Oncogenic Biogenesis of pri-miR-17∼92 Reveals Hierarchy and Competition among Polycistronic MicroRNAs.

The microRNAs encoded by the miR-17∼92 polycistron are commonly overexpressed in cancer and orchestrate a wide range of oncogenic functions. Here, we identify a mechanism for miR-17∼92 oncogenic function through the disruption of endogenous microRNA (miRNA) processing. We show that, upon oncogenic overexpression of the miR-17∼92 primary transcript (pri-miR-17∼92), the microprocessor complex remains associated with partially processed intermediates that aberrantly accumulate. These intermediates reflect a series of hierarchical and conserved steps in the early processing of the pri-miR-17∼92 transcript. Encumbrance of the microprocessor by miR-17∼92 intermediates leads to the broad but selective downregulation of co-expressed polycistronic miRNAs, including miRNAs derived from tumor-suppressive miR-34b/c and from the Dlk1-Dio3 polycistrons. We propose that the identified steps of polycistronic miR-17∼92 biogenesis contribute to the oncogenic re-wiring of gene regulation networks. Our results reveal previously unappreciated functional paradigms for polycistronic miRNAs in cancer.

Investigating brain-gut microbiota dynamics and inflammatory processes in an autistic-like rat model using MRI biomarkers during childhood and adolescence.

Autism spectrum disorder (ASD) is characterized by social interaction deficits and repetitive behaviors. Recent research has linked that gut dysbiosis may contribute to ASD-like behaviors. However, the exact developmental time point at which gut microbiota alterations affect brain function and behavior in patients with ASD remains unclear. We hypothesized that ASD-related brain microstructural changes and gut dysbiosis induce metabolic dysregulation and proinflammatory responses, which collectively contribute to the social behavioral deficits observed in early childhood. We used an autistic-like rat model that was generated via prenatal valproic acid exposure. We analyzed brain microstructural changes using diffusion tensor imaging (DTI) and examined microbiota, blood, and fecal samples for inflammation biomarkers. The ASD model rats exhibited significant brain microstructural changes in the anterior cingulate cortex, hippocampus, striatum, and thalamus; reduced microbiota diversity (Prevotellaceae and Peptostreptococcaceae); and altered metabolic signatures. The shift in microbiota diversity and density observed at postnatal day (PND) 35, which is a critical developmental period, underscored the importance of early ASD interventions. We identified a unique metabolic signature in the ASD model, with elevated formate and reduced acetate and butyrate levels, indicating a dysregulation in short-chain fatty acid (SCFA) metabolism. Furthermore, increased astrocytic and microglial activation and elevated proinflammatory cytokines-interleukin-1 beta (IL-1ß), interleukin-6 (IL-6), interferon-gamma (IFN-γ), and tumor necrosis factor-alpha (TNF-α)-were observed, indicating immune dysregulation. This study provided insights into the complex interplay between the brain and the gut, and indicated DTI metrics as potential imaging-based biomarkers in ASD, thus emphasizing the need for early childhood interventions.

Robotic Versus Conventional or Endoscopic-assisted Nipple-sparing Mastectomy and Immediate Prosthesis Breast Reconstruction in the Management of Breast Cancer: A Prospectively Designed Multicenter Trial Comparing Clinical Outcomes, Medical Cost, and Patient-reported Outcomes (RCENSM-P).

OBJECTIVE: To compare the clinical and patient-reported outcomes of minimal access and conventional nipple-sparing mastectomy (C-NSM). The secondary outcomes investigated included medical costs and oncological safety. BACKGROUND: Minimal-access NSM has been increasingly applied in the treatment of patients with breast cancer. However, prospective multicenter trials comparing robotic-assisted NSM (R-NSM) versus C-NSM or endoscopic-assisted NSM (E-NSM) are lacking. METHODS: A prospectively designed 3-arm multicenter, nonrandomized trial (NCT04037852) was conducted from October 1, 2019 to December 31, 2021, to compare R-NSM with C-NSM or E-NSM. RESULTS: A total of 73 R-NSM, 74 C-NSM, and 84 E-NSM procedures were enrolled. The median wound length and operation time of C-NSM was (9 cm, 175 minutes), (4 cm, and 195 minutes) in R-NSM, and (4 cm and 222 minutes) in E-NSM. Complications were comparable among the groups. Better wound healing was observed in the minimal-access NSM group. The R-NSM procedure was 4000 and 2600 United States Dollars more expensive than C-NSM and E-NSM, respectively. Wound/scar and postoperative acute pain evaluation favored the use of minimal access NSM over C-NSM. Quality of life in terms of chronic breast/chest pain, mobility, and range of motion of the upper extremity showed no significant differences. The preliminary oncologic results showed no differences among the 3 groups. CONCLUSIONS: R-NSM or E-NSM is a safe alternative if compared with C-NSM in terms of perioperative morbidities, especially with better wound healing. The advantage of minimal access groups was higher wound-related satisfaction. Higher costs remain one of the major limiting factors in the widespread adoption of R-NSM.

Distinct, opposing functions for CFIm59 and CFIm68 in mRNA alternative polyadenylation of Pten and in the PI3K/Akt signalling cascade.

Precise maintenance of PTEN dosage is crucial for tumor suppression across a wide variety of cancers. Post-transcriptional regulation of Pten heavily relies on regulatory elements encoded by its 3'UTR. We previously reported the important diversity of 3'UTR isoforms of Pten mRNAs produced through alternative polyadenylation (APA). Here, we reveal the direct regulation of Pten APA by the mammalian cleavage factor I (CFIm) complex, which in turn contributes to PTEN protein dosage. CFIm consists of the UGUA-binding CFIm25 and APA regulatory subunits CFIm59 or CFIm68. Deep sequencing analyses of perturbed (KO and KD) cell lines uncovered the differential regulation of Pten APA by CFIm59 and CFIm68 and further revealed that their divergent functions have widespread impact for APA in transcriptomes. Differentially regulated genes include numerous factors within the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signalling pathway that PTEN counter-regulates. We further reveal a stratification of APA dysregulation among a subset of PTEN-driven cancers, with recurrent alterations among PI3K/Akt pathway genes regulated by CFIm. Our results refine the transcriptome selectivity of the CFIm complex in APA regulation, and the breadth of its impact in PTEN-driven cancers.

Ochratoxin A detoxification potentials of basil, chan, and chia seeds.

The most toxic of the ochratoxins is ochratoxin A (OTA), which is primarily produced by species of Aspergillus and Penicillium that can be found in maize, wheat, coffee, red wine, and various grains. OTA induces immunotoxicity, nephrotoxicity, hepatotoxicity, teratogenicity, and carcinogenicity in both animals and humans. Thus, there is a need to identify mycotoxin detoxification agents that can effectively decontaminate OTA. Seeds of basil (Ocimum basilicum L.), chan (Hyptis suaveolens L.), and chia (Salvia hispanica L.) are functional foods capable of eliminating harmful substances. Despite this potential, the impact of these seeds on OTA detoxification remains unclear. This study reveals that milled basil, chan, and chia seeds adsorb significant levels of OTA, with chia demonstrating the highest adsorption capacity, followed by chan and basil seeds showing the least efficiency. Furthermore, milled basil, chan, and chia seeds effectively reduced OTA residues in artificial gastric and intestinal fluids, where they achieved up to 93% OTA adsorption in the former. In addition, these milled seeds were able to remove OTAs from canned, drip, and instant coffee. This study is the first to report the OTA elimination potential of basil, chan, and chia seeds.

Bicolor Tuning and Hyper-Reflective Color Switching Based on Two Stacked Cholesteric Liquid Crystal Cells with Asymmetric Electrothermal Optical Responses.

We propose a double-cell cholesteric liquid crystal (CLC) device composed of a left-handed (LH) CLC cell with a pair of sheet electrodes and a right-handed (RH) CLC cell with a tri-electrode configuration characterized by a sheet electrode on the top and an interdigitated electrode on the bottom substrates. Bi-reflected color tuning and hyper-reflective color switching are revealed from this cell stack via the electrothermal control of the central wavelengths of the LH- and RH-bandgaps by voltage-induced pseudo-dielectric heating. The two CLCs are thermally sensitive and exhibit overlapped bandgaps in the field-off state with nearly identical temperature dependence, resulting in a hyper-reflective color at 720 nm at 23.4 °C and 380 nm at 29.8 °C. Upon the application of 4 Vrms at 2 MHz across the stacked device to induce pseudo-dielectric heating, two reflective colors can be resolved due to asymmetrical temperature elevations. Accordingly, the difference in wavelength between the two colors increases with increasing voltage through a series cell connection, while maintaining approximately constant via a parallel connection. This study provides a feasible pathway to developing a multifunctional device with electrothermally tunable bi-reflected and hyper-reflective states based on two conventional cell geometries, which is promising for lasers and color-related display applications.

Relationship Between Chronic Kidney Disease, Glucose Homeostasis, and Plasma Osteocalcin Carboxylation and Fragmentation.

OBJECTIVE: Chronic kidney disease (CKD) is associated with reduced insulin sensitivity, through mechanisms that are not well understood. Low vitamin K intake and incomplete carboxylation of the vitamin K-dependent protein osteocalcin may promote insulin resistance. We assessed relationships of osteocalcin concentration, carboxylation, and fragmentation with CKD and glucose homeostasis in a cross-sectional study. METHODS: We included 87 participants without diabetes: 50 (27 female) with moderate to severe CKD (estimated glomerular filtration rate <60 mL/min/1.73 m2 not treated with dialysis) and 37 (17 female) healthy controls. Total osteocalcin was measured by immunoassay, and osteocalcin carboxylation and fragmentation status by liquid chromatography-electrospray ionization-based mass spectrometric immunoassay. Endpoints included glucose tolerance (based on 2-hour oral glucose tolerance test), insulin sensitivity (hyperinsulinemic-euglycemic clamp), and pancreatic beta-cell function (intravenous glucose tolerance test). RESULTS: The total plasma osteocalcin concentration was higher in the CKD group (mean [standard deviation] 102.9 [147.5]) than that in the control group (53.6 [51.1] ng/mL, P = .03), and more osteocalcin was circulating as fragments. The extent of osteocalcin carbocylation did not differ between individuals with and without CKD. Osteocalcin concentration, carboxylation, and fragmentation were not associated with any measure of glucose homeostasis in multivariable-adjusted analyses. CONCLUSIONS: In CKD, circulating osteocalcin concentrations are elevated, in part due to larger proportions of fragmented forms. However, osteocalcin carboxylation status is not significantly different between individuals with and without CKD. Our data also do not provide support for the hypothesis that differences in osteocalcin carboxylation may explain reduced insulin sensitivity in individuals with CKD.

Urban-rural differences in the prevalence and correlates of mild cognitive impairment in community-dwelling older adults in Taiwan: The EMCIT study.

BACKGROUND: Taiwan is a rapidly aging society. The elderly with mild cognitive impairment (MCI) have increased risk of dementia, and this is a population-based report using standard neuropsychological tests and expert consensus diagnosis to assess the MCI prevalence and its associated factors in Taiwan. METHOD: The Epidemiology of Mild Cognitive Impairment study in Taiwan (EMCIT) is a community-based, prospective cohort study. Independently-living individuals aged ≧60 years in a rural area (n = 122) and in an urban area (n = 348) of New Taipei City, Taiwan, completed detailed neuropsychological tests at the cohort baseline. Diagnosis of MCI was ascertained through expert consensus based on 2011 NIA-AA criteria. RESULTS: Of 470 participants recruited between 2017 and 2019 (mean age 71.2 ± 5.4 years), the prevalence of MCI was higher in the rural area than in the urban area (25.1% vs. 10.8%, p < 0.001) after standardized for age, gender, and level of education. Having lower education and having depression symptoms were consistently associated with increased risk of MCI in both urban and rural areas (p < 0.05). Being male and diabetes were additionally associated with MCI prevalence in urban areas. CONCLUSION: In this community-based prospective cohort study in Taiwan, the prevalence of MCI in the rural community was much higher than that in the urban community. Different strategies may be needed to targeted different types of communities.

Co-targeting bromodomain and extra-terminal proteins and MCL1 induces synergistic cell death in melanoma.

The treatment of melanoma has been markedly improved by the introduction of targeted therapies and checkpoint blockade immunotherapy. Unfortunately, resistance to these therapies remains a limitation. Novel anticancer therapeutics targeting the MCL1 anti-apoptotic protein have shown impressive responses in haematological cancers but are yet to be evaluated in melanoma. To assess the sensitivity of melanoma to new MCL1 inhibitors, we measured the response of 51 melanoma cell lines to the novel MCL1 inhibitor, S63845. Additionally, we assessed combination of this drug with inhibitors of the bromodomain and extra-terminal (BET) protein family of epigenetic readers, which we postulated would assist MCL1 inhibition by downregulating anti-apoptotic targets regulated by NF-kB such as BCLXL, BCL2A1 and XIAP, and by upregulating pro-apoptotic proteins including BIM and NOXA. Only 14% of melanoma cell lines showed sensitivity to S63845, however, combination of S63845 and I-BET151 induced highly synergistic apoptotic cell death in all melanoma lines tested and in an in vivo xenograft model. Cell death was dependent on caspases and BAX/BAK. Although the combination of drugs increased the BH3-only protein, BIM, and downregulated anti-apoptotic proteins such as BCL2A1, the importance of these proteins in inducing cell death varied between cell lines. ABT-199 or ABT-263 inhibitors against BCL2 or BCL2 and BCLXL, respectively, induced further cell death when combined with S63845 and I-BET151. The combination of MCL1 and BET inhibition appears to be a promising therapeutic approach for metastatic melanoma, and presents opportunities to add further BCL2 family inhibitors to overcome treatment resistance.

A prominent environmental endocrine disruptor, 4-nonylphenol, promotes endometriosis development via plasmacytoid dendritic cells.

Endometriosis is an estrogen-dependent chronic inflammatory disease and is associated etiologically with environmental endocrine disruptor (EED) exposure. 4-nonylphenol (NP), a widely found EED, has weak estrogenic activity and modulates plasmacytoid dendritic cell (pDC) function in vitro and in vivo. We aimed to elucidate the immunomodulatory effect of NP on the development of endometriosis, particularly focusing on pDCs. This study established a surgically induced endometriosis murine model (C57BL/6) under conditions of NP treatment that are relevant to the level and route of human exposure. Multi-parametric flow cytometry was used for analysis of infiltrated immune cell subsets in lesions. The results showed that NP exposure significantly promoted endometriotic lesion growth, survival and angiogenesis development of lesions as well as pDC accumulation in the lesions in mice. Adoptive transfer of NP-conditioned pDCs into mice significantly enhanced lesion development and local pDC infiltration, whereas NP-conditioned conventional dendritic cells did not affect lesion growth. In vitro functional analysis showed that NP-conditioned pDCs in lesions expressed high levels of CD36, a scavenger receptor and NP-conditioned splenic pDCs secreted an enhanced level of IL-10 in response to apoptotic cell recognition in a CD36-dependent manner. Furthermore, we observed that local treatment with blocking antibodies against IL-10 and CD36 on the day of surgery significantly inhibited lesion development. NP exposure also altered the estrous cycle in mice. The results suggest that chronic and low-dose exposure to NP enhances endometriotic lesion growth by altering pDC homeostasis and function. This study has important implications for understanding the environment-innate immunity interaction in human endometriosis.

Alternative polyadenylation confers Pten mRNAs stability and resistance to microRNAs.

Fine regulation of the phosphatase and tensin homologue (PTEN) phosphatase dosage is critical for homeostasis and tumour suppression. The 3'-untranslated region (3'-UTR) of Pten mRNA was extensively linked to post-transcriptional regulation by microRNAs (miRNAs). In spite of this critical regulatory role, alternative 3'-UTRs of Pten have not been systematically characterized. Here, we reveal an important diversity of Pten mRNA isoforms generated by alternative polyadenylation sites. Several 3'-UTRs are co-expressed and their relative expression is dynamically regulated. In spite of encoding multiple validated miRNA-binding sites, longer isoforms are largely refractory to miRNA-mediated silencing, are more stable and contribute to the bulk of PTEN protein and signalling functions. Taken together, our results warrant a mechanistic re-interpretation of the post-transcriptional mechanisms involving Pten mRNAs and raise concerns on how miRNA-binding sites are being validated.

Dedicated cone-beam breast CT using laterally-shifted detector geometry: Quantitative analysis of feasibility for clinical translation.

BACKGROUND: High-resolution, low-noise detectors with minimal dead-space at chest-wall could improve posterior coverage and microcalcification visibility in the dedicated cone-beam breast CT (CBBCT). However, the smaller field-of-view necessitates laterally-shifted detector geometry to enable optimizing the air-gap for x-ray scatter rejection. OBJECTIVE: To evaluate laterally-shifted detector geometry for CBBCT with clinical projection datasets that provide for anatomical structures and lesions. METHODS: CBBCT projection datasets (n = 17 breasts) acquired with a 40×30 cm detector (1024×768-pixels, 0.388-mm pixels) were truncated along the fan-angle to emulate 20.3×30 cm, 22.2×30 cm and 24.1×30 cm detector formats and correspond to 20, 120, 220 pixels overlap in conjugate views, respectively. Feldkamp-Davis-Kress (FDK) algorithm with 3 different weighting schemes were used for reconstruction. Visual analysis for artifacts and quantitative analysis of root-mean-squared-error (RMSE), absolute difference between truncated and 40×30 cm reconstructions (Diff), and its power spectrum (PSDiff) were performed. RESULTS: Artifacts were observed for 20.3×30 cm, but not for other formats. The 24.1×30 cm provided the best quantitative results with RMSE and Diff (both in units of µ, cm-1) of 4.39×10-3±1.98×10-3 and 4.95×10-4±1.34×10-4, respectively. The PSDiff (>0.3 cycles/mm) was in the order of 10-14µ2mm3 and was spatial-frequency independent. CONCLUSIONS: Laterally-shifted detector CBBCT with at least 220 pixels overlap in conjugate views (24.1×30 cm detector format) provides quantitatively accurate and artifact-free image reconstruction.

The smut fungus Ustilago esculenta has a bipolar mating system with three idiomorphs larger than 500 kb.

Zizania latifolia Turcz., which is mainly distributed in Asia, has had a long cultivation history as a cereal and vegetable crop. On infection with the smut fungus Ustilago esculenta, Z. latifolia becomes an edible vegetable, water bamboo. Two main cultivars, with a green shell and red shell, are cultivated for commercial production in Taiwan. Previous studies indicated that cultivars of Z. latifolia may be related to the infected U. esculenta isolates. However, related research is limited. The infection process of the corn smut fungus Ustilago maydis is coupled with sexual development and under control of the mating type locus. Thus, we aimed to use the knowledge of U. maydis to reveal the mating system of U. esculenta. We collected water bamboo samples and isolated 145 U. esculenta strains from Taiwan's major production areas. By using PCR and idiomorph screening among meiotic offspring and field isolates, we identified three idiomorphs of the mating type locus and found no sequence recombination between them. Whole-genome sequencing (Illumina and PacBio) suggested that the mating system of U. esculenta was bipolar. Mating type locus 1 (MAT-1) was 552,895 bp and contained 44% repeated sequences. Sequence comparison revealed that U. esculenta MAT-1 shared high gene synteny with Sporisorium reilianum and many repeats with Ustilago hordei MAT-1. These results can be utilized to further explore the genomic diversity of U. esculenta isolates and their application for water bamboo breeding.

H2B ubiquitylation and the histone chaperone Asf1 cooperatively mediate the formation and maintenance of heterochromatin silencing.

Heterochromatin is a heritable form of gene repression, with critical roles in development and cell identity. Understanding how chromatin factors results in such repression is a fundamental question. Chromatin is assembled and disassembled during transcription, replication and repair by anti-silencing function 1 (Asf1), a highly conserved histone chaperone. Transcription and DNA replication are also affected by histone modifications that modify nucleosome dynamics, such as H2B ubiquitylation (H2Bub). We report here that H2Bub and Asf1 cooperatively promote transcriptional silencing at yeast telomeres and mating loci. Through real time monitoring of HML (Hidden MAT Left) locus silencing, we found that transcriptional repression was slowly initiated and never fully established in mutants lacking both Asf1 and H2Bub. These findings are consistent with impaired HML silencer-binding and spreading of repressor proteins, Sir2 and Sir3. In addition, mutants lacking H2Bub and Asf1 show defects in both nucleosome assembly and higher-order heterochromatin organization at the HML locus. Our findings reveal a novel role for H2Bub and Asf1 in epigenetic silencing at mating loci. Thus, the interplay between H2Hbub and Asf1 may fine-tune nucleosome dynamics and SIR protein recruitment, and represent an ongoing requirement for proper formation and maintenance of heterochromatin.

3D-printed CuO nanoparticle-functionalized flow reactor enables online fluorometric monitoring of glucose.

An online flow reactor was fabricated by using a fused deposition modeling three-dimensional printing (3DP) technology along with thermoplastic poly(lactic acid) filaments incorporating copper oxide nanoparticles (CuO NPs). In the presence of glucose, the flow reactor displays multi-catalytic activities because accelerates the oxidation of 2',7'-dichlorodihydrofluorescein to form fluorescein which displays green fluorescence under 480 nm excitation (emission wavelength: 530 nm). The CuO NPs exert two functions to mediate electron transfer at a basic reaction condition, viz. direct oxidation of glucose to generate reactive oxygen species (ROS), and prompting the ROS to oxidize 2',7'-dichlorofluorescin diacetate. The flow reactor coupled to a microdialysis sampler and a fluorometer was applied for online fluorometric monitoring of brain extracellular glucose levels in living rats based on scanning of time-resolved fluorescence intensities. After optimization of (a) the manufacture of the flow reactor, (b) the reaction conditions (pH 10; 50 °C), and (c) the online analytical system, the detection limit of the method (when using 10-µL samples of microdialysate) is as low as 6.1 µM (linear range: 0.05-5 mM) with a sampling frequency of 7.5 h-1. To illustrate the method's applicability, analyses of spiked off-line-collected rat brain microdialysates were conducted. In addition, rat brain extracellular glucose levels were monitored in-vivo and online upon neuronal depolarization triggered by perfusing a high-K+ medium. The results demonstrate that functionalizing raw 3DP materials with appropriate nanomaterials can simplify the manufacturing of analytical devices and related analytical procedures. This will extend the diversity and adaptability of current 3DP-enabling analytical strategies. Graphical abstract Schematic presentation of an online flow reactor fabricated using a fused deposition modeling 3D printer along with poly(lactic acid) (PLA) filaments incorporating CuO NPs. The manufactured flow reactor displays multi-catalytic activities and simplifies online fluorometric monitoring of living rat brain extracellular glucose.

Cognitive impairment in patients with end-stage renal disease: Accelerated brain aging?

BACKGROUND: Chronic kidney disease exhibits a prominent premature aging phenotype in many different organ systems, including the brain. Nevertheless, a comprehensive characterization of brain aging in non-demented patients with end-stage renal disease (ESRD) is lacking and it remains unclear if the collective changes of cognitive functions and brain structures in ESRD is compatible with aging. METHODS: We compared 56 non-demented, independently living dialysis patients (mean age 59.4 ± 11.0 years; mean dialysis vintage of 5.9 years) and 60 non-dialysis controls on a battery of neuropsychological tests, brain MRI T1 imaging and diffusion tensor imaging. Participants with diagnosis of dementia, Mini-Mental State Examination <24, medical history of stroke, or recent hospitalization within 1 month were excluded. RESULTS: Dialysis patients showed significantly worse performance in attention/information processing speed and executive function adjusted for age, sex, education, diabetes and depression. Reduced total brain volume and subcortical volume including hippocampus were found in dialysis patients. Vertex-wise analysis showed cortical thinning in middle frontal, lateral occipital and precuneus region. Furthermore, decreased white matter integrity was found primarily in bilateral anterior thalamic tract, fronto-occipital fasciculus, forceps minor and uncinate tract after correction for multiple comparisons. CONCLUSION: Overall, differences in cognitive functions, cortical volumes/thickness and white matter integrity associated with dialysis are also cognitive domains and brain structure changes associated with normal aging. In other words, non-demented, independently living dialysis patients present an accelerated brain aging phenotype even after taking into account effects of age, diabetes and depression.

Which nasopharyngeal cancer patients need adaptive radiotherapy?

BACKGROUND: Adaptive radiotherapy (ART) has potential benefits in patients with nasopharyngeal cancer (NPC). This retrospective study aimed to identify the factors favoring ART. MATERIALS AND METHODS: Forty NPC patients were retrospectively included in this study. All patients received two-phase, volumetric modulated arc radiotherapy (VMAT) and underwent a second computed tomography (CT) for the phase II ART. We generated phantom, non-ART plans by a hybrid method for comparison with ART plans. A paired t-test was used to evaluate the dose differences between these two plans. A subgroup analysis through a paired t-test was used to evaluate the factors favoring ART. RESULTS: The second CT images were captured at the median 22 fractions. The median total dose of the planning target volume-one (PTV-1) was 72 Gy, and the phase II dose was 16 Gy. The volumes of the ipsilateral parotid gland (23.2 vs. 19.2 ml, p <  0.000), contralateral parotid gland (23.0 vs. 18.4 ml, p <  0.000), clinical target volume-1 (CTV-1, 32.2 vs. 20.9 ml, p <  0.000), and PTV-1 (125.8 vs. 107.3 ml, p <  0.000) all shrunk significantly between these two CT simulation procedures. Among the nearby critical organs, only the ipsilateral parotid gland displayed significant dose reduction by the ART plan (5.3 vs. 6.0 Gy, p = 0.004). Compared to the phantom plan, the ART could significantly improve the PTV-1 target volume coverage of D98 (15.4 vs. 12.3 Gy, p < 0.000). Based on the D98 of PTV-1, the factors of a large initial weight (> 60 kg, p < 0.000), large body mass index (BMI) (> 21.5, p < 0.000), obvious weight loss (> 2.8 kg, p < 0.000), concurrent chemoradiotherapy (p < 0.000), and stages III-IV (p < 0.000) favored the use of ART. CONCLUSIONS: ART could significantly reduce the mean dose to the ipsilateral parotid gland. ART has dosimetrical benefit for patients with a heavy initial weight, large BMI, obvious weight loss, concurrent chemoradiotherapy, and cancer in stages III-IV.

H2B mono-ubiquitylation facilitates fork stalling and recovery during replication stress by coordinating Rad53 activation and chromatin assembly.

The influence of mono-ubiquitylation of histone H2B (H2Bub) on transcription via nucleosome reassembly has been widely documented. Recently, it has also been shown that H2Bub promotes recovery from replication stress; however, the underling molecular mechanism remains unclear. Here, we show that H2B ubiquitylation coordinates activation of the intra-S replication checkpoint and chromatin re-assembly, in order to limit fork progression and DNA damage in the presence of replication stress. In particular, we show that the absence of H2Bub affects replication dynamics (enhanced fork progression and reduced origin firing), leading to γH2A accumulation and increased hydroxyurea sensitivity. Further genetic analysis indicates a role for H2Bub in transducing Rad53 phosphorylation. Concomitantly, we found that a change in replication dynamics is not due to a change in dNTP level, but is mediated by reduced Rad53 activation and destabilization of the RecQ helicase Sgs1 at the fork. Furthermore, we demonstrate that H2Bub facilitates the dissociation of the histone chaperone Asf1 from Rad53, and nucleosome reassembly behind the fork is compromised in cells lacking H2Bub. Taken together, these results indicate that the regulation of H2B ubiquitylation is a key event in the maintenance of genome stability, through coordination of intra-S checkpoint activation, chromatin assembly and replication fork progression.