Sequential phosphorylation of CST subunits by different cyclin-Cdk1 complexes orchestrate telomere replication

<p>Telomeres are nucleoprotein structures that cap the ends of linear chromosomes. Telomere homeostasis is central to maintaining genomic integrity. In budding yeast, Cdk1 phosphorylates the telomere-specific binding protein, Cdc13, promoting the recruitment of telomerase to telomere and thereby telomere elongation. Cdc13 is also an integral part of the CST (<u>C</u>dc13-<u>S</u>tn1-<u>T</u>en1) complex that is essential for telomere capping and counteracting telomerase-dependent telomere elongation. Therefore, telomere length homeostasis is a balance between telomerase-extendable and CST-unextendable states. In our earlier work, we showed that Cdk1 also phosphorylates Stn1 which occurs sequentially following Cdc13 phosphorylation during cell cycle progression. This stabilizes the CST complex at the telomere and results in telomerase inhibition. Hence Cdk1-dependent phosphorylations of Stn1 acts like a molecular switch that drives Cdc13 to complex with Stn1-Ten1 rather than with telomerase. However, the underlying mechanism of how a single cyclin-dependent kinase phosphorylates Cdc13 and Stn1 in temporally distinct windows is largely unclear. Here, we show that S phase cyclins are necessary for telomere maintenance. The S phase and mitotic cyclins facilitate Cdc13 and Stn1 phosphorylation respectively, to exert opposing outcomes at the telomere. Thus, our results highlight a previously unappreciated role for cyclins in telomere replication.</p

Azo Coupling Reaction Induced Macromolecular Self-Assembly in Aqueous Solution

This communication reported azo coupling reaction induced macromolecular self-assembly in aqueous solution. Diblock copolymer (PEG-<i>b</i>-PSNHBoc) consisting of a hydrophilic PEG block, and a hydrophobic N-Boc protected poly­(<i>p</i>-vinylaniline) block was synthesized by RAFT polymerization. Then double hydrophilic diblock copolymer (PEG-<i>b</i>-PSN₂⁺) composed of PEG and PS based macromolecular diazonium salts was prepared by the diazotization of PEG-<i>b</i>-PSNH₂, which was obtained by deprotection of PEG-<i>b</i>-PSNHBoc. As <i>N</i>,<i>N</i>-dimethylaniline was gradually added into the freshly prepared PEG-<i>b</i>-PSN₂⁺ aqueous solution, the azo coupling reaction between <i>N</i>,<i>N</i>-dimethylaniline and diazonium salts took place, which would lead to the generation of azobenzene pendants. Due to the poor solubility of azobenzene pendants in water, the formed hydrophobic polymeric chains aggregated to form the self-assembly colloidal particles. By incorporating a fluorescent group into the aniline, the aggregates formed through azo coupling reaction induced macromolecular self-assembly showed enzyme-triggered fluorescent behaviors

Synthesis and Characterization of Photoprocessable Lignin-Based Azo Polymer

Lignin-based azo polymer-bearing pseudo-stilbene-type azo chromophores have been synthesized by a post-azo coupling reaction between the modified alkali lignin and diazonium salts of 4-aminobenzoic acid in DMF with high yield. The polymer synthesized was characterized by using spectroscopic methods. Significant dichroism and surface relief patterns could be photoinscribed on the prepared azo polymer films by using appropriate laser beams. Self-assembly of the lignin-based polymer in selective solvents (THF/H₂O) can give uniform colloidal spheres, which can be elongated along the polarization direction of the irradiation light. This novel synthesized lignin-based azo polymer could potentially be used for applications such as reversible optical data storage, photoswitching, sensors, and other photo-driven devices, which provide a simple strategy for value-added utilization of lignin biomass resources

Discovery of Novel Class I Histone Deacetylase Inhibitors with Promising in Vitro and in Vivo Antitumor Activities

A successful structure-based design of novel cyclic depsipeptides that selectively target class I HDAC isoforms is described. Compound <b>11</b> has an IC₅₀ of 2.78 nM for binding to the HDAC1 protein, and the prodrugs <b>12</b> and <b>13</b> also exhibit promising antiproliferative activities in the nanomolar range against various cancer cell lines. Compounds <b>12</b> and <b>13</b> show more than 20-fold selectivity toward human cancer cells over human normal cells in comparison with romidepsin (FK228), demonstrating low probability of toxic side effects. In addition, compound <b>13</b> exhibits excellent in vivo anticancer activities in a human prostate carcinoma (Du145) xenograft model with no observed toxicity. Thus, prodrug <b>13</b> has therapeutic potential as a new class of anticancer agent for further clinical translation

<i>IFNAR2</i>-dependent gene expression profile induced by IFN-α in <i>Pteropus alecto</i> bat cells and impact of <i>IFNAR2</i> knockout on virus infection

<div><p>Bats are important reservoirs of many viruses, which are capable of infecting the host without inducing obvious clinical diseases. Interferon and the downstream interferon regulated genes (IRGs) are known to act as the first line of defense against viral infections. Little is known about the transcriptional profile of genes being induced by interferon in bats and their role in controlling virus infection. In this study, we constructed <i>IFNAR2</i> knockout bat cell lines using CRISPR technology and further characterized gene expression profiles induced by the most abundant IFN-α (IFN-α3). Firstly, we demonstrated that the CRISPR/Cas9 system is <a href="http://dict.cn/practicable" target="_blank">applicable</a> for bat cells as this represents the first CRIPSR knockout cell line for bats. Our results showed the pleiotropic effect of IFN-α3 on the bat kidney cell line, PaKiT03. As expected, we confirmed that <i>IFNAR2</i> is indispensable for IFN-a signaling pathway and plays an important role in antiviral immunity. Unexpectedly, we also identified novel <i>IFNAR2</i>-dependent IRGs which are enriched in pathways related to cancer. To our knowledge, this seems to be bat-specific as no such observation has been reported for other mammalian species. This study expands our knowledge about bat immunology and the cell line established can provide a powerful tool for future study into virus-bat interaction and cancer biology.</p></div

Discovery of Novel Class I Histone Deacetylase Inhibitors with Promising in Vitro and in Vivo Antitumor Activities

A successful structure-based design of novel cyclic depsipeptides that selectively target class I HDAC isoforms is described. Compound <b>11</b> has an IC₅₀ of 2.78 nM for binding to the HDAC1 protein, and the prodrugs <b>12</b> and <b>13</b> also exhibit promising antiproliferative activities in the nanomolar range against various cancer cell lines. Compounds <b>12</b> and <b>13</b> show more than 20-fold selectivity toward human cancer cells over human normal cells in comparison with romidepsin (FK228), demonstrating low probability of toxic side effects. In addition, compound <b>13</b> exhibits excellent in vivo anticancer activities in a human prostate carcinoma (Du145) xenograft model with no observed toxicity. Thus, prodrug <b>13</b> has therapeutic potential as a new class of anticancer agent for further clinical translation

The top-ranked diseases enriched by IPA analysis for the unknown IRGs.

<p>The top-ranked diseases enriched by IPA analysis for the unknown IRGs.</p

Effect of <i>IFNAR2</i> KO on H1N1 infection in bat cells.

<p>Cells were treated with IFN-α3 for 2 hrs before infected with H1N1 at MOI of 0.1. Cells and Culture supernatants were harvested at 48 hrs post infection. Gene expression was determined by measuring mRNA level using qPCR (A) and the data were normalized against the expression level of the housekeeping gene SNRPD3. Virus titers were determined by plaque assay in BHK cells (B). Error bars indicate standard deviations from three independent experiments.</p

Verification of <i>IFNAR2</i> KO in two cell lines obtained from two independent gRNAs.

<p>(A) Sanger sequencing was performed to validate the location and nature of the deletion events. Left: The location of gRNA and PAM motif are given in blue and red, respectively. The deletion regions are highlighted in gray. Right: Chromatogram of DNA sequence spanning the deletion region. Quantitative RT-PCR (B) and western blot (C) analyses were performed to confirm the functional phenotype of the clones.</p

Number of DEGs induced by IFN-α3 in wild-type PaKiT03 and <i>IFNAR2</i> knockout cell lines.

<p>Number of DEGs induced by IFN-α3 in wild-type PaKiT03 and <i>IFNAR2</i> knockout cell lines.</p