A team led by a Baylor University researcher has published a groundbreaking article that provides a better understanding of the dynamic process by which sunlight-induced DNA damage from the molecular repair machinery in cells is recognized as being in need of repair.
Sun ultraviolet light is a ubiquitous carcinogen that can cause structural damage to cellular DNA. Since DNA contains important cell function blueprints, failure to remove and restore damaged portions of DNA can be detrimental and lead to skin cancer in humans, said lead author Jung-Hyun Min, Ph.D., associate professor of chemistry and biochemistry at Baylor's College of Arts & Sciences.
Min and her team demonstrated how the repair protein Rad4 / XPC would bind to such a UV-induced DNA damage 6-4 photoproduct to mark the damaged site along the DNA in preparation for the rest of the NER. Process (nucleotide excision repair) in cells.
The study – "Structure and Mechanism of Detection of Pyrimidine-Pyrimidone (6-4) Photoproducts by the Rad4 / XPC-NER Complex" – is published in the journal Nucleic Acids Research (NAR ) as a "breakthrough article".
Breakthrough articles present meaningful studies that answer longstanding questions in the field of nucleic acid research and / or open up new reasons and mechanistic hypotheses for investigation. These are the best-of-breed releases published in the NAR, accounting for 1 to 2 percent of reviews received by the journal.
UV light threatens the integrity of the genome by causing cellular DNA damage known as intra-strand crosslink damage, Min said. Two major types of these lesions are cyclobutane-pyrimidine dimer (CPD), which accounts for approximately 70% Percent of this damage. and 6-4 photoproduct (6-4PP), which accounts for about 30 percent.
The cellular DNA repair system (NER), which is responsible for the elimination of these lesions, is much faster in 6-4PP than CPD, Min said. This is because a DNA damage-sensing protein (Rad4 / XPC), initiates the NER, recognizes 6-4PP more efficiently than CPD.
Once a lesion is bound by Rad4 / XPC, it can be removed by the NER pathway. NER works in all organisms from yeast to man. It remains unclear how the Rad4 / XPC protein recognizes the lesions and what causes the differences in recognition efficiency, Min said.
The team first determined a 3-D structure of the Rad4 protein attached to a DNA substrate which contains a 6-4PP lesion using a technique called X-ray crystallography. The structure showed that the proteins rotate the parts of the DNA that contain the 6-4PP outward, "opening" the DNA double helix. This was accompanied by a strong twisting and bending of the DNA strands.
However, it was not the damaged part of the DNA that the protein contacted directly, Min said.
Instead, the protein bound specifically to the healthy parts of the DNA opposite the lesion. This shows that the protein could in principle bind to both the CPD and other environmental DNA lesions known to be recognized by Rad4 / XPC. However, it could not be directly explained why the detection efficiency between the lesions may be different.
To remedy this, Min then collaborated with Suse Broyde, Ph.D., at New York University, using molecular dynamics to simulate the process, with Rad4 initially binding to DNA with 6-4PP or CPD can.
The simulation studies showed that the protein readily engages with 6-4PP to loosen, bend, and partially "open" the DNA at the lesion site. Remarkably, the CPD-containing DNA resisted twisting and bending 6-4PP.
Overall, the team succeeded in creating a three-dimensional molecular trajectory that maps the key steps during the DNA "opening." was performed by Rad4 / XPC and revealed the reasons for the different recognition of 6-4PP and CPD.
Min believes that the discovery of these mechanisms could offer advantages to nucleotide excision repair that go beyond understanding UV-induced damage, as NER also plays a role. Important repair pathway for much of the environmental DNA damage – including caused by industrial pollutants, cigarette smoke and even some chemotherapeutic agents.
"The hallmark of NER is that it repairs a very wide range of DNA damage, which is quite important in terms of how our genomes are protected against environmental DNA damage," Min said.
"During It has been known for many decades that this Rad4 / XPC protein can recognize 6-4PP very efficiently. y, there is no structure that shows how it really binds to the lesion and why its recognition is so efficient compared to lesions like CPD is, "she said. "Basically, our study fills this gap well and tells us what this mechanism must be."
While this research has shown how Rad4 / XPC can bind to damage in a DNA duplex, it is still unknown how the protein can find such a damage if it is DNA that is tightly organized, such as it is the case in cells (called chromatin).
Min said most of the DNAs in chromatin are spun around proteins called histones, and how Rad4 / XPC can get around to find a lesion is another puzzle
She also said it was not known how Rad4 / XPC would recruit the next player of the repair pathway, the transcription factor II-H complex (TFIIH), which is important for checking the damage before other proteins arrive and in fact the damaged part fails.
"We hope the knowledge uncovered can be helpful in solving serious health problems," said Min. "So we imagine we can help – understanding how to do things with full 3D Structure details work. "
A new mechanism for accessing damaged DNA
Debamita Paul et al. Structure and Mechanism of the Detection of Pyrimidine-Pyrimidone (6-4) Photoproducts by the Rad4 / XPC Complex for the Repair of Nucleotide Excisions, Nucleic Acids Research (2019). DOI: 10.1093 / nar / gkz359
Study provides insight into sun-induced DNA damage and cell repair (2019, 14 July)
retrieved on 15 July 2019
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