Deregulated Cdc6 inhibits DNA replication and suppresses Cdc7-mediated phosphorylation of Mcm2–7 complex

Mcm2–7 is recruited to eukaryotic origins of DNA replication by origin recognition complex, Cdc6 and Cdt1 thereby licensing the origins. Cdc6 is essential for origin licensing during DNA replication and is readily destabilized from chromatin after Mcm2–7 loading. Here, we show that after origin licensing, deregulation of Cdc6 suppresses DNA replication in Xenopus egg extracts without the involvement of ATM/ATR-dependent checkpoint pathways. DNA replication is arrested specifically after chromatin binding of Cdc7, but before Cdk2-dependent pathways and deregulating Cdc6 after this step does not impair activation of origin firing or elongation. Detailed analyses revealed that Cdc6 deregulation leads to strong suppression of Cdc7-mediated hyperphosphorylation of Mcm4 and subsequent chromatin loading of Cdc45, Sld5 and DNA polymerase α. Mcm2 phosphorylation is also repressed although to a lesser extent. Remarkably, Cdc6 itself does not directly inhibit Cdc7 kinase activity towards Mcm2–4–6–7 in purified systems, rather modulates Mcm2–7 phosphorylation on chromatin context. Taken together, we propose that Cdc6 on chromatin acts as a modulator of Cdc7-mediated phosphorylation of Mcm2–7, and thus destabilization of Cdc6 from chromatin after licensing is a key event ensuring proper transition to the initiation of DNA replication

The PAF1 complex is involved in embryonic epidermal morphogenesis in Caenorhabditis elegans

AbstractThe PAF1 complex (PAF1C) is an evolutionarily conserved protein complex involved in transcriptional regulation and chromatin remodeling. How the PAF1C is involved in animal development is still not well understood. Here, we report that, in the nematode Caenorhabditis elegans, the PAF1C is involved in epidermal morphogenesis in late embryogenesis. From an RNAi screen we identified the C. elegans ortholog of a component of the PAF1C, CTR-9, as a gene whose depletion caused various defects during embryonic epidermal morphogenesis, including epidermal cell positioning, ventral enclosure and epidermal elongation. RNAi of orthologs of other four components of the PAF1C (PAFO-1, LEO-1, CDC-73 and RTFO-1) caused similar epidermal defects. In these embryos, whereas the number and cell fate determination of epidermal cells were apparently unaffected, their position and shape were severely disorganized. PAFO-1::mCherry, mCherry::LEO-1 and GFP::RTFO-1 driven by the authentic promoters were detected in the nuclei of a wide range of cells. Nuclear localization of GFP::RTFO-1 was independent of other PAF1C components, while PAFO-1::mCherry and mCherry::LEO-1 dependent on other components except RTFO-1. Epidermis-specific expression of mCherry::LEO-1 rescued embryonic lethality of the leo-1 deletion mutant. Thus, although the PAF1C is universally expressed in C. elegans embryos, its epidermal function is crucial for the viability of this animal

Accurate deep learning model using semi-supervised learning and Noisy Student for cervical cancer screening in low magnification images.

Deep learning technology has been used in the medical field to produce devices for clinical practice. Deep learning methods in cytology offer the potential to enhance cancer screening while also providing quantitative, objective, and highly reproducible testing. However, constructing high-accuracy deep learning models necessitates a significant amount of manually labeled data, which takes time. To address this issue, we used the Noisy Student Training technique to create a binary classification deep learning model for cervical cytology screening, which reduces the quantity of labeled data necessary. We used 140 whole-slide images from liquid-based cytology specimens, 50 of which were low-grade squamous intraepithelial lesions, 50 were high-grade squamous intraepithelial lesions, and 40 were negative samples. We extracted 56,996 images from the slides and then used them to train and test the model. We trained the EfficientNet using 2,600 manually labeled images to generate additional pseudo labels for the unlabeled data and then self-trained it within a student-teacher framework. Based on the presence or absence of abnormal cells, the created model was used to classify the images as normal or abnormal. The Grad-CAM approach was used to visualize the image components that contributed to the classification. The model achieved an area under the curve of 0.908, accuracy of 0.873, and F1-score of 0.833 with our test data. We also explored the optimal confidence threshold score and optimal augmentation approaches for low-magnification images. Our model efficiently classified normal and abnormal images at low magnification with high reliability, making it a promising screening tool for cervical cytology

R-CHOP with dose-attenuated radiation therapy could induce good prognosis in gastric diffuse large B cell lymphoma

<p>Abstract</p> <p>Background</p> <p>The treatment strategy for gastric diffuse large cell lymphoma (DLBCL) has not been standardized in such as to the cycles of chemotherapy, dose of radiation, or necessity for the surgery. Although the results of CHOP or R-CHOP treatments have demonstrated the good prognosis, the treatments have been controversial in many cases.</p> <p>Methods</p> <p>We retrospectively analyzed 40 gastric DLBCL patients receiving chemotherapy with or without radiation in our institute. Those in stages II-IV were treated with six cycles of R-CHOP without radiation; for those in stage I, we administered three cycles of R-CHOP with radiation.</p> <p>Results</p> <p>The three-year overall survival (OS) and progression-free survival (PFS) rates were 95.2 and 91.8%, respectively. Those in stage I obtained 100% of OS. The radiation dose prescribed was 30.6 Gy for CR cases and 39.6 to 40 Gy for PR after chemotherapy. Although survival rates tended to correlate with staging groups or age-adjusted IPI classifications, multivariate statistical analysis did not show clear differences. All 14 patients with initial bleeding were successfully managed without surgery during treatment.</p> <p>Conclusion</p> <p>R-CHOP therapy was very effective for gastric DLBCL. It may be not necessary to use more than 30.6 Gy of radiotherapy in the highly chemo-sensitive cases. Less toxic treatments should be made available to gastric DLBCL patients.</p