Sequence-dependent kinetic model for transcription elongation by RNA polymerase.

We present a kinetic model for the sequence-dependent motion of RNA polymerase (RNAP) during transcription elongation. For each NTP incorporation, RNAP has a net forward translocation of one base-pair along the DNA template. However, this process may involve the exploration of back-tracked and forward-tracked translocation modes. In our model, the kinetic rates for the reaction pathway, calculated based on the stabilities of the transcription elongation complex (TEC), necessarily lead to sequence-dependent NTP incorporation rates. Simulated RNAP elongation kinetics is in good agreement with data from transcription gels and single-molecule studies. The model provides a kinetic explanation for well-known back-tracked pauses at transcript positions with unstable TECs. It also predicts a new type of pause caused by an energetically unfavorable transition from pre to post-translocation modes.

[1]  R. Sousa,et al.  T7 RNA polymerase elongation complex structure and movement. , 2000, Journal of molecular biology.

[2]  N. Sugimoto,et al.  Thermodynamic parameters to predict stability of RNA/DNA hybrid duplexes. , 1995, Biochemistry.

[3]  Thomas A Steitz,et al.  The Structural Mechanism of Translocation and Helicase Activity in T7 RNA Polymerase , 2004, Cell.

[4]  J. SantaLucia,et al.  Thermodynamic parameters for DNA sequences with dangling ends. , 2000, Nucleic acids research.

[5]  P. V. von Hippel,et al.  A thermodynamic analysis of RNA transcript elongation and termination in Escherichia coli. , 1991, Biochemistry.

[6]  Nancy R. Forde,et al.  Using mechanical force to probe the mechanism of pausing and arrest during continuous elongation by Escherichia coli RNA polymerase , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[7]  Jeffrey W. Roberts,et al.  Mechanism of intrinsic transcription termination and antitermination. , 1999, Science.

[8]  Shigeyuki Yokoyama,et al.  Structural Basis for Substrate Selection by T7 RNA Polymerase , 2004, Cell.

[9]  S. Benkovic,et al.  Elementary steps in the DNA polymerase I reaction pathway. , 1983, Biochemistry.

[10]  K. Johnson,et al.  Conformational coupling in DNA polymerase fidelity. , 1993, Annual review of biochemistry.

[11]  Elio A. Abbondanzieri,et al.  Ubiquitous Transcriptional Pausing Is Independent of RNA Polymerase Backtracking , 2003, Cell.

[12]  S. Darst,et al.  A Structural Model of Transcription Elongation , 2000 .

[13]  E. Nudler,et al.  Spatial organization of transcription elongation complex in Escherichia coli. , 1998, Science.

[14]  J. Roberts,et al.  RNA polymerase pausing and transcript release at the lambda tR1 terminator in vitro. , 1983, The Journal of biological chemistry.

[15]  Deletion analysis of the lambda tR1 termination region. Effect of sequences near the transcript release sites, and the minimum length of rho-dependent transcripts. , 1994, Journal of molecular biology.

[16]  D. Turner,et al.  Predicting thermodynamic properties of RNA. , 1995, Methods in enzymology.

[17]  D. Court,et al.  The relationship between function and DNA sequence in an intercistronic regulatory region in phage λ , 1978, Nature.

[18]  P. V. von Hippel,et al.  Reaction pathways in transcript elongation. , 2002, Biophysical chemistry.

[19]  J. SantaLucia,et al.  Improved nearest-neighbor parameters for predicting DNA duplex stability. , 1996, Biochemistry.

[20]  E. Geiduschek,et al.  Analysis of RNA chain elongation and termination by Saccharomyces cerevisiae RNA polymerase III. , 1994, Journal of molecular biology.

[21]  Michelle D. Wang,et al.  Force and velocity measured for single molecules of RNA polymerase. , 1998, Science.

[22]  T. Elston,et al.  Force generation in RNA polymerase. , 1998, Biophysical journal.

[23]  E. Nudler,et al.  The RNA–DNA Hybrid Maintains the Register of Transcription by Preventing Backtracking of RNA Polymerase , 1997, Cell.

[24]  M. Kashlev,et al.  RNA Polymerase Switches between Inactivated and Activated States By Translocating Back and Forth along the DNA and the RNA* , 1997, The Journal of Biological Chemistry.

[25]  William H. Press,et al.  Numerical recipes in C , 2002 .

[26]  F. Jülicher,et al.  Motion of RNA polymerase along DNA: a stochastic model. , 1998, Biophysical journal.

[27]  M. Kashlev,et al.  Transcriptional arrest: Escherichia coli RNA polymerase translocates backward, leaving the 3' end of the RNA intact and extruded. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[28]  Smita S. Patel,et al.  Pre-steady-state kinetic analysis of processive DNA replication including complete characterization of an exonuclease-deficient mutant. , 1991, Biochemistry.

[29]  K. Severinov,et al.  Crystal Structure of Thermus aquaticus Core RNA Polymerase at 3.3 Å Resolution , 1999, Cell.

[30]  D. Erie,et al.  Allosteric Binding of Nucleoside Triphosphates to RNA Polymerase Regulates Transcription Elongation , 2001, Cell.

[31]  Hiroshi Handa,et al.  NTP-driven Translocation by Human RNA Polymerase II* , 2003, The Journal of Biological Chemistry.

[32]  P. V. Hippel,et al.  An Integrated Model of the Transcription Complex in Elongation, Termination, and Editing , 1998 .

[33]  R. Sousa,et al.  A model for the mechanism of polymerase translocation. , 1997, Journal of molecular biology.

[34]  M. Chamberlin,et al.  Ribonucleic acid chain elongation by Escherichia coli ribonucleic acid polymerase. I. Isolation of ternary complexes and the kinetics of elongation. , 1974, The Journal of biological chemistry.

[35]  R. Landick,et al.  Pausing by bacterial RNA polymerase is mediated by mechanistically distinct classes of signals. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[36]  Michelle D. Wang,et al.  Single molecule analysis of RNA polymerase elongation reveals uniform kinetic behavior , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[37]  P. V. von Hippel,et al.  Multiple RNA polymerase conformations and GreA: control of the fidelity of transcription. , 1993, Science.

[38]  E. Nudler,et al.  The mechanism of intrinsic transcription termination. , 1999, Molecular cell.