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Structure-specific endonucleases (SSEs) play key roles in DNA replication, recombination, and repair. SSEs must be tightly regulated to ensure genome stability but their regulatory mechanisms remain incompletely understood. Here, we show that in the fission yeast Schizosaccharomyces pombe, the activities of two SSEs, Dna2 and Rad16 (ortholog of human XPF), are temporally controlled during the cell cycle by the CRL4Cdt2 ubiquitin ligase. CRL4Cdt2 targets Pxd1, an inhibitor of Dna2 and an activator of Rad16, for degradation in S phase. The ubiquitination and degradation of Pxd1 is dependent on CRL4Cdt2, PCNA, and a PCNA-binding degron motif on Pxd1. CRL4Cdt2-mediated Pxd1 degradation prevents Pxd1 from interfering with the normal S-phase functions of Dna2. Moreover, Pxd1 degradation leads to a reduction of Rad16 nuclease activity in S phase, and restrains Rad16-mediated single-strand annealing, a hazardous pathway of repairing double-strand breaks. These results demonstrate a new role of the CRL4Cdt2 ubiquitin ligase in genome stability maintenance and shed new light on how SSE activities are regulated during the cell cycle., Author summary Structure-specific endonucleases are enzymes that process DNA intermediates generated in DNA replication, recombination, and repair. Proper regulation of these enzymes is critical for maintaining genome stability. Dna2 and XPF are two such enzymes present across eukaryotes, from yeasts to humans. Here, we show that in the fission yeast Schizosaccharomyces pombe, the activities of Dna2 and Rad16 (the equivalent of human XPF) are temporally controlled during the cell cycle by the CRL4Cdt2 ubiquitin E3 ligase. In the S phase of the cell cycle, CRL4Cdt2 promotes the degradation of Pxd1, which is an inhibitor of Dna2 and an activator of Rad16. Through targeting Pxd1 for degradation, CRL4Cdt2 increases the activity of Dna2 in S phase and is important for the normal S-phase function of Dna2. Meanwhile, the degradation of Pxd1 reduces the activity of Rad16 in S phase, and curtails Rad16-dependent single-strand annealing, a mutagenic DNA repair pathway. Our findings uncover a new mechanism regulating two important endonucleases during the cell cycle, and reveal a new way of coordinating endonucleases to safeguard genome stability.

As one of the most aggressive malignancies, cervical cancer (CC) which mainly affects females has high risks of relapse and death. Long non-coding RNA (lncRNA) RP11-552M11.4 is verified to promote the progression of ovarian cancer; nevertheless, its role and the probable molecular mechanisms in CC remain unclear until the present study. Herein, we unveiled that the expression of lncRNA RP11-552M11.4 tested by qRT-PCR was enhanced in tumor tissues compared with the para-carcinoma tissues and related to FIGO Stage, lymph node metastasis, vascular invasion and distant metastasis in CC. Additionally, CC patients with high lncRNA RP11-552M11.4 level suffered from poor clinical outcomes. Moreover, silenced lncRNA RP11-552M11.4 restrained cell proliferation, migration and invasion in CC cells. Subsequently, the mechanistic studies revealed that lncRNA RP11-552M11.4 functioned as a ceRNA of ATF1 in CC by acting as the endogenous sponge for miR-3941, which was identified as a tumor suppressor in CC. Moreover, both miR-3941 inhibition and ATF1 overexpression restored the impacts of inhibited lncRNA RP11-552M11.4 on cellular processes in CC cells. Our observations elucidated the carcinogenic role of lncRNA RP11-552M11.4 in CC was mediated through miR-3941/ATF1 axis, giving a new insight into the effective target for the treatment and prognosis of cervical cancer.

The human dynamin-like protein, HdynIV, has recently been cloned and shown to be involved in the formation and trafficking of coated vesicles. In particular, one of the HdynIV variant overexpressions has been suggested to contribute to the pathogenesis of brain tumors. In this paper, we report on the genomic organization of the human HdynIV gene. The gene was found to correspond to 20 exons of genomic sequence on human chromosome 12, distributed over 64 kb of genomic DNA. The two exons, numbers 15 and 16, are subjected to differential splicing, generating four different transcripts of a perfect match to our recent report on the four different spliced HdynIV variants [DNA Cell Biol. 19 (2000) 189]. We have also characterized the 5′ regulatory region of the HdynIV gene in order to understand the molecular mechanisms regulating its expression. The transcriptional initiation site was identified by 5′-RACE. The 5′-flanking sequence of the HdynIV gene contains three GC boxes that concatenate Ap2- and Sp1-binding motifs, but that does not contain either the TATA or CAAT consensus sequence. A region between −140 and +92 contributed to high promoter activity. Deletion analysis demonstrated that the minimal promoter activity required the region of −110 to −100. Electrophoretic mobility shift assay demonstrated that a putative transcriptional factor bound to the region of −119 to −90. Site-directed mutagenesis analysis of this region revealed that nucleotides at −108 to −100 were essential for transactivation mediated by this transcriptional factor. In conclusion, we have characterized the minimal HdynIV promoter and shown that CTCCCAGCA (−108 to −100) sequence may act as a novel transcriptional element for regulating HdynIV gene expression.

Dynamin-like protein, a large GTP-binding protein, has recently been cloned, and studies have suggested that it is involved in the formation of coated vesicles. In this report, the differential expression of four human dynamin-like protein splice variants (HdynIV-wildtype [WT], -11, -26, and -37) from various brain tumors was identified by reverse transcription/polymerase chain reaction (RT-PCR). One novel variant (HdynIV-11), not described previously, was identified. The four alternatively spliced variants exhibited tissue specificity in normal tissues. The HdynIV-WT was strongly expressed in the brain, whereas HdynIV-37 was expressed in all tissues examined. Moreover, HdynIV-26 was dominant in the liver and apparently overexpressed in all astrocytomas and most meningiomas and adenomas. This report suggests that HdynIV-26 may cause aberrant protein trafficking and alter vesicle formation in brain tumors. Our results also suggest that dynamin-like protein is associated with various brain tumors and, more importantly, that aberrant expression of the HdynIV-26 variant may play a role in brain tumorigenesis.