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PubMed2026

Dual Roles of RAD23b and RAD4 on the Desiccation Tolerance of Germinated Seeds.

Desiccation tolerance (DT) is a survival trait enabling orthodox seeds to withstand extremely low water content. While some protective factors are characterised, it remains mechanistically obscure. Here, based on PEG-induced DT re-establishment in germinated Brassica napus L. seeds, we investigated the dual functions of nucleotide excision repair (NER) components RAD23b and RAD4 in DNA repair and transcriptional regulation of root development. PEG pre-treatment alleviated dehydration-induced DNA damage and activated NER genes, suggesting the involvement of NER in seed DT. Unexpectedly, Arabidopsis atrad23b mutant and BnRAD23b/BnRAD4 over-expressing seeds all exhibited significantly decreased DT after dry back, which evoked a hypothesis that BnRAD23b-BnRAD4 functions beyond NER. Normally, BnRAD4 interacted with BnRAD23b and repressed the expression of root development genes NAC103, EMB1444, RRA1 by directly binding to STRE elements within their promoters. Dehydration stress alleviated this repression, drove transcriptional reprogramming and might redirect the complex to execute DNA repair. Genetic analyses revealed that germinated seeds of atnac103, atemb1444, and atrra1 single mutants all exhibited reduced DT, and double mutants under atrad23b background almost abolished DT. This study suggests that RAD23b and RAD4 may regulate DT re-establishment of germinated seeds through balancing genome integrity and radicle development, with implications for DT study broadly.

Brassica napus L.↗

PubMed2024

Comparative Studies on Bulky DNA Damage Binding by Nucleotide Excision Repair Proteins Using Surface Plasmon Resonance, Differential Scanning Fluorometry, and DNase I Footprinting.

Nucleotide excision repair is a crucial cellular mechanism that ensures genomic stability, thereby preventing mutations that can lead to cancer. The human XPC and its yeast ortholog Rad4 protein complexes are central to this process and were the focus of the study. We used surface plasmon resonance and differential scanning fluorimetry to study the binding characteristics of XPC and Rad4 when bound to the bulky cluster di-FAAF-containing 55-mer duplex DNA. Our findings revealed that XPC binds 10 times more significant affinity to control and di-FAAF-modified DNA than Rad4 with greater protein-DNA interactions. Differential scanning fluorimetry indicates that Rad4 causes comparatively more significant conformational changes upon complexation with the damaged DNA. We conducted DNase I footprinting of the Rad4/DNA complex for the first time by determining the regions protected from DNase I digestion. The DNA at the lesion is entirely resistant to digestion by DNase I in the absence of Rad4 several nucleotides to the 3'-side of the first FAAF lesion. The lack of DNase I cleavage at the lesions did not change upon adding Rad4. However, in the presence of Rad4, a footprint is observed on the 7-nucleotide region (5'-TGGTGAT-3') of the complementary strand to the 3' side of the lesion.

Surface Plasmon Resonance↗

PubMed1978

Responses of radiation-sensitive mutants of Saccharomyces cerevisiae to lethal effects of bleomycin.

Haploid and diploid strains of yeast containing genes conferring radiation-sensitivity were studied under growing and nongrowing experimental conditions for their relative sensitivities to growth-inhibitory effects of bleomycin (BM). The rad1, rad2, rad3, rad4, rad5 (and allelic rev2), rad7, rad10, rad11, rad 12, rad14, rad15, rad16 and rev3 strains exhibited responses similar to normal (Rad+) yeast strains. It is concluded from these findings that the excision-repair function deficient in several of these mutant strains is not important for repair of bleomycin-induced damages in yeast. The sensitive strains contained rad6, rad9, rad18, rad22, rad50, rad51, rad52, rad53, rad54, rad55, rad56, rad57 and rs1. Strains bearing rad8 or rad19 could not be classified unambiguously. With one exception, all rad mutants found very sensitive to BM were sensitive to X-rays, suggesting that some aspect of the repair of BM- and X-ray-induced damages in yeast may be similar. Sensitivities to BM and radiation co-segregated in pedigrees following meiosis, and several BM-resistant revertants isolated from two rad6 mutant strains sensitive to BM, X-rays and UV were cross-resistant to all three agents. These results confirm that the rad mutants were responsible for the cross-sensitivities in the original strains.

Bleomycin↗

PubMed1977

Isolation and characterization of MMS-sensitive mutants of Saccharomyces cerevisiae.

We have isolated mutants sensitive to methyl methanesulfonate (MMS) in Saccharomyces cerevisiae. Alleles of rad1, rad4, rad52, rad55 and rad57 were found amoung these mms mutants. Twenty-nine of the mms mutants which complement the existing radiation-sensitive (rad and rev) mutants belong to 22 new complementation groups. Mutants from five complementation groups are sensitive only to MMS. Mutants of 11 complementation groups are sensitive to UV or X rays in addition to MMS, mutants of six complementation groups are sensitive to all three agents. The cross-sensitivities of these mms mutants to UV and X rays are discussed in terms of their possible involvement in DNA repair. Sporulation is reduced or absent in homozygous diploids of mms mutants from nine complementation groups.

Alleles↗