Complex formed by complementary RNA stem-loops and its stabilization by a protein: Function of ColE1 Rom protein

A small plasmid-specified RNA (RNA I) inhibits formation of the RNA primer for CoIE1 DNA replication by binding to its precursor (RNA II). Binding is modulated by the plasmid-specified Rom protein. Both in the presence and absence of Rom, binding starts with interaction between loops of RNAs. To understand the mechanism of binding, we examined the interactions of pairs of single stem-loops that are complementary fragments of RNA I and RNA II. We found that these complementary single stem-loops bind to each other at their loops, forming an RNAase V1-sensitive structure. Rom protects the complex from cleavage and from alkylation of phosphate groups by ethyinitrosourea. A single dimer of Rom binds to the complex by recognizing the structure rather than its exact nucleotide sequence. Rom enhances complex formation by decreasing the rate of dissociation of the complex. Structures of RNA complexes formed in the presence and absence of Rom are proposed.

[1]  J. Tomizawa.  Control of cole 1 plasmid replication: The process of binding of RNA I to the primer transcript , 1984, Cell.

[2]  B. Polisky,et al.  Suppression of ColE1 RNA-RNA mismatch mutations in vivo by the ColE1 Rop protein. , 1987, Plasmid.

[3]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[4]  G. Cesareni,et al.  Interaction between RNA1 and the primer precursor in the regulation of Co1E1 replication. , 1983, Journal of molecular biology.

[5]  L. Castagnoli,et al.  Control of initiation of pMB1 replication: Purified rop protein and RNA I affect primer formation in vitro , 1984, Cell.

[6]  B. Polisky,et al.  Alternative conformations of the ColE1 replication primer modulate its interaction with RNA I , 1985, Cell.

[7]  O. Uhlenbeck,et al.  Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA templates. , 1987, Nucleic acids research.

[8]  W. Gilbert,et al.  Chemical probes for higher-order structure in RNA. , 1980, Proceedings of the National Academy of Sciences of the United States of America.

[9]  D. Crothers,et al.  Studies of the complex between transfer RNAs with complementary anticodons. I. Origins of enhanced affinity between complementary triplets. , 1976, Journal of molecular biology.

[10]  G. Cesareni,et al.  Base pairing of RNA I with its complementary sequence in the primer precursor inhibits ColE1 replication , 1981, Nature.

[11]  J. Tomizawa.  Control of ColE1 plasmid replication: Binding of RNA I to RNA II and inhibition of primer formation , 1986, Cell.

[12]  H. Masukata,et al.  Factor-independent termination of transcription in a stretch of deoxyadenosine residues in the template DNA , 1987, Cell.

[13]  H. Masukata,et al.  Control of primer formation for ColE1 plasmid replication: Conformational change of the primer transcript , 1986, Cell.

[14]  M. Helmer-Citterich,et al.  Control of ColE1 replication: low affinity specific binding of Rop (Rom) to RNAI and RNAII. , 1988, The EMBO journal.

[15]  A. Oka,et al.  The structure of a transcriptional unit on colicin E1 plasmid. , 1979, European journal of biochemistry.

[16]  M Kokkinidis,et al.  Structure of the ColE1 rop protein at 1.7 A resolution. , 1987, Journal of molecular biology.

[17]  T. Itoh,et al.  Plasmid ColE1 incompatibility determined by interaction of RNA I with primer transcript. , 1981, Proceedings of the National Academy of Sciences of the United States of America.

[18]  J. Tomizawa.  Control of cole1 plasmid replication: Initial interaction of RNA I and the primer transcript is reversible , 1985, Cell.

[19]  S. Dasgupta,et al.  Multiple mechanisms for initiation of ColE1 DNA replication: DNA synthesis in the presence and absence of ribonuclease H , 1987, Cell.

[20]  L. Castagnoli,et al.  Genetic and structural analysis of the ColE1 Rop (Rom) protein. , 1989, The EMBO journal.

[21]  V. Vlassov,et al.  Tertiary structure of tRNAs in solution monitored by phosphodiester modification with ethylnitrosourea. , 1981, European journal of biochemistry.

[22]  G. Cesareni,et al.  Control of ColE1 DNA replication: the rop gene product negatively affects transcription from the replication primer promoter. , 1982, Proceedings of the National Academy of Sciences of the United States of America.

[23]  S. Dasgupta,et al.  Transcriptional activation of ColE1 DNA synthesis by displacement of the nontranscribed strand , 1987, Cell.

[24]  J. Tomizawa,et al.  Control of cole 1 plasmid replication: Enhancement of binding of RNA I to the primer transcript by the rom protein , 1984, Cell.

[25]  G. Cesareni,et al.  Regulation of plasmid copy number by complementary RNAs , 1985 .

[26]  J. Tomizawa.  Control of ColE1 plasmid replication. Intermediates in the binding of RNA I and RNA II. , 1990, Journal of molecular biology.