9. A connection between transcription and repair

20.9 A connection between transcription and repair

In both bacteria and eukaryotes, there is a direct link from RNA polymerase to the activation of repair. The basic phenomenon was first observed because transcribed genes are preferentially repaired. Then it was discovered that it is only the template strand of DNA that is the target Xthe nontemplate strand is repaired at the same rate as bulk DNA.

In bacteria, the repair activity is provided by the uvr excision-repair system (see 14 Recombination and repair). Preferential repair is abolished by mutations in the gene mfd, whose product provides the link from RNA polymerase to the Uvr enzymes (for review see 224).

Figure 20.14 Mfd recognizes a stalled RNA polymerase and directs DNA repair to the damaged template strand.

Figure 20.14 shows a model for the link between transcription and repair. When RNA polymerase encounters DNA damage in the template strand, it stalls because it cannot use the damaged sequences as a template to direct complementary base pairing. This explains the specificity of the effect for the template strand (damage in the nontemplate strand does not impede progress of the RNA polymerase).

Figure 14.28 The Uvr system operates in stages in which UvrAB recognizes damage, UvrBC nicks the DNA, and UvrD unwinds the marked region.
Animated figure

The Mfd protein has two roles. First, it displaces the ternary complex of RNA polymerase from DNA. Second, it causes the UvrABC enzyme to bind to the damaged DNA. This leads to repair of DNA by the excision-repair mechanism (see Figure 14.28). After the DNA has been repaired, the next RNA polymerase to traverse the gene is able to produce a normal transcript (661).

A similar mechanism, although relying on different components, is used in eukaryotes. The template strand of a transcribed gene is preferentially repaired following UV-induced damage. The general transcription factor TFIIH is involved. TFIIH is found in alternative forms, which consist of a core associated with other subunits.

TFIIH has a common function in both initiating transcription and repairing damage. The same helicase subunit (XPD) creates the initial transcription bubble and melts DNA at a damaged site. Its other functions differ between transcription and repair, as provided by the appropriate form of the complex.

Figure 20.15 The TFIIH core may associate with a kinase at initiation and associate with a repair complex when damaged DNA is encountered.

Figure 20.15 shows that the basic factor involved in transcription consists of a core (of 5 subunits) associated with other subunits that have a kinase activity; this complex also includes a repair subunit. The kinase catalytic subunit that phosphorylates the CTD of RNA polymerase belongs to a group of kinases that are involved in cell cycle control (see 27 Cell cycle and growth regulation). It is possible that this connection influences transcription in response to the stage of the cell cycle.

Figure 14.37 A helicase unwinds DNA at a damaged site, endonucleases cut on either side of the lesion, and new DNA is synthesized to replace the excised stretch.
Animated figure

The alternative complex consists of the core associated with a large group of proteins that are coded by repair genes. These include a subunit (XPC) that recognizes damaged DNA, which provides the coupling function that enables a template strand to be preferentially repaired when RNA polymerase becomes stalled at damaged DNA. Other proteins associated with the complex include endonucleases (XPG, XPF, ERCC1). Homologous proteins are found in the complexes in yeast (where they are often identified by rad mutations that are defective in repair) and in man (where they are identified by mutations that cause diseases resulting from deficiencies in repairing damaged DNA) (662, 663). (Subunits with the name XP are coded by genes in which mutations cause the disease xeroderma pigmentosum (see 14.18 Eukaryotic repair systems). The basic model for repair is animated in Figure 14.37.

The kinase complex and the repair complex can associate and dissociate reversibly from the core TFIIH. This suggests a model in which the first form of TFIIH is required for initiation, but may be replaced by the other form (perhaps in response to encountering DNA damage). TFIIH dissociates from RNA polymerase at an early stage of elongation (after transcription of ~50 bp); its reassociation at a site of damaged DNA may require additional coupling components.

The repair function may require modification or degradation of RNA polymerase. The large subunit of RNA polymerase is degraded when the enzyme stalls at sites of UV damage. This process is deficient in cells from patients with Cockayne’s syndrome (a repair disorder). Cockayne’s syndrome is caused by mutations in either of two genes (CSA and CSB), both of whose products appear to be part of or bound to TFIIH. We do not yet understand the connection between the transcription/repair apparatus as such and the degradation of RNA polymerase. It is possible that removal of the polymerase is necessary once it has become stalled (664).

Reviews
224:	Selby, C. P. and Sancar, A. (1994). Mechanisms of transcription-repair coupling and mutation frequency decline. Microbiol. Rev. 58, 317-329.

Research
661:	Selby, C. P. and Sancar, A. (1993). Molecular mechanism of transcription-repair coupling. Science 260, 53-58.
662:	Schaeffer, L. et al. (1993). DNA repair helicase: a component of BTF2 (TFIIH) basic transcription factor. Science 260, 58-63.
663:	Svejstrup, J. Q. et al. (1995). Different forms of TFIIH for transcription and DNA repair: holo-TFIIH and a nucleotide excision repairosome. Cell 80, 21-28.
664:	Bregman, D. et al. (1996). UV-induced ubiquitination of RNA polymerase II: a novel modification deficient in Cockayne syndrome cells. Proc. Nat. Acad. Sci. USA 93, 11586-11590.