OZF is a Claspin-interacting protein essential to maintain the replication fork progression rate under replication stress.

DNA replication is essential for cell proliferation and is one of the cell cycle stages where DNA is more vulnerable. Replication stress is a prominent property of tumor cells and an emerging target for cancer therapy. Although it is not directly involved in nucleotide incorporation, Claspin is a protein with relevant functions in DNA replication. It harbors a DNA-binding domain that interacts preferentially with branched or forked DNA molecules. It also acts as a platform for the interaction of proteins related to DNA damage checkpoint activation, DNA repair, DNA replication origin firing, and fork progression. In order to find new proteins potentially involved in the regulation of DNA replication, we performed a two-hybrid screen to discover new Claspin-binding proteins. This system allowed us to identify the zinc-finger protein OZF (ZNF146) as a new Claspin-interacting protein. OZF is also present at replication forks and co-immunoprecipitates not only with Claspin but also with other replisome components. Interestingly, OZF depletion does not affect DNA replication in a normal cell cycle, but its depletion induces a reduction in the fork progression rate under replication stress conditions. Our results suggest that OZF is a Claspin-binding protein with a specific function in fork progression under replication stress.

CircPIP5K1A facilitates gastric cancer progression via miR-376c-3p/ZNF146 axis.

BACKGROUND: Recently, many emerging circular RNAs (circRNAs) have been studied in human malignancies, including gastric cancer (GC). Researches concerning cancers have revealed that aberrant expression of circRNAs play a big part in tumorigenesis and development of diverse malignant tumors. Although hsa_circ_0014130 (circPIP5K1A) has been confirmed to be closely related to non-small cell lung cancer (NSCLC) progression, the knowledge of its function on GC progression remains unclear. Therefore, it is of great interest to uncover the underlying role of circPIP5K1A in GC. METHODS: The expression and characteristic of circPIP5K1A were separately analyzed by RT-qPCR, nucleic acid electrophoresis, RNase R and Actinomycin D treatment. CCK-8, colony formation, EdU, transwell, TUNEL, flow cytometry, luciferase reporter, RIP and RNA pull-down assays were employed to testify the regulatory role of circPIP5K1A in GC. RESULTS: In current study, circPIP5K1A, featured with closed-loop structure, was proved to be highly expressed in tissues and cells of GC. Loss-of-function assays depicted that silencing circPIP5K1A suppressed GC development. Follow-up mechanism tests unveiled that circPIP5K1A bound with miR-376c-3p and inhibition of miR-376c-3p reversed circPIP5K1A downregulation-mediated effect on GC progression. Additionally, ZNF146 was verified to be the downstream molecule of circPIP5K1A/miR-376c-3p axis in modulating GC progression. CONCLUSIONS: circPIP5K1A stimulates GC progression by sponging miR-376c-3p to upregulate ZNF146 expression.

ZNF146/OZF and ZNF507 target LINE-1 sequences.

Repetitive sequences including transposable elements and transposon-derived fragments account for nearly half of the human genome. While transposition-competent transposable elements must be repressed to maintain genomic stability, mutated and fragmented transposable elements comprising the bulk of repetitive sequences can also contribute to regulation of host gene expression and broader genome organization. Here, we analyzed published ChIP-seq data sets to identify proteins broadly enriched on transposable elements in the human genome. We show 2 of the proteins identified, C2H2 zinc finger-containing proteins ZNF146 (also known as OZF) and ZNF507, are targeted to distinct sites within LINE-1 ORF2 at thousands of locations in the genome. ZNF146 binding sites are found at old and young LINE-1 elements. In contrast, ZNF507 preferentially binds at young LINE-1 sequences correlated to sequence changes in LINE-1 elements at ZNF507's binding site. To gain further insight into ZNF146 and ZNF507 function, we disrupt their expression in HEK293 cells using CRISPR/Cas9 and perform RNA sequencing, finding modest gene expression changes in cells where ZNF507 has been disrupted. We further identify a physical interaction between ZNF507 and PRMT5, suggesting ZNF507 may target arginine methylation activity to LINE-1 sequences.

Hsa_circ_NOTCH3 regulates ZNF146 through sponge adsorption of miR-875-5p to promote tumorigenesis of hepatocellular carcinoma.

BACKGROUND: To explore the specific mechanism of circular RNA (circRNA) in the occurrence and development of hepatocellular carcinoma (HCC), and provide new ideas for its diagnosis and treatment. METHODS: Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was for evaluating the expression of circ_NOTCH3 in liver cancer tissues and matched normal tissues and related cell lines. After overexpression or co-expression of circ_NOTCH3 or microRNA (miRNA) in cells, the changes in cell function were analyzed. Bioinformatics analysis and dual luciferase report analysis were utilized to predict and verify the binding site between circ_NOTCH3 and miRNA. Western blotting was applied to detect gene expression alterations. Additionally, in vivo tumor growth was also utilized to further assess the influence of knocking-down circ_NOTCH3 on the progression of HCC. RESULTS: It was confirmed circ_NOTCH3 was highly expressed in HCC specimens and cells. The proliferation, migration, invasion, and oxaliplatin-resistance potential of HCC could be restrained by silencing circ_NOTCH3 or by ectopic expression of miR-875-5p in vitro. In terms of mechanism, circ_NOTCH3 directly binds to miR-875-5p, regulating its activity by targeting the 3'-UTR of ZNF146. Overexpression of circ_NOTCH3 evidently overturned the diminishing influence of miR-875-5p mimics on HCC cells. CONCLUSIONS: As an oncogene, circ_NOTCH3 can trigger the proliferation, invasion, migration, and oxaliplatin resistance of HCC cells through the miR-875-5p/ZNF146 axis, and may be a promising target for the treatment of HCC.

LncRNA KCNQ1OT1 acts as miR-216b-5p sponge to promote colorectal cancer progression via up-regulating ZNF146.

Long non-coding RNAs (lncRNAs) have shown to act as important regulators in cancer biology. The aim of this study was to investigate the role and mechanism of lncRNA KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1) in colorectal cancer (CRC) progression. The abundance of KCNQ1OT1, microRNA-216b-5p (miR-216b-5p) and zinc finger protein 146 (ZNF146) messenger RNA (mRNA) was measured by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation was analyzed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and colony formation assay. Cell migration and invasion abilities were assessed by transwell assays. Western blot assay was performed for determination of protein levels. LncBase v.2 of DIANA Tool and StarBase software were used to predict the targets of KCNQ1OT1 and miR-216b-5p, respectively. Dual-luciferase reporter assay was implemented to confirm the target interaction between miR-216b-5p and KCNQ1OT1 or ZNF146. KCNQ1OT1 expression was higher in CRC tissues and cell lines. KCNQ1OT1 interference restrained the proliferation, migration and invasion of CRC cells. MiR-216b-5p was a target of KCNQ1OT1 in CRC cells, and KCNQ1OT1 knockdown-induced effects in CRC cells were partly overturned by miR-216b-5p silencing. MiR-216b-5p bound to the 3' untranslated region (3'UTR) of ZNF146, and ZNF146 overexpression partly attenuated miR-216b-5p overexpression-mediated influences in CRC cells. KCNQ1OT1 up-regulated the abundance of ZNF146 through sequestering miR-216b-5p in CRC cells. KCNQ1OT1 accelerated the proliferation and motility of CRC cells through elevating ZNF146 expression via sponging miR-216b-5p. KCNQ1OT1/miR-216b-5p/ZNF146 axis might be underlying target for the diagnosis and treatment of CRC patients.