Weaver/Molecular Biology

Coping with DNA Damage Without Repairing It

The direct reversal and excision repair mechanisms described so far are all true repair processes. They eliminate the defective DNA entirely. However, cells have other means of coping with damage that do not remove it but simply skirt around it. These are sometimes called repair mechanisms, even though they really are not. A better term might be damage tolerance mechanism.

Recombination Repair

Recombination repair is the most important of these mechanisms. It is also sometimes called postreplication repair because replication of a pyrimidine dimer can leave a gap opposite the dimer that must be repaired. Figure 10 shows how recombination repair works. First, the DNA is replicated. This creates a problem for DNA with pyrimidine dimers because the dimers stop the replication machinery. Nevertheless, after a pause, replication continues, leaving a gap (a daughter strand gap) across from the dimer. (A new primer is presumably required to restart DNA synthesis.) Next, recombination occurs between the gapped strand and its homologue on the other daughter DNA duplex. This recombination depends on the recA gene product, which exchanges the homologous DNA strands (chapter 22). The net effect of this recombination is to fill in the gap across from the pyrimidine dimer and to create a new gap in the other DNA duplex. However, since the other duplex has no dimer, the gap can easily be filled in by DNA polymerase and ligase. Note that the DNA damage still exists, but the cell has at least managed to replicate its DNA. Sooner or later, true DNA repair could presumably occur.

Error-Prone Bypass

So-called error-prone bypass is another way of dealing with damage without really repairing it. In E. coli, this pathway is induced by DNA damage, including ultraviolet damage, and depends on the product of the recA gene. We have encountered recA before -- in the preceding paragraph and in our discussion of the induction of a lambda prophage during the SOS response (chapter 8) -- and we will encounter it again in our consideration of recombination in chapter 22. Indeed, error-prone repair is also part of the SOS response. The chain of events seems to be as follows (Figure 11): Ultraviolet light or another mutagenic treatment somehow activates the RecA co-protease activity. This co-protease has several targets. One we have studied already is the lambda repressor, but its main target is the product of the lexA gene. This product, LexA, is a repressor for many genes, including repair genes; when it is stimulated by RecA co-protease to cleave itself, all these genes are induced.

Two of the newly induced genes are umuC and umuD, which make up a single operon (umuDC). The product of the umuD gene (UmuD) is clipped by a protease to form UmuD’, which associates with the umuC product, UmuC, to form a complex UmuD’₂C. This complex has DNA polymerase activity, so it is also referred to as DNA pol V. (DNA pol IV is the product of another SOS response gene, dinB.) Pol V can cause error-prone bypass of thymine dimers in vitro on its own, but pol III holoenzyme and RecA protein stimulate this process considerably. Such bypass involves replication of DNA across from the pyrimidine dimer even though correct "reading" of the defective strand is impossible. This avoids leaving a gap, but it usually puts the wrong bases into the new DNA strand (hence the name "error-prone"). When the DNA replicates again, these errors will be perpetuated.

Wild-type E. coli cells can tolerate as many as fifty pyrimidine dimers in their genome without ill effect because of their active repair mechanisms. Bacteria lacking one of the uvr genes cannot carry out excision repair, so their susceptibility to ultraviolet damage is greater. However, they are still somewhat resistant to DNA damage. On the other hand, double mutants in uvr and recA can perform neither excision repair nor recombination repair, and they are very sensitive to ultraviolet damage, perhaps because they have to rely on error-prone repair. Under these conditions, only one to two pyrimidine dimers per genome is a lethal dose.

Summary: Cells can employ nonrepair methods to circumvent DNA damage. One of these is recombination repair, in which the gapped DNA strand across from a damaged strand recombines with a normal strand in the other daughter DNA duplex after replication. This solves the gap problem but leaves the original damage unrepaired. Another mechanism to deal with DNA damage, at least in E. coli, is to induce the SOS response, which causes the DNA to replicate even though the damaged region cannot be read correctly. This results in errors in the newly made DNA, so the process is called error-prone bypass.

Error-Prone and Error-Free Bypass in Humans

Human cells also have an error-prone system to bypass pyrimidine dimers. It involves a distinct DNA polymerase known as DNA polymerase x (zeta) that inserts bases at random to get past the pyrimidine dimer. But human cells also have a relatively error-free bypass system that automatically inserts two AMPs into the DNA strand across from a pyrimidine dimer. Thus, even though the bases in the dimer cannot base-pair, this system is able to make the correct choice if both bases in the dimer are thymines -- which is usually the case. The DNA polymerase involved in this relatively error-free repair is DNA polymerase h (eta).

In 1999, Fumio Hanaoka and colleagues discovered that the defective gene in patients with the variant form of XP (XP-V) is the gene that codes for DNA polymerase h . Thus, these patients cannot carry out the comparatively error-free bypass of pyrimidine dimers catalyzed by DNA polymerase h , and must therefore rely on the error-prone bypass catalyzed by DNA polymerase x . This error-prone system introduces mutations during replication of pyrimidine dimers not removed by the excision repair system. However, because these patients have normal excision repair, few dimers are left for the error-prone system to deal with. This argument accounts for the relatively low sensitivity of XP-V cells to ultraviolet radiation.

Summary: Humans have a relatively error-free bypass system that inserts AMPs across from a pyrimidine dimer, thus replicating thymine dimers (but not dimers involving cytosines) correctly. This system uses DNA polymerase. When the gene for this enzyme is defective, DNA polymerase takes over. But this polymerase inserts random nucleotides across from a pyrimidine dimer, so it is error prone. These errors in correcting UV damage lead to a variant form of XP known as XP-V.

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