Chemotherapy failure is the major cause of recurrence and poor prognosis in colorectal cancer (CRC) patients. The role of the differentially expressed lncRNAs in 5-Fluorouracil chemoresistance has not fully explained. Here, we observed lncRNA H19 was associated with the 5-Fu resistance in CRC. Quantitative analysis indicated that H19 was significantly increased in recurrent CRC patient samples. Kaplan–Meier survival analysis indicated that high H19 expression in CRC tissues was significantly associated with poor recurrent free survival. Our functional studies demonstrated that H19 promoted colorectal cells 5-Fu resistance. Mechanistically, H19 triggered autophagy via SIRT1 to induce cancer chemoresistance. Furthermore, bioinformatics analysis showed that miR-194–5p could directly bind to H19, suggesting H19 might work as a ceRNA to sponge miR-194–5p, which was confirmed by Dual-luciferase reporter assay and Immunoprecipitation assay. Extensively, our study also showed that SIRT1 is the novel direct target of miR-194–5p in CRC cells. Taken together, our study suggests that H19 mediates 5-Fu resistance in CRC via SIRT1 mediated autophagy. Our finding provides a novel mechanistic role of H19 in CRC chemoresistance, suggesting that H19 may function as a marker for prediction of chemotherapeutic response to 5-Fu.

DNAThe DNAs used for these studies were the following oligonucleotides (Integrated DNA Technologies) of: a 45-mer lagging-strand oligo, (5ʹ-GGCAGGCAGGCAGGCACACACTCTCCAATTA/iBiodT/CACTTCCTACTCTA-3ʹ) and a 70-mer leading-strand oligo (5ʹ-TAGAGTAGGAAGTGA/iBiodT/AATTGGAGAGTGTGTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT*T*T*T*T*T-3ʹ). The asterisks are residues containing a phosphothio linkage. The two oligos were annealed in equimolar amounts by heating to 90°C followed by slow (1 hour) cooling to room temperature. The hybrid was purified from a 8% native PAGE then a 1 molar equivalent of streptavidin was added, enabling a single streptavidin to cross-link the two diametrically opposed biotins. ProteinsMcm10 and CMG were purified as previously described (Langston et al., 2017), with the following modifications. The Mcm10 contained a N-terminal hexahistidine tag and a C-terminal 3X FLAG tag. Briefly, 48 L E. coli cells carrying the Mcm10 T7 based E. coli expression vector were grown to OD 0.6 at 37 o C, then cooled to 15 o C and induced upon adding IPTG for an additional 8 h at 15 o C. The cells were harvested by slow speed centrifugation and lysed using a continuous flow high speed homogenizer. Cell debris was removed by centrifugation and applied to a 10 ml Chelating Sepharose Fast Flow column (GE Healthcare) charged with 50 mM NiSO 4 in Buffer A (20 mM Tris-Cl pH 7.9, 5 mM imidazole, 500 mM NaCl, 0.01% NP-40). The column was washed with Buffer A, then eluted with 375 mM imidazole in Buffer A. The eluated material was applied to a 6 ml anti-FLAG M2 affinity gel (Sigma) equilibrated in Buffer B (20 mM Tris-Cl pH 7.5, 10% glycerol, 500 mM NaCl, 1 mM DTT, 1 mM MgCl 2 , 0.01% NP-40) and then washed with 20 column volumes of Buffer B before eluting with Buffer B containing 0.2 mg/ml FLAG peptide (EZ Biolab, Carmel, Indiana USA) using two 6 ml pulses of 20 min each and collecting 1.5 ml fractions. Eluted fractions were analyzed by SDS-PAGE, protein concentration was determined using Bradford Protein Stain (Sigma) wtih BSA as a standard. Proteins were then aliquoted, snap frozen in liquid nitrogen and stored at -80 o C.The CMG-Mcm10 complex was reconstituted by mixing 765 pmol CMG with 3.1 nmol Mcm10 in 0.7 ml Buffer C (10 mM Tris-Cl pH 7.5, 200 mM KCl, 2mM DTT, 2 mM MgCl 2 ) for 30 minutes on ice. The mixture was then applied to a 0.1 ml MonoQ column, equilibrated in Buffer C. The column was washed using the same buffer and eluted with a 2.5 ml gradient of Buffer C from 0.2 M KCl to 0.6 M KCl. Fractions of 0.1 ml were collected, analysed by Bradford and SDS-PAGE, pooled and dialyzed against 50 mM K-glutamate in 25 mM Tris-acetate pH 7.5, 2 mM Mg acetate, and 1 mM DTT. Dialysed material was analyzed again by Bradford stain for protein concentation , then aliquoted, snap frozen in liquid nitrogen and stored at -80 o C.

Sustainable water-splitting hydrogen production has long been considered one of the most promising energy conversion technologies, but enormous challenges remain: for instance, water electrolysis suffers from high overpotential and over energy consumption under neutral pH conditions. Here, taking advantage of the memory effect of layered double hydroxide (LDH), we report an energy-efficient neutral water electrolyzer material based on LDH with multiple vacancy defects. Benefiting from the improved electrical conductivity, larger electrochemical surface area (ECSA), and faster charge transfer, the NiFe LDH with O, Ni, and Fe vacancies exhibits a low overpotential of 87 mV at 10 mA/cm 2 for hydrogen evolution reaction (HER) in a pH 7 buffer electrolyte. Impressively, the as-fabricated vacancy-containing NiFe LDH (v-NiFe LDH) splits water with a current density of 10 mA/cm 2 at ∼1.60 V in a two-electrode device, outperforming most other water-splitting catalysts in neutral media. Such an electrolyzer setup could be powered by a commercial 2.0 V solar cell, producing hydrogen at a current density as high as 100 mA/cm 2 .

For nearly a century, oxygen has been widely accepted as the key element that triggers photoresponse in polycrystalline PbSe photoconductive detectors. Our photoluminescence and responsivity studies on PbSe samples, however, suggest that oxygen only serves as an effective sensitization improver and it is iodine rather than oxygen that plays the key role in triggering the photo-response. These studies shed light on the sensitization process for detector applications and ways to passivate defects in IV-VI semiconductors. As a result, high peak detectivity of 2.8 Â 10 10 cm Á Hz 1/2 Á W À1 was achieved at room temperature. V

The current view is that eukaryotic replisomes are independent. Here we show that Ctf4 tightly dimerizes CMG helicase, with an extensive interface involving Psf2, Cdc45, and Sld5. Interestingly, Ctf4 binds only one Pol α-primase. Thus, Ctf4 may have evolved as a trimer to organize two helicases and one Pol α-primase into a replication factory. In the 2CMG–Ctf43–1Pol α-primase factory model, the two CMGs nearly face each other, placing the two lagging strands toward the center and two leading strands out the sides. The single Pol α-primase is centrally located and may prime both sister replisomes. The Ctf4-coupled-sister replisome model is consistent with cellular microscopy studies revealing two sister forks of an origin remain attached and are pushed forward from a protein platform. The replication factory model may facilitate parental nucleosome transfer during replication.