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Transcription cycle. Microcin j25. TRCF (mfd). GreA GreB. Rifampicin. Anti-  's. Activators. Transcription cycle. TRCF (mfd). Miller spread. Transcription Repair Coupling Factor (TRCF, mfd). !. Selby & Sancar (1993) Science 260 , 53. Mahdi et al. (2003) EMBO J . 22 , 724.

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Transcription cycle
Transcription cycle

Microcin j25

TRCF

(mfd)

GreA

GreB

Rifampicin

Anti-'s

Activators




Transcription cycle

Transcription Repair Coupling Factor (TRCF, mfd)

!

Selby & Sancar (1993) Science260, 53.


Transcription cycle

Mahdi et al. (2003) EMBO J. 22, 724.

Chambers et al. (2003) Nucleic Acids Res. 31, 6409.

Selby & Sancar (1993) Science260, 53.

Selby & Sancar (1995) J. Biol. Chem. 270, 4882.


Transcription cycle

Rescue

• Transcript cleavage (stimulated by GreA/GreB)

• Slide forward



Transcription cycle

RecG

Singleton et al. (2001) Cell 107, 79

TRCF

Park et al. (2002) Cell 109, 757


Transcription cycle

Alexandra Deaconescu

Collaborators

Anna L. Chambers, Abigail J. Smith, Nigel J. Savery (University of Bristol, Bristol, UK)

Bryce E. Nickels, Ann Hochschild (Harvard Medical School)


Transcription cycle

Data collection and model refinement statistics for apo Mfd

Experimental electrondensity map

(after solvent modification)

Deaconescu et al. (2006) Cell124, 507.




Transcription cycle

APS NE-CAT-8BM

APS SBC-19ID

Deaconescu et al. (2006) Cell124, 507.

R/Rfree = 0.25/0.29 (3.2 Å resolution)


Transcription cycle

Singleton et al. (2001) Cell 107, 79

Deaconescu et al. (2006) Cell124, 507.


Transcription cycle

Truglio et al. (2002) EMBO J. 23, 2498

UvrB homology module, UvrA recruitment, and the role of D7


Transcription cycle

• necessary for RNAP binding (Selby & Sancar, 1995)

• binds RNAP b subunit 19-142 (Park et al., 2002)


Transcription cycle

DNA-binding and ATPase activity of TRCF-L499R mutant intact

Deaconescu et al. (2006) Cell124, 507.



Transcription cycle

A. L. Chambers, A.J. Smith, N. J. Savery

B. Nickels, A. Hochschild

Leu499

Smith & Savery (2005) Nucleic Acids Res. 33, 755

Park et al. (2002) Cell 109, 757

Park et al. (2002) Cell 109, 757


Transcription cycle

Dürr et al. (2005) Cell 121, 363


Transcription cycle

• TRCF - compact arrangement of structural domains linked by long, flexible linkers - appears ‘primed’ for large scale conformational changes (translocation module, UvrA binding surface).

? Nature and timing of TRCF conformational changes during the steps of TRCF-mediated transcription-coupled repair reaction? (recognition of a stalled RNAP; forward translocation of the RNAP to the transcription block; RNAP release and transcript termination; recruitment of the NER machinery to the site)

? Details of protein/protein interactions involving TRCF? (TRCF/RNAP and TRCF/NER machinery)

? Termination mechanism?


Transcription cycle

Anna L. Chambers, Abigail J. Smith, Nigel J. Savery (University of Bristol)

Bryce E. Nickels, Ann Hochschild (Harvard Medical School)

Chris Roberts, Jeff Roberts (Cornell University)

Chris Selby, Aziz Sancar (University of North Carolina, Chapel Hill)

N. Sukumar , M. Capel, S. Ealick (APS NE-CAT-8BM)

A. Joachimiak, S. Grinnell (APS SBC-19ID)

Darst Lab

Matthew Bick

Elizabeth Campbell

Linda (Madge) Champagnie

Alexandra Deaconescu

Raji Edayathumangalam

Deepti Jain

Valerie Lamour

William Lane

Fred Leon

Natacha Opalka

Georgia Patikoglou

Sheng Wang

Lars Westblade

Kelly-Anne Wilson


Transcription cycle

Bacterial transcription termination (University of Bristol)

Figure 1

A. Intrinsic

terminator

B. Rho

C. Mfd


Transcription cycle

Roadblock repression assay (University of Bristol)

(Chambers et al., 2003, Nucleic Acids Res. 31, 6409)


Transcription cycle

DNA-binding activity of TRCF-L499R mutant intact (University of Bristol)

(also ATPase activity)


Transcription cycle

Epshtein & Nudler (2003) Cooperation between RNA polymerase molecules in transcription elongation, Science300, 801.