ZmRAD17 Is Required for Accurate Double-Strand Break Repair During Maize Male Meiosis

RAD17, a replication factor C (RFC)-like DNA damage sensor protein, is involved in DNA checkpoint control and required for both meiosis and mitosis in yeast and mammals. In plant, the meiotic function of RAD17 was only reported in rice so far. Here, we identified and characterized the RAD17 homolog in maize. The Zmrad17 mutants exhibited normal vegetative growth but male was partially sterile. In Zmrad17 pollen mother cells, non-homologous chromosome entanglement and chromosome fragmentation were frequently observed. Immunofluorescence analysis manifested that DSB formation occurred as normal and the loading pattern of RAD51 signals was similar to wild-type at the early stage of prophase I in the mutants. The localization of the axial element ASY1 was normal, while the assembly of the central element ZYP1 was severely disrupted in Zmrad17 meiocytes. Surprisingly, no obvious defect in female sterility was observed in Zmrad17 mutants. Taken together, our results suggest that ZmRAD17 is involved in DSB repair likely by promoting synaptonemal complex assembly in maize male meiosis. These phenomena highlight a high extent of divergence from its counterpart in rice, indicating that the RAD17 dysfunction can result in a drastic dissimilarity in meiotic outcome in different plant species.

Exploration of Prognostic Biomarkers among Replication Factor C Family in the Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is one of the common cancers with a high incidence and mortality. The human replication factor C (RFC) family contains 5 subunits that play an important role in DNA replication and DNA damage repair. RFCs are abnormally expressed in a variety of cancers; some of them are differentially expressed in HCC tissues and related to tumor growth. However, the expression, prognostic value, and effect targets of the whole RFC family in HCC are still unclear. To address these issues, we performed a multidimensional analysis of RFCs in HCC patients by Oncomine, UALCAN, GEPIA, Human protein atlas, Kaplan-Meier plotter, cBioPortal, GeneMANIA, String, and LinkedOmics. mRNA expression of RFCs was significantly increased in HCC tissues. There was a significant correlation between the expression of RFC2/3/4/5 and tumor stage of HCC patients. Besides, high mRNA expression of RFC2/4 was associated with worse overall survival (OS). Moreover, genetic alterations of RFCs were associated with worse OS in HCC patients. We found that genes co-expressed with RFC2/4 were mainly involved in biological processes, such as chromosome segregation, mitotic cell cycle phase transition, and telomere organization and they activated the cell cycle and spliceosome pathways. The gene set is mainly enriched in cancer-related kinases AURKA, ATR, CDK1, PLK1, and CHEK1. E2F family members were the key transcription factors for RFCs. Our results suggest that differentially expressed RFC2 and RFC4 are potential prognostic biomarkers in HCC and may act on E2F transcription factors and some kinase targets to dysregulate the cell cycle pathway. These efforts may provide new research directions for prognostic biomarkers and therapeutic targets in HCC.

BTF3 confers oncogenic activity in prostate cancer through transcriptional upregulation of Replication Factor C

High levels of Basic Transcription Factor 3 (BTF3) have been associated with prostate cancer. However, the mechanisms underlying the role of BTF3 as an oncogenic transcription factor in prostate tumorigenesis have not been explored. Herein, we report that BTF3 confers oncogenic activity in prostate cancer cells. Mechanistically, while both BTF3 splicing isoforms (BTF3a and BTF3b) promote cell growth, BTF3b, but not BTF3a, regulates the transcriptional expression of the genes encoding the subunits of Replication Factor C (RFC) family that is involved in DNA replication and damage repair processes. BTF3 knockdown results in decreased expression of RFC genes, and consequently attenuated DNA replication, deficient DNA damage repair, and increased G2/M arrest. Furthermore, knockdown of the RFC3 subunit diminishes the growth advantage and DNA damage repair capability conferred by ectopic overexpression of BTF3b. Importantly, we show that enforced BTF3 overexpression in prostate cancer cells induces substantial accumulation of cisplatin-DNA adducts and render the cells more sensitive to cisplatin treatment both in vitro and in vivo. These findings provide novel insights into the role of BTF3 as an oncogenic transcription factor in prostate cancer and suggest that BTF3 expression levels may serve as a potential biomarker to predict cisplatin treatment response.

CANVAS: a late onset ataxia due to biallelic intronic AAGGG expansions

The ataxias are a group of disorders that manifest with balance, movement, speech and visual problems. They can arise due to dysfunction of the cerebellum, the vestibular system and/or the sensory neurons. Genetic defects are a common cause of chronic ataxia, particularly common are repeat expansions in this group of conditions. Co-occurrence of cerebellar ataxia with neuropathy and vestibular areflexia syndrome has been termed CANVAS. Although CANVAS is a rare syndrome, on discovery of biallelic expansions in the second intron of replication factor C subunit 1 (RFC1) gene, we and others have found the phenotype is broad and RFC1 expansions are a common cause of late-onset progressive ataxia. We aim to provide a review and update on recent developments in CANVAS and populations, where the disorder has been reported. We have also optimised a protocol for RFC1 expansion screening which is described herein and expanded phenotype after analysing late-onset ataxia patients from around the world.

Up-regulated RFC2 Predicts Unfavorably Progression in Hepatocellular Carcinoma

Background: Replication factor C (RFC) is closely related to tumor progression and metastasis. However, the functional significance of RFC2 in hepatocellular carcinoma remains unclear.Materials and methods: In order to solve this problem, the expression of RFC2 in liver cancer patients was analyzed through ONCOMINE, UALCAN, human protein atlas. Survival analysis was conducted using Kaplan-Meier plotter and GEPIA. GO and KEGG enrichment analyses were carried out. The protein-protein interaction (PPI) network was performed through Metascape.Result: The transcription and protein level of RFC2 in HCC were overexpressed, which was significantly related to the clinical individual cancer stage and pathological tumor grade of HCC patients. In addition, in patients with liver cancer, higher RFC2 expression was found to be significantly correlated with shorter OS and DFS. Furthermore, the function of RFC2 in liver cancer was DNA replication, and its main mechanism was the phase transition of the cell cycle.Conclusion: RFC2 might promote the development of liver cancer. It could also be used as a novel biomarker for the prognosis of liver cancer.

ATAD5 deficiency alters DNA damage metabolism and sensitizes cells to PARP inhibition

Replication factor C (RFC), a heteropentamer of RFC1-5, loads PCNA onto DNA during replication and repair. Once DNA synthesis has ceased, PCNA must be unloaded. Recent findings assign the uloader role primarily to an RFC-like (RLC) complex, in which the largest RFC subunit, RFC1, has been replaced with ATAD5 (ELG1 in Saccharomyces cerevisiae). ATAD5-RLC appears to be indispensable, given that Atad5 knock-out leads to embryonic lethality. In order to learn how the retention of PCNA on DNA might interfere with normal DNA metabolism, we studied the response of ATAD5-depleted cells to several genotoxic agents. We show that ATAD5 deficiency leads to hypersensitivity to methyl methanesulphonate (MMS), camptothecin (CPT) and mitomycin C (MMC), agents that hinder the progression of replication forks. We further show that ATAD5-depleted cells are sensitive to poly(ADP)ribose polymerase (PARP) inhibitors and that the processing of spontaneous oxidative DNA damage contributes towards this sensitivity. We posit that PCNA molecules trapped on DNA interfere with the correct metabolism of arrested replication forks, phenotype reminiscent of defective homologous recombination (HR). As Atad5 heterozygous mice are cancer-prone and as ATAD5 mutations have been identified in breast and endometrial cancers, our finding may open a path towards the therapy of these tumours.