0022-2836(71)90129-x. [DOI] [PubMed] [Google Scholar]
  12. Itakura K., Hirose T., Crea R., Riggs A. D., Heyneker H. L., Bolivar F., Boyer H. W. Expression in Escherichia coli of a chemically synthesized gene for the hormone somatostatin. Science. 1977 Dec 9;198(4321):1056–1063. doi: 10.1126/science.412251. [DOI] [PubMed] [Google Scholar]
  13. Johnsen M., Christensen T., Dennis P. P., Fiil N. P. Autogenous control: ribosomal protein L10-L12 complex binds to the leader sequence of its mRNA. EMBO J. 1982;1(8):999–1004. doi: 10.1002/j.1460-2075.1982.tb01284.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Karam J. D., Bowles M. G. Mutation to overproduction of bacteriophage T4 gene products. J Virol. 1974 Feb;13(2):428–438. doi: 10.1128/jvi.13.2.428-438.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Karam J., Bowles M., Leach M. Expression of bacteriophage T4 genes 45, 44, and 62. I. Discoordinate synthesis of the T4 45- and 44-proteins. Virology. 1979 Apr 15;94(1):192–203. doi: 10.1016/0042-6822(79)90449-5. [DOI] [PubMed] [Google Scholar]
  16. Karam J., Gold L., Singer B. S., Dawson M. Translational regulation: identification of the site on bacteriophage T4 rIIB mRNA recognized by the regA gene function. Proc Natl Acad Sci U S A. 1981 Aug;78(8):4669–4673. doi: 10.1073/pnas.78.8.4669. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Karam J., McCulley C., Leach M. Genetic control of mRNA decay in T4 phage-infected Escherichia coli. Virology. 1977 Feb;76(2):685–700. doi: 10.1016/0042-6822(77)90251-3. [DOI] [PubMed] [Google Scholar]
  18. Kozak M., Nathans D. Translation of the genome of a ribonucleic acid bacteriophage. Bacteriol Rev. 1972 Mar;36(1):109–134. doi: 10.1128/br.36.1.109-134.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kyte J., Doolittle R. F. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. doi: 10.1016/0022-2836(82)90515-0. [DOI] [PubMed] [Google Scholar]
  20. Lemaire G., Gold L., Yarus M. Autogenous translational repression of bacteriophage T4 gene 32 expression in vitro. J Mol Biol. 1978 Nov 25;126(1):73–90. doi: 10.1016/0022-2836(78)90280-2. [DOI] [PubMed] [Google Scholar]
  21. Mathews C. K., North T. W., Prem veer Reddy G. Multienzyme complexes in DNA precursor biosynthesis. Adv Enzyme Regul. 1978;17:133–156. doi: 10.1016/0065-2571(79)90011-6. [DOI] [PubMed] [Google Scholar]
  22. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  23. Merle P., Kadenbach B. The subunit composition of mammalian cytochrome c oxidase. Eur J Biochem. 1980 Apr;105(3):499–507. doi: 10.1111/j.1432-1033.1980.tb04525.x. [DOI] [PubMed] [Google Scholar]
  24. Meyer F., Weber H., Weissmann C. Interactions of Q beta replicase with Q beta RNA. J Mol Biol. 1981 Dec 15;153(3):631–660. doi: 10.1016/0022-2836(81)90411-3. [DOI] [PubMed] [Google Scholar]
  25. Model P., McGill C., Mazur B., Fulford W. D. The replication of bacteriophage f1: gene V protein regulates the synthesis of gene II protein. Cell. 1982 Jun;29(2):329–335. doi: 10.1016/0092-8674(82)90149-0. [DOI] [PubMed] [Google Scholar]
  26. Napoli C., Gold L., Singer B. S. Translational reinitiation in the rIIB cistron of bacteriophage T4. J Mol Biol. 1981 Jul 5;149(3):433–449. doi: 10.1016/0022-2836(81)90480-0. [DOI] [PubMed] [Google Scholar]
  27. Nossal N. G. DNA replication with bacteriophage T4 proteins. Purification of the proteins encoded by T4 genes 41, 45, 44, and 62 using a complementation assay. J Biol Chem. 1979 Jul 10;254(13):6026–6031. [PubMed] [Google Scholar]
  28. Oppenheim D. S., Yanofsky C. Translational coupling during expression of the tryptophan operon of Escherichia coli. Genetics. 1980 Aug;95(4):785–795. doi: 10.1093/genetics/95.4.785. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Piperno J. R., Alberts B. M. An ATP stimulation of T4 DNA polymerase mediated via T4 gene 44/62 and 45 proteins. The requirement for ATP hydrolysis. J Biol Chem. 1978 Jul 25;253(14):5174–5179. [PubMed] [Google Scholar]
  30. Russel M., Gold L., Morrissett H., O'Farrell P. Z. Translational, autogenous regulation of gene 32 expression during bacteriophage T4 infection. J Biol Chem. 1976 Nov 25;251(22):7263–7270. [PubMed] [Google Scholar]
  31. Sacerdot C., Fayat G., Dessen P., Springer M., Plumbridge J. A., Grunberg-Manago M., Blanquet S. Sequence of a 1.26-kb DNA fragment containing the structural gene for E.coli initiation factor IF3: presence of an AUU initiator codon. EMBO J. 1982;1(3):311–315. doi: 10.1002/j.1460-2075.1982.tb01166.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Schneider T. D., Stormo G. D., Haemer J. S., Gold L. A design for computer nucleic-acid-sequence storage, retrieval, and manipulation. Nucleic Acids Res. 1982 May 11;10(9):3013–3024. doi: 10.1093/nar/10.9.3013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Schneider T. D., Stormo G. D., Yarus M. A., Gold L. Delila system tools. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):129–140. doi: 10.1093/nar/12.1part1.129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Spahr P. F., Farber M., Gesteland R. F. Binding site on R17 RNA for coat protein. Nature. 1969 May 3;222(5192):455–458. doi: 10.1038/222455a0. [DOI] [PubMed] [Google Scholar]
  35. Steege D. A. 5'-Terminal nucleotide sequence of Escherichia coli lactose repressor mRNA: features of translational initiation and reinitiation sites. Proc Natl Acad Sci U S A. 1977 Oct;74(10):4163–4167. doi: 10.1073/pnas.74.10.4163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Stormo G. D., Schneider T. D., Gold L., Ehrenfeucht A. Use of the 'Perceptron' algorithm to distinguish translational initiation sites in E. coli. Nucleic Acids Res. 1982 May 11;10(9):2997–3011. doi: 10.1093/nar/10.9.2997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Tinoco I., Jr, Borer P. N., Dengler B., Levin M. D., Uhlenbeck O. C., Crothers D. M., Bralla J. Improved estimation of secondary structure in ribonucleic acids. Nat New Biol. 1973 Nov 14;246(150):40–41. doi: 10.1038/newbio246040a0. [DOI] [PubMed] [Google Scholar]
  38. Tomizawa J. I., Anraku N., Iwama Y. Molecular mechanisms of genetic recombination in bacteriophage. VI. A mutant defective in the joining of DNA molecules. J Mol Biol. 1966 Nov 14;21(2):247–253. doi: 10.1016/0022-2836(66)90095-7. [DOI] [PubMed] [Google Scholar]
  39. Trimble R. B., Maley Level of specific prereplicative mRNA's during bacteriophage T4 regA-, 43- and T4 43- infection of Escherichia coli B. J Virol. 1976 Feb;17(2):538–549. doi: 10.1128/jvi.17.2.538-549.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  41. Weber H., Billeter M. A., Kahane S., Weissmann C., Hindley J., Porter A. Molecular basis for repressor activity of Q replicase. Nat New Biol. 1972 Jun 7;237(75):166–170. doi: 10.1038/newbio237166a0. [DOI] [PubMed] [Google Scholar]
  42. Wiberg J. S., Mendelsohn S., Warner V., Hercules K., Aldrich C., Munro J. L. SP62, a viable mutant of bacteriophage T4D defective in regulation of phage enzyme synthesis. J Virol. 1973 Oct;12(4):775–792. doi: 10.1128/jvi.12.4.775-792.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Wilson G. G., Murray N. E. Molecular cloning of the DNA ligase gene from bacteriophage T4. I. Characterisation of the recombinants. J Mol Biol. 1979 Aug 15;132(3):471–491. doi: 10.1016/0022-2836(79)90270-5. [DOI] [PubMed] [Google Scholar]
  44. Yen T. S., Webster R. E. Translational control of bacteriophage f1 gene II and gene X proteins by gene V protein. Cell. 1982 Jun;29(2):337–345. doi: 10.1016/0092-8674(82)90150-7. [DOI] [PubMed] [Google Scholar]

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