Scientists discover new mechanisms for repairing plant DNA damage

The study of the response mechanism of DNA damage is one of the most basic biological problems. Recently, the team of Yan Shunping, professor of the College of Life Science and Technology of Huazhong Agricultural University and Hubei Hongshan Laboratory, published a research paper in the international journal EMBO Journal. The paper found that the SMC5/6 complex mediates monoubiquitination of histone H2B at the DSB site by recruiting the PAF1 complex, thereby promoting homologous recombination repair. This not only revealed the new mechanism of SMC5/6 regulating DNA damage repair, but also found that PAF1C has a new function other than transcriptional regulation – DNA damage repair, which provides new genetic resources for the use of homologous recombinant repair mechanism to improve gene targeting efficiency.

The SMC5/6 complex recruits the PAF1 complex to promote DNA damage repair. Photo courtesy of Huazhong Agricultural University

DNA is the carrier of genetic information, and its accurate replication and transmission is the basis for the survival of organisms. However, many exogenous factors (e.g., ultraviolet rays, ionizing radiation, etc.) and endogenous factors (e.g., reactive oxygen species, replication errors, etc.) can cause DNA damage. If these damages cannot be repaired, it will affect the most basic biological processes such as replication and transcription, and then affect the growth and development of organisms and species reproduction.

DNA double-strand breaks (DSBs) are considered the most serious type of DNA damage. Organisms repair DNA double-strand breaks mainly through two pathways, namely non-homologous terminal ligation (NHEJ) and homologous recombination (HR). The repair pathway of non-homologous end connection is a relatively fast but error-prone process, while the repair pathway of homologous recombination is a relatively slow but precise repair pathway.

DNA double-strand break repair is one of the foundations of gene editing using technologies such as CRISPR. Gene knockout can be achieved by relying on non-homologous terminal linkage pathways, and gene knock-in can be achieved by relying on homologous recombination pathways. At present, one of the main bottlenecks of gene editing technology is the low efficiency of gene knock-in, which cannot meet the needs of basic research and biological breeding. Among them, one of the most important reasons is the low efficiency of homologous recombination of plants. Therefore, there is an urgent desire to improve the efficiency of gene knock-in by improving the efficiency of homologous recombination.

The chromosome structure maintenance complex SMC5/6 is highly conserved in eukaryotes and is necessary for homologous recombination repair, but its mechanism of regulating homologous recombination repair needs to be further studied. The PAF1 complex is also highly conserved in eukaryotes, and its main function is to regulate the transcription of genes. Previously, whether the PAF1 complex was involved in DNA damage repair was still a mystery.

In this study, the researchers used forward genetics to screen Arabidopsis DNA damage response mutants and found that ddrm4 mutants are very sensitive to DNA double-strand break induction reagents. DDRM4 encodes PAF1, the core subunit of the PAF1 complex. Through biochemistry, cell biology, genetics and other means, the researchers found that the SMC5/6 complex recruited the PAF1 complex to the DNA double-strand break site, and the PAF1 complex further recruited the E2 ubiquitin-binding enzyme UBC1/2 and E3 ubiquitin ligase HUB1/2 to mediate the single ubiquitination of the histone H2B at the DNA double-strand break site, thereby promoting homologous recombination repair.

Li Cunliang and Guo Yuyu, doctoral students in Yan Shunping’s group, are the co-first authors of the paper, Yan Shunping is the corresponding author, and researcher Wang Lili also participated in the study. The research was supported by the National Natural Science Foundation of China and the Huazhong Agricultural University-Shenzhen Institute of Agricultural Genomics, Chinese Academy of Agricultural Sciences.

It is worth mentioning that this is the third DDRM protein that the team recently discovered that regulates homologous recombinant repair. In previous studies, the team identified two plant-specific regulatory proteins, DDRM1 and DDRM2. As an E3 ubiquitin ligase, DDRM1 is a single ubiquitinated transcription factor SOG1, which improves the protein stability of SOG1; SOG1 promotes DDRM2 transcription; DDRM2 promotes the binding of the homologous recombinase RAD51 to the DSB site, thereby completing homologous recombinant repair. (Source: Li Chen, China Science News)

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