| content |
Contents |
Uvr Proteins
The process of nucleotide excision repair is controlled in E. coli by the UvrABC endonuclease enzyme complex, which consists of four Uvr proteins: UvrA, UvrB, UvrC, and DNA helicase II (sometimes also known as UvrD in this complex). First, a UvrA-UvrB complex scans the DNA, with the UvrA subunit recognizing distortions in the helix, caused for example by thymine dimers and other cyclobutyl dimers. When the complex recognizes such a distortion, UvrA exits and UvrB melts the base pairs between the two DNA strands. UvrB then recruits UvrC, which incises the DNA 8 nucleotides away from the distortion on the 5’ end and 4 nucleotides away on the 3’ end, creating a 12-nucleotide long single-stranded DNA that is excised with the help of the DNA helicase II. The resultant gap is then filled in using DNA polymerase I and DNA ligase. The basic excision process is very similar in higher cells, but these cells usually involve many more proteins – E.coli is a simple example.
Nucleotide Excision Repair in Eukaryotes
Nucleotide excision repair has more complexity in eukaryotes. But the general principles upon which it operates are similar. There are 9 major proteins involved in NER in mammalian cells and their names come from the diseases associated with the deficiencies in those proteins. XPA,XPB XPC,XPD,XPE,XPF, XPG all derive from Xeroderma pigmentosum and CSA and CSB represents proteins linked to Cockayne syndrome. Additionally, the proteins ERCCI, RPA, Rad23, and others also participate in nucleotide excision repair.
As described below, nucleotide excision repair can be categorized into two classes, global genome NER (GG-NER) and Transcription Coupled NER (TC-NER). Two different sets of proteins are involved in the distortion and recognition of the DNA damage in the two types of NER. In GG-NER, the XPC-Rad23B complex is responsible for distortion recognition, and DDB1 and DDB2 (XPE) can also recognize some types of damage caused by UV light. Additionally, XPA performs a function in damage recognition that is as yet poorly defined. In TC-NER, CS proteins CSA and CSB bind some types of DNA damage instead of XPC-Rad23B.
The subsequent steps in GG-NER and TC-NER are similar to each other and to those in NER in prokaryotes. XPB and XPD, which are subunits in transcription factor TFIIH have helicase activity and unwind the DNA at the sites of damages. XPG protein has a structure-specific endonuclease activity, which makes an incision 3’ to the damaged DNA. Subsequently XPF protein, which is associated with ERCC1 makes the 5' incision during the NER. The dual-incision leads to the removal of a ssDNA with a single strand gap of 25~30 nucleotides.
The resulting gap in DNA is filled by DNA pol δ or ε by copying the undamaged strand. Proliferating Cell Nuclear Antigen (PCNA) assists the DNA polymerase in the reaction and Replication Protein A (RPA) protects the other DNA strand from degradation during the NER. Finally, DNA ligase seals the nicks to finish NER.
Global genomic NER
Global genomic NER repairs damage in both transcribed and untranscribed DNA strands in active and inactive genes throughout the genome. This pathway employs several 'damage sensing' proteins including the DNA-damage binding (DDB) and XPC-Rad23B complexes that constantly scan the genome and recognize helix distortions. Upon identification of a damaged site, subsequent repair proteins are then recruited to the damaged DNA to verify presence of DNA damage, excise the damaged DNA surrounding the lesion then fill in the repair patch.
A 2002 paper published by Frit et al showed that Global Genome Repair can be enhanced near transcribed regions. Their main conclusion is that the binding of transcription factors led to an increase in nucleotide excision repair on nearby regions of DNA. They showed that this enhancement was distinct from Transcription Coupled Repair in two ways: Repair was occuring on both the template and non-template strands. Secondly, when Pol-II activity was blocked, either by α-amanitin or modification of the TATA box, the repair still occurred. Through digestions with micrococcal nuclease they determined that the binding of transcription factors led to local chromatin remodeling, likely giving NER proteins greater access to the DNA.
Transcription coupled repair
One of the interesting aspects of nucleotide excision repair is the transcription coupled repair mechanism. When RNA polymerase II is transcribing a gene and encounters a helix distorting thymine dimer, for example, it cannot continue transcription and stalls, recruiting nucleotide excision repair proteins to itself. This has two important consequences: (1) the RNA polymerase itself becomes a tool for recognizing DNA damage and (2) genes that are being actively transcribed are given more attention by the repair mechanisms. This is an especially important mechanism as persistently stalled RNA polymerase II results in either arrest of the cell cycle or pre-programmed cell death (apoptosis).
See also
External links
References
- DNA repair and Mutagenesis (2nd edition), Errol C.Friedberg et.al
- Frit et al. Transcriptional Activators Stimulate DNA Repair. Molecular Cell, Vol. 10, 1391–1401, December, 2002
