E3 ubiquitin-protein ligase RNF8 is an enzyme that in humans is encoded by the RNF8 gene. [5] [6] [7] RNF8 has activity both in immune system functions [8] and in DNA repair.
E3 ubiquitin-protein ligase RNF8 is an enzyme that in humans is encoded by the RNF8 gene. [5] [6] [7] RNF8 has activity both in immune system functions [8] and in DNA repair.
The protein encoded by this gene contains a RING finger motif and an FHA domain. This protein has been shown to interact with several class II ubiquitin-conjugating enzymes (E2), including UBE2E1/UBCH6, UBE2E2, and UBE2E3, and may act as a ubiquitin ligase (E3) in the ubiquitination of certain nuclear proteins. Alternatively spliced transcript variants encoding distinct isoforms have been reported. [7]
RNF8 promotes repair of DNA damage through three DNA repair pathways: homologous recombinational repair (HRR), [9] non-homologous end joining (NHEJ), [10] [11] and nucleotide excision repair (NER). [10] DNA damage is considered to be the primary cause of cancer, and deficiency in DNA repair can cause mutations leading to cancer. [12] A deficiency in RNF8 predisposes mice to cancer. [13] [14]
After the occurrence of a double-strand break in DNA, the chromatin needs to be relaxed to allow DNA repair, either by HRR or by NHEJ. There are two pathways that result in chromatin relaxation, one initiated by PARP1 and one initiated by γH2AX (the phosphorylated form of the H2AX protein) (see Chromatin remodeling). Chromatin remodeling initiated by γH2AX depends on RNF8, as described below.
The histone variant H2AX constitutes about 10% of the H2A histones in human chromatin. [15] At the site of a DNA double-strand break, the extent of chromatin with phosphorylated γH2AX is about two million base pairs. [15]
γH2AX does not, by itself, cause chromatin decondensation, but within seconds of irradiation the protein “Mediator of the DNA damage checkpoint 1” (MDC1) specifically attaches to γH2AX. [16] [17] This is accompanied by simultaneous accumulation of RNF8 protein and the DNA repair protein NBS1 which bind to MDC1. [18] RNF8 mediates extensive chromatin decondensation through its subsequent interaction with CHD4 protein, [19] a component of the nucleosome remodeling and deacetylase complex NuRD.
DNA end resection is a pivotal step in HRR repair that produces 3’ overhangs that provide a platform to recruit proteins involved in HRR repair. The MRN complex, consisting of Mre11, Rad50 and NBS1, carries out the initial steps of this end resection. [20] RNF8 ubiquitinates NBS1 (both before and after DNA damage occurs), and this ubiquitination is required for effective homologous recombinational repair. [9] Ubiquitination of NBS1 by RNF8 is, however, not required for the role of NBS1 in another DNA repair process, the error-prone microhomology-mediated end joining DNA repair. [9]
RNF8 appears to have other roles in HRR as well. RNF8, acting as a ubiquitin ligase, mono-ubiquitinates γH2AX to tether DNA repair molecules at DNA lesions. [21] In particular, RNF8 activity is required to recruit BRCA1 for homologous recombination repair. [22]
Ku protein is a dimeric protein complex, a heterodimer of two polypeptides, Ku70 and Ku80. Ku protein forms a ring structure. An early step in non-homologous end joining DNA repair of a double-strand break is the slipping of a Ku protein (with its ring protein structure) over each end of the broken DNA. The two Ku proteins, one on each broken end, bind to each other and form a bridge. [23] [24] This protects the DNA ends and forms a platform for further DNA repair enzymes to operate. After the broken ends are rejoined, the two Ku proteins still encircle the now intact DNA and can no longer slip off an end. The Ku proteins must be removed or they cause loss of cell viability. [25] The removal of Ku protein is performed either by RNF8 ubiquitination of Ku80, allowing it to be released from the Ku protein ring, [26] or else by NEDD8 promoted ubiquitination of Ku protein, causing its release from DNA. [25]
UV-induced formation of pyrimidine dimers in DNA can lead to cell death unless the lesions are repaired. Most repair of these lesions is by nucleotide excision repair. [27] After UV-irradiation, RNF8 is recruited to sites of UV-induced DNA damage and ubiquitinates chromatin component histone H2A. These responses provide partial protection against UV irradiation. [10] [28]
Spermatogenesis is the process in which spermatozoa are produced from spermatogonial stem cells by way of mitosis and meiosis. A major function of meiosis is homologous recombinational repair of this germline DNA.[ citation needed ] RNF8 plays an essential role in signaling the presence of DNA double-strand breaks. Male mice with a gene knockout for RNF8 have impaired spermatogenesis, apparently due to a defect in homologous recombinational repair. [13]
RNF8 has been shown to interact with Retinoid X receptor alpha. [29]
Chromatin is a complex of DNA and protein found in eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser structures. This prevents the strands from becoming tangled and also plays important roles in reinforcing the DNA during cell division, preventing DNA damage, and regulating gene expression and DNA replication. During mitosis and meiosis, chromatin facilitates proper segregation of the chromosomes in anaphase; the characteristic shapes of chromosomes visible during this stage are the result of DNA being coiled into highly condensed chromatin.
Ubiquitin is a small regulatory protein found in most tissues of eukaryotic organisms, i.e., it is found ubiquitously. It was discovered in 1975 by Gideon Goldstein and further characterized throughout the late 1970s and 1980s. Four genes in the human genome code for ubiquitin: UBB, UBC, UBA52 and RPS27A.
DNA repair is a collection of processes by which a cell identifies and corrects damage to the DNA molecules that encode its genome. In human cells, both normal metabolic activities and environmental factors such as radiation can cause DNA damage, resulting in tens of thousands of individual molecular lesions per cell per day. Many of these lesions cause structural damage to the DNA molecule and can alter or eliminate the cell's ability to transcribe the gene that the affected DNA encodes. Other lesions induce potentially harmful mutations in the cell's genome, which affect the survival of its daughter cells after it undergoes mitosis. As a consequence, the DNA repair process is constantly active as it responds to damage in the DNA structure. When normal repair processes fail, and when cellular apoptosis does not occur, irreparable DNA damage may occur, including double-strand breaks and DNA crosslinkages. This can eventually lead to malignant tumors, or cancer as per the two hit hypothesis.
Non-homologous end joining (NHEJ) is a pathway that repairs double-strand breaks in DNA. NHEJ is referred to as "non-homologous" because the break ends are directly ligated without the need for a homologous template, in contrast to homology directed repair(HDR), which requires a homologous sequence to guide repair. NHEJ is active in both non-dividing and proliferating cells, while HDR is not readily accessible in non-dividing cells. The term "non-homologous end joining" was coined in 1996 by Moore and Haber.
Homologous recombination is a type of genetic recombination in which genetic information is exchanged between two similar or identical molecules of double-stranded or single-stranded nucleic acids.
Mouse double minute 2 homolog (MDM2) also known as E3 ubiquitin-protein ligase Mdm2 is a protein that in humans is encoded by the MDM2 gene. Mdm2 is an important negative regulator of the p53 tumor suppressor. Mdm2 protein functions both as an E3 ubiquitin ligase that recognizes the N-terminal trans-activation domain (TAD) of the p53 tumor suppressor and as an inhibitor of p53 transcriptional activation.
Nibrin, also known as NBN or NBS1, is a protein which in humans is encoded by the NBN gene.
H2A histone family member X is a type of histone protein from the H2A family encoded by the H2AFX gene. An important phosphorylated form is γH2AX (S139), which forms when double-strand breaks appear.
Histone-modifying enzymes are enzymes involved in the modification of histone substrates after protein translation and affect cellular processes including gene expression. To safely store the eukaryotic genome, DNA is wrapped around four core histone proteins, which then join to form nucleosomes. These nucleosomes further fold together into highly condensed chromatin, which renders the organism's genetic material far less accessible to the factors required for gene transcription, DNA replication, recombination and repair. Subsequently, eukaryotic organisms have developed intricate mechanisms to overcome this repressive barrier imposed by the chromatin through histone modification, a type of post-translational modification which typically involves covalently attaching certain groups to histone residues. Once added to the histone, these groups elicit either a loose and open histone conformation, euchromatin, or a tight and closed histone conformation, heterochromatin. Euchromatin marks active transcription and gene expression, as the light packing of histones in this way allows entry for proteins involved in the transcription process. As such, the tightly packed heterochromatin marks the absence of current gene expression.
Chromatin remodeling is the dynamic modification of chromatin architecture to allow access of condensed genomic DNA to the regulatory transcription machinery proteins, and thereby control gene expression. Such remodeling is principally carried out by 1) covalent histone modifications by specific enzymes, e.g., histone acetyltransferases (HATs), deacetylases, methyltransferases, and kinases, and 2) ATP-dependent chromatin remodeling complexes which either move, eject or restructure nucleosomes. Besides actively regulating gene expression, dynamic remodeling of chromatin imparts an epigenetic regulatory role in several key biological processes, egg cells DNA replication and repair; apoptosis; chromosome segregation as well as development and pluripotency. Aberrations in chromatin remodeling proteins are found to be associated with human diseases, including cancer. Targeting chromatin remodeling pathways is currently evolving as a major therapeutic strategy in the treatment of several cancers.
NEDD8 is a protein that in humans is encoded by the NEDD8 gene. This ubiquitin-like (UBL) protein becomes covalently conjugated to a limited number of cellular proteins, in a process called NEDDylation similar to ubiquitination. Human NEDD8 shares 60% amino acid sequence identity to ubiquitin. The primary known substrates of NEDD8 modification are the cullin subunits of cullin-based E3 ubiquitin ligases, which are active only when NEDDylated. Their NEDDylation is critical for the recruitment of E2 to the ligase complex, thus facilitating ubiquitin conjugation. NEDD8 modification has therefore been implicated in cell cycle progression and cytoskeletal regulation.
Cullin-4A is a protein that in humans is encoded by the CUL4A gene. CUL4A belongs to the cullin family of ubiquitin ligase proteins and is highly homologous to the CUL4B protein. CUL4A regulates numerous key processes such as DNA repair, chromatin remodeling, spermatogenesis, haematopoiesis and the mitotic cell cycle. As a result, CUL4A has been implicated in several cancers and the pathogenesis of certain viruses including HIV. A component of a CUL4A complex, Cereblon, was discovered to be a major target of the teratogenic agent thalidomide.
DNA damage-binding protein 2 is a protein that in humans is encoded by the DDB2 gene.
Mediator of DNA damage checkpoint protein 1 is a 2080 amino acid long protein that in humans is encoded by the MDC1 gene located on the short arm (p) of chromosome 6. MDC1 protein is a regulator of the Intra-S phase and the G2/M cell cycle checkpoints and recruits repair proteins to the site of DNA damage. It is involved in determining cell survival fate in association with tumor suppressor protein p53. This protein also goes by the name Nuclear Factor with BRCT Domain 1 (NFBD1).
E3 ubiquitin-protein ligase RING2 is an enzyme that in humans is encoded by the RNF2 gene.
Ubiquitin-conjugating enzyme E2 D3 is a protein that in humans is encoded by the UBE2D3 gene.
Sirtuin 6 is a stress responsive protein deacetylase and mono-ADP ribosyltransferase enzyme encoded by the SIRT6 gene. In laboratory research, SIRT6 appears to function in multiple molecular pathways related to aging, including DNA repair, telomere maintenance, glycolysis and inflammation. SIRT6 is member of the mammalian sirtuin family of proteins, which are homologs to the yeast Sir2 protein.
Homology-directed repair (HDR) is a mechanism in cells to repair double-strand DNA lesions. The most common form of HDR is homologous recombination. The HDR mechanism can only be used by the cell when there is a homologous piece of DNA present in the nucleus, mostly in G2 and S phase of the cell cycle. Other examples of homology-directed repair include single-strand annealing and breakage-induced replication. When the homologous DNA is absent, another process called non-homologous end joining (NHEJ) takes place instead.
DNA damage is an alteration in the chemical structure of DNA, such as a break in a strand of DNA, a nucleobase missing from the backbone of DNA, or a chemically changed base such as 8-OHdG. DNA damage can occur naturally or via environmental factors, but is distinctly different from mutation, although both are types of error in DNA. DNA damage is an abnormal chemical structure in DNA, while a mutation is a change in the sequence of base pairs. DNA damages cause changes in the structure of the genetic material and prevents the replication mechanism from functioning and performing properly. The DNA damage response (DDR) is a complex signal transduction pathway which recognizes when DNA is damaged and initiates the cellular response to the damage.
The human KDM2B gene encodes the protein lysine (K)-specific demethylase 2B.
PDB gallery | |
|---|---|
|
