JMJD5 couples with CDK9 to release the paused RNA polymerase II.

More than 30% of genes in higher eukaryotes are regulated by RNA polymerase II (Pol II) promoter proximal pausing. Pausing is released by the positive transcription elongation factor complex (P-TEFb). However, the exact mechanism by which this occurs and whether phosphorylation of the carboxyl-terminal domain of Pol II is involved in the process remains unknown. We previously reported that JMJD5 could generate tailless nucleosomes at position +1 from transcription start sites (TSS), thus perhaps enable progression of Pol II. Here we find that knockout of JMJD5 leads to accumulation of nucleosomes at position +1. Absence of JMJD5 also results in loss of or lowered transcription of a large number of genes. Interestingly, we found that phosphorylation, by CDK9, of Ser2 within two neighboring heptad repeats in the carboxyl-terminal domain of Pol II, together with phosphorylation of Ser5 within the second repeat, HR-Ser2p (1, 2)-Ser5p (2) for short, allows Pol II to bind JMJD5 via engagement of the N-terminal domain of JMJD5. We suggest that these events bring JMJD5 near the nucleosome at position +1, thus allowing JMJD5 to clip histones on this nucleosome, a phenomenon that may contribute to release of Pol II pausing.

Upregulation of NOD1 and NOD2 contribute to cancer progression through the positive regulation of tumorigenicity and metastasis in human squamous cervical cancer.

BACKGROUND: Metastatic cervical squamous cell carcinoma (CSCC) has poor prognosis and is recalcitrant to the current treatment strategies, which warrants the necessity to identify novel prognostic markers and therapeutic targets. Given that CSCC is a virus-induced malignancy, we hypothesized that the pattern recognition receptors (PRRs) involved in the innate immune response likely play a critical role in tumor development. METHODS: A bioinformatics analysis, qPCR, IHC, immunofluorescence, and WB were performed to determine the expression of NOD1/NOD2. The biological characteristics of overexpression NOD1 or NOD2 CSCC cells were compared to parental cells: proliferation, migration/invasion and cytokines secretion were examined in vitro through CCK8/colony formation/cell cycle profiling/cell counting, wound healing/transwell, and ELISA assays, respectively. The proliferative and metastatic capacity of overexpression NOD1 or NOD2 CSCC cells were also evaluated in vivo. FCM, mRNA and protein arrays, ELISA, and WB were used to identify the mechanisms involved, while novel pharmacological treatment were evaluated in vitro and in vivo. Quantitative variables between two groups were compared by Student's t test (normal distribution) or Mann-Whitney U test (non-normal distribution), and one-way or two-way ANOVA was used for comparing multiple groups. Pearson χ2 test or Fisher's exact test was used to compare qualitative variables. Survival curves were plotted by the Kaplan-Meier method and compared by the log-rank test. P values of < 0.05 were considered statistically significant. RESULTS: NOD1 was highly expressed in CSCC with lymph-vascular space invasion (LVSI, P < 0.01) and lymph node metastasis (LM, P < 0.01) and related to worse overall survival (OS, P = 0.016). In vitro and in vivo functional assays revealed that the upregulation of NOD1 or NOD2 in CSCC cells promoted proliferation, invasion, and migration. Mechanistically, NOD1 and NOD2 exerted their oncogenic effects by activating NF-κb and ERK signaling pathways and enhancing IL-8 secretion. Inhibition of the IL-8 receptor partially abrogated the effects of NOD1/2 on CSCC cells. CONCLUSIONS: NOD1/2-NF-κb/ERK and IL-8 axis may be involved in the progression of CSCC; the NOD1 significantly enhanced the progression of proliferation and metastasis, which leads to a poor prognosis. Anti-IL-8 was identified as a potential therapeutic target for patients with NOD1high tumor.

Concurrent definitive chemoradiation incorporating intensity-modulated radiotherapy followed by adjuvant chemotherapy in high risk locally advanced cervical squamous cancer: a phase II study.

BACKGROUND: Although the prognosis of locally advanced cervical cancer has improved dramatically, survival for those with stage IIIB-IVA disease or lymph nodes metastasis remains poor. It is believed that the incorporation of intensity-modulated radiotherapy into the treatment of cervical cancer might yield an improved loco-regional control, whereas more cycles of more potent chemotherapy after the completion of concurrent chemotherapy was associated with a diminished distant metastasis. We therefore initiated a non-randomized prospective phaseII study to evaluate the feasibility of incorporating both these two treatment modality into the treatment of high risk locally advanced cervical cancer. OBJECTIVES: To determine whether the incorporation of intensity-modulated radiotherapy and the addition of adjuvant paclitaxel plus cisplatin regimen into the treatment policy for patients with high risk locally advanced cervical cancer might improve their oncologic outcomes. STUDY DESIGN: Patients were enrolled if they had biopsy proven stage IIIA-IVA squamous cervical cancer or stage IIB disease with metastatic regional nodes. Intensity-modulated radiotherapy was delivered with dynamic multi-leaf collimators using 6MV photon beams. Prescription for PTV ranged from 45.0 ~ 50.0 Gy at 1.8 Gy ~ 2.0 Gy/fraction in 25 fractions. Enlarged nodes were contoured separately and PTV-nodes were boosted simultaneously to a total dose of 50.0-65 Gy at 2.0- 2.6 Gy/fraction in 25 fractions. A total dose of 28 ~ 35 Gy high-dose- rate brachytherapy was prescribed to point A in 4 ~ 5 weekly fractions using an iridium- 192 source. Concurrent weekly intravenous cisplatin at 30 mg/m2 was initiated on the first day of radiotherapy for over 1-h during external-beam radiotherapy. Adjuvant chemotherapy was scheduled within 4 weeks after the completion of concurrent chemo-radiotherapy and repeated 3 weeks later. Paclitaxel 150 mg/m2 was given as a 3-h infusion on day1, followed by cisplatin 35 mg/m2 with 1-h infusion on day1-2 (70 mg/m2 in total). RESULTS: Fifty patients achieved complete response 4 weeks after the completion of the treatment protocol, whereas 2 patients had persistent disease. After a median follow-up period of 66 months, loco-regional (including 2 persistent disease), distant, and synchronous treatment failure occurred in 4,5, and 1, respectively. The 5-year disease-free survival, loco-regional recurrence-free survival, distant-metastasis recurrence-free survival was 80.5%, 90.3%, and 88.0%, respectively. Four of the patients died of the disease, and the 5-year overall survival was 92.1%. Most of the toxicities reported during concurrent chemo-radiotherapy were mild and transient. The occurrence of hematological toxicities elevated mildly during adjuvant chemotherapy, as 32% (16/50) and 4% (2/50) patients experienced grade 3-4 leukopenia and thrombocytopenia, respectively. Grade 3-4 late toxicities were reported in 3 patients. CONCLUSIONS: The incorporation of intensity-modulated radiotherapy and adjuvant paclitaxel plus cisplatin chemotherapy were highly effective and well-tolerated in the treatment of high-risk locally advanced cervical cancer. The former yields an improved loco-regional control, whereas distant metastases could be effectively eradicated with mild toxicities when adjuvant regimen was prescribed.

The Receptor Binding Domain of SARS-CoV-2 Lambda Variant Has a Better Chance Than the Delta Variant in Evading BNT162b2 COVID-19 mRNA Vaccine-Induced Humoral Immunity.

The SARS-CoV-2 Delta and Lambda variants had been named variants of concern (VOC) and variants of interest (VOI), respectively, by the World Health Organization (WHO). Both variants have two mutations in the spike receptor binding domain (RBD) region, with L452R and T478K mutations in the Delta variant, and L452Q and F490S mutations in the Lambda variant. We used surface plasmon resonance (SPR)-based technology to evaluate the effect of these mutations on human angiotensin-converting enzyme 2 (ACE2) and Bamlanivimab binding. The affinity for the RBD ligand, ACE2, of the Delta RBD is approximately twice as strong as that of the wild type RBD, an increase that accounts for the increased infectivity of the Delta variant. On the other hand, in spite of its amino acid changes, the Lambda RBD has similar affinity to ACE2 as the wild type RBD. The protective anti-wild type RBD antibody Bamlanivimab binds very poorly to the Delta RBD and not at all to the Lambda RBD. Nevertheless, serum antibodies from individuals immunized with the BNT162b2 vaccine were found to bind well to the Delta RBD, but less efficiently to the Lambda RBD in contrast. As a result, the blocking ability of ACE2 binding by serum antibodies was decreased more by the Lambda than the Delta RBD. Titers of sera from BNT162b2 mRNA vaccinated individuals dropped 3-fold within six months of vaccination regardless of whether the target RBD was wild type, Delta or Lambda. This may account partially for the fall off with time in the protective effect of vaccines against any variant.

The potential underlying mechanism of the leukemia caused by MLL-fusion and potential treatments.

A majority of infant and pediatric leukemias are caused by the mixed-lineage leukemia gene (MLL) fused with a variety of candidates. Several underlying mechanisms have been proposed. One currently popular view is that truncated MLL1 fusion and its associated complex constitutively hijacks super elongation complex, including positive transcription elongation factor b, CDK9, and cyclin T1 complex and DOT1L, to enhance the expression of transcription factors that maintain or restore stemness of leukocytes, as well as prevent the differentiation of hematopoietic progenitor cells. An alternative emerging view proposes that MLL1-fusion promotes the recruitment of TATA binding protein and RNA polymerase II (Pol II) initiation complex, so as to increase the expression levels of target genes. The fundamental mechanism of both theories are gain of function for truncated MLL1 fusions, either through Pol II elongation or initiation. Our recent progress in transcription regulation of paused Pol II through JMJD5, JMJD6, and JMJD7, combined with the repressive role of H3K4me3 revealed by others, prompted us to introduce a contrarian hypothesis: the failure to shut down transcribing units by MLL-fusions triggers the transformation: loss of function of truncated MLL1 fusions coupled with the loss of conversion of H3K4me1 to H3K4me3, leading to the constitutive expression of transcription factors that are in charge of maintenance of hematopoietic progenitor cells, may trigger the transformation of normal cells into cancer cells. Following this track, a potential treatment to eliminate these fusion proteins, which may ultimately cure the disease, is proposed.

Detection of sentinel lymph node in laparoscopic surgery for uterine cervical cancer using carbon nanoparticles.

BACKGROUND AND OBJECTIVES: Carbon nanoparticles (CNPs) has been widely confirmed the efficiency in sentinel lymph node (SLN) mapping for various solid tumors. This study aims to explore the feasibility and effectiveness of CNPs during laparoscopic surgery for cervical cancer. METHODS: We analyzed 45 women with stage IB1-IIA1 cervical cancer who underwent SLN mapping using CNPs during laparoscopic surgery. The effectiveness of CNPs was evaluated by the detection rate and accuracy parameters. Factors associated with SLN laterality and SLNs localizations were analyzed. RESULTS: The overall and bilateral detection rate was 93.3% (42/45) and 60.0% (27/45), respectively. Elevated body mass index was associated with decreased bilateral detection rate (P = .015). A total of 225 SLNs were harvested, with a mean number of 5.0 ± 3.6. A total of 81.3% of SLNs were in expected localizations including external iliac (39.1%), internal iliac (25.8%), and obturator (16.4%) regions, while 18.7% in unusual localizations including common iliac (10.7%), parametrial (7.6%), and presarcal (0.4%) regions. None positive lymph node was found in non-SLNs with a false-negative rate of 0%. CONCLUSION: Laparoscopic SLN mapping with CNPs appears to be simple and efficient for patients with early-stage cervical cancer.

Clipping of arginine-methylated histone tails by JMJD5 and JMJD7.

Two of the unsolved, important questions about epigenetics are: do histone arginine demethylases exist, and is the removal of histone tails by proteolysis a major epigenetic modification process? Here, we report that two orphan Jumonji C domain (JmjC)-containing proteins, JMJD5 and JMJD7, have divalent cation-dependent protease activities that preferentially cleave the tails of histones 2, 3, or 4 containing methylated arginines. After the initial specific cleavage, JMJD5 and JMJD7, acting as aminopeptidases, progressively digest the C-terminal products. JMJD5-deficient fibroblasts exhibit dramatically increased levels of methylated arginines and histones. Furthermore, depletion of JMJD7 in breast cancer cells greatly decreases cell proliferation. The protease activities of JMJD5 and JMJD7 represent a mechanism for removal of histone tails bearing methylated arginine residues and define a potential mechanism of transcription regulation.

Separate lateral parametrial lymph node dissection improves detection rate of parametrial lymph node metastasis in early-stage cervical cancer: 10-year clinical evaluation in a single center in China.

OBJECTIVE: To investigate the clinical significance of separate lateral parametrial lymph node dissection (LPLND) in improving parametrial lymph node (PLN) and its metastasis detection rate during radical hysterectomy for early-stage cervical cancer. METHODS: From July 2007 to August 2017, 2,695 patients with cervical cancer in stage IB1-IIA2 underwent radical hysterectomy were included. Of these patients, 368 underwent separate dissection of PLNs using the LPLND method, and 2,327 patients underwent conventional radical hysterectomy (CRH). We compared the surgical parameters, PLN detection rate and PLN metastasis rate between the two groups. RESULTS: Compared with CRH group, the rate of laparoscopic surgery was higher (60.3% vs. 15.9%, P<0.001), and the blood transfusion rate was lower (19.0%vs. 29.0%, P<0.001) in the LPLND group. PLNs were detected in 356 cases (96.7%) in the LPLND group, and 270 cases (11.6%) in the CRH group (P<0.001), respectively. The number of PLNs detected in the LPLND group was higher than that in the CRH group (median 3vs. 1, P<0.001). The PLN metastases were detected in 25 cases (6.8%) in the LPLND group, and 18 cases (0.8%) in the CRH group (P<0.001), respectively. In multivariable analysis, LPLND is an independent factor not only for PLN detection [odds ratio (OR)=228.999, 95% confidence interval (95% CI): 124.661-420.664; P<0.001], but also for PLN metastasis identification (OR=10.867, 95% CI: 5.381-21.946; P<0.001). CONCLUSIONS: LPLND is feasible and safe. The surgical method significantly improves the detection rate of PLN and avoids omission of PLN metastasis during radical hysterectomy for early-stage cervical cancer.

Neoadjuvant Bleomycin, Etoposide, and Cisplatin (BEP) Chemotherapy in the Treatment of Extensively Advanced Yolk Sac Tumors: A Single Center Experience.

OBJECTIVE: This study aimed to investigate the role of neoadjuvant bleomycin, etoposide, and cisplatin (BEP) regimen in patients with extensively advanced yolk sac tumors (YSTs). METHODS: Between July 1982 and December 2015, a total of 58 patients with YST were initially treated at our institution, among which 18 were evaluated to be inoperable and received neoadjuvant BEP regimen. They were either too debilitated by the disease [Eastern Cooperative Oncology Group Performance Status Scale (ECOG ps) ≥2] to undergo a major surgery or were with too extensively disseminated lesions to be optimally debulked. This cohort of patients was retrospectively reviewed. RESULTS: One or 2 cycles of BEP regimen were prescribed to the majority of patients preoperatively. At the completion of neoadjuvant chemotherapy, 17 of them had ECOG ps of 1 or less. Seventeen (94.4%) exhibited clinical partial tumor regression, and 1 (5.6%) had clinical stable disease. Pathological complete tumor regression was observed in 2 (11.1%) patients, whereas the remaining 16 (88.9%) had nearly complete pathological regression. Seventeen patients were cytoreduced to no macroscopic residual disease; the remaining 1 was cytoreduced to macroscopic residual disease of 2 cm or less. No major surgical complications occurred. After a median follow-up of 83.5 months, 17 patients were free of recurrence. Five-year disease-free survival and overall survival were both 94.4%. Fertility-sparing surgery was carried out in all the 17 patients with the desire to preserve their fertility, and 5 infants were delivered in 6 patients who attempted conception. CONCLUSIONS: One or 2 cycles of neoadjuvant BEP regimen followed by cytoreductive surgery offer a chance for cure in extensively advanced patients with YSTs and help pave the way for fertility-sparing surgery.

Limited energy parametrial resection/dissection during modified laparoscopic nerve-sparing radical hysterectomy.

OBJECTIVE: The aim of this study was to investigate the feasibility of using ultrasonic scalpel combined with vascular clip in parametrial management, called limited energy parametrial resection/dissection (LEPRD), in laparoscopic nerve plane-sparing radical hysterectomy (NPSRH), a modified nerve-sparing radical hysterectomy (NSRH); and to evaluate its effectiveness in pelvic autonomic nerve preservation. METHODS: From July 2012 to January 2016, 257 consecutive patients with stage IB1 to IIA2 cervical cancer who underwent NPSRH were included in this study. Patients were divided into three cohorts according to the different parametrial resection modality. The clinical, pathological and surgery-related parameters were compared between the three groups. Short- and long-term postoperative bladder functions were evaluated. RESULTS: LEPRD was attempted in 94 patients, and was successful in 65 (69.1%) patients (LEPRD group). The remaining 29 (30.9%) patients required bipolar coagulation after failure of vascular clipping (combined modality group). Routine bipolar cautery was used in the other 163 patients during the parametrial resection (bipolar group). The blood loss in the LEPRD group was significantly lower than those in the other two groups (P<0.001). The rate of successful Foley removal on postoperative day 7 was significantly higher in the LEPRD group than in the bipolar group (P=0.022). The incidence of chronic voiding dysfunction was significantly lower in the LEPRD group than in the bipolar group (P=0.019). CONCLUSIONS: It is feasible to perform LEPRD in NPSRH for cervical cancers. This kind of limited energy surgical technique is associated with less blood loss, and leads to improved postoperative bladder function.

Structural characterization of the pulmonary innate immune protein SPLUNC1 and identification of lipid ligands.

The short palate, lung and nasal epithelial clone 1 (SPLUNC1) protein is a member of the palate, lung, and nasal epithelium clone (PLUNC) family, also known as bactericidal/permeability-increasing (BPI) fold-containing protein, family A, member 1 (BPIFA1). SPLUNC1 is an abundant protein in human airways, but its function remains poorly understood. The lipid ligands of SPLUNC1 as well as other PLUNC family members are largely unknown, although some reports provide evidence that lipopolysaccharide (LPS) could be a lipid ligand. Unlike previous hypotheses, we found significant structural differences between SPLUNC1 and BPI. Recombinant SPLUNC1 produced in HEK 293 cells harbored several molecular species of sphingomyelin and phosphatidylcholine as its ligands. Significantly, in vitro lipid-binding studies failed to demonstrate interactions between SPLUNC1 and LPS, lipoteichoic acid, or polymyxin B. Instead, one of the major and most important pulmonary surfactant phospholipids, dipalmitoylphosphatidylcholine (DPPC), bound to SPLUNC1 with high affinity and specificity. We found that SPLUNC1 could be the first protein receptor for DPPC. These discoveries provide insight into the specific determinants governing the interaction between SPLUNC1 and lipids and also shed light on novel functions that SPLUNC1 and other PLUNC family members perform in host defense.

The structural basis of urea-induced protein unfolding in ß-catenin.

Although urea and guanidine hydrochloride are commonly used to denature proteins, the molecular underpinnings of this process have remained unclear for a century. To address this question, crystal structures of ß-catenin were determined at various urea concentrations. These structures contained at least 105 unique positions that were occupied by urea molecules, each of which interacted with the protein primarily via hydrogen bonds. Hydrogen-bond competition experiments showed that the denaturing effects of urea were neutralized when polyethylene glycol was added to the solution. These data suggest that urea primarily causes proteins to unfold by competing and disrupting hydrogen bonds in proteins. Moreover, circular-dichroism spectra and nuclear magnetic resonance (NMR) analysis revealed that a similar mechanism caused protein denaturation in the absence of urea at pH levels greater than 12. Taken together, the results led to the conclusion that the disruption of hydrogen bonds is a general mechanism of unfolding induced by urea, high pH and potentially other denaturing agents such as guanidine hydrochloride. Traditionally, the disruption of hydrophobic interactions instead of hydrogen bonds has been thought to be the most important cause of protein denaturation.

[Application of laparoscopy in the modified nerve plane-sparing radical hysterectomy of cervical cancer].

OBJECTIVE: The aim of this study was to assess the feasibility and safety of laparoscopic nerve plane-sparing radical hysterectomy (NPSRH) and compare with that of open NPSRH. METHODS: One hundred and thirty-four patients with FIGO stage Ib1-IIa2 cervical cancer were enrolled in the study. Thirty-three patients underwent laparoscopic NPSRH. During the operation, the pelvic autonomic nerve plane which is directly underneath the ureter was integrally preserved by dissecting the pelvic spaces laparoscopically. The vessels around the nerve plane were controlled by Hem-o-lok polymer clips. One hundred and one patients underwent open NPSRH without special instruments. The clinical, pathological and surgery-related parameters were compared between the two groups. Moreover, postoperative short-term bladder function of these patients was also analyzed. RESULTS: There was no significant difference between the laparoscopic group and open group in terms of age, body mass index, previous surgery, FIGO stage, pathologic type, etc. (P > 0.05). The mean duration of surgery in the laparoscopic group was significantly longer [(303.8 ± 67.5) min vs. (272.4 ± 57.5) min] (P < 0.01). But, the laparoscopic group had less blood loss [177.0 ml vs. 474.5 ml, P < 0.01] and blood transfusion rate [ 6.1% (2/33 cases) vs. 49.5% (50/101 cases), P < 0.001]. There was no significant difference regarding the proportion of patients who firstly passed the post-void residual urine volume (PVR) test (P > 0.05). The median time of catheterization between the two groups were also comparable (P > 0.05). However, the postoperative hospital stay was significantly shorter in the laparoscopic group [median postoperative hospital stay 9.2 days vs. 11.0 days, P < 0.001]. CONCLUSIONS: Laparoscopic NPSRH is feasible. It seems to be comparable with open NPSRH in terms of preserving pelvic nerve function, but is more favorable in terms of blood loss and postoperative recovery.

[Clinical analysis of ovarian preservation for stage I endometrial carcinomas in women aged 40 years and younger].

OBJECTIVE: To investigate the safety of ovarian preservation for stage I endometrial carcinomas in women aged 40 years and younger. METHODS: Seventy-five cases of stage I endometrial cancer aged 40 years and younger from Jan 1999 to Jan 2012 were treated in Cancer Hospital, Chinese Academy of Medical Sciences. They were further divided into two groups: 20 patients who underwent ovarian preservation (group A) and 55 patients who underwent oophorectomy (group B). Clinical and pathological recordings of these patients were reviewed and compared. RESULTS: In the group A, there were 13 patients preserved both ovaries, and 7 patients preserved a single ovary. While there were no significant differences in the age, body mass index, surgical staging, histology, grade, cytology of peritoneal lavage or ascites, and postoperative treatment between two groups (all P > 0.05). The differences in the level of CA125 [25% (5/20) versus 18% (10/55)] and number of patients underwent pelvic lymphadenectomy [35% (7/20) versus 84% (46/55)] were statistically significant between two groups (all P < 0.05). Of seventy-five cases, only two patients relapsed and all survived after a median follow-up time of 31.7 months (range: 0 to 160 months). Kaplan-Meier analysis revealed no difference in overall survival (100.0% versus 100.0%) and disease free survival (90.0% versus 95.5%) between two groups (P = 0.579). CONCLUSIONS: Ovarian preservation has no statistically significant impact on the survival of young patients with stage Ia, well differentiated endometrial cancer. Large-scale, prospective clinical studies are needed to validate the safety of ovarian preservation for those patients.

Molecular insight into interactions between the Taf14, Yng1 and Sas3 subunits of the NuA3 complex.

The NuA3 complex is a major regulator of gene transcription and the cell cycle in yeast. Five core subunits are required for complex assembly and function, but it remains unclear how these subunits interact to form the complex. Here, we report that the Taf14 subunit of the NuA3 complex binds to two other subunits of the complex, Yng1 and Sas3, and describe the molecular mechanism by which the extra-terminal domain of Taf14 recognizes the conserved motif present in Yng1 and Sas3. Structural, biochemical, and mutational analyses show that two motifs are sandwiched between the two extra-terminal domains of Taf14. The head-to-toe dimeric complex enhances the DNA binding activity of Taf14, and the formation of the hetero-dimer involving the motifs of Yng1 and Sas3 is driven by sequence complementarity. In vivo assays in yeast demonstrate that the interactions of Taf14 with both Sas3 and Yng1 are required for proper function of the NuA3 complex in gene transcription and DNA repair. Our findings suggest a potential basis for the assembly of three core subunits of the NuA3 complex, Taf14, Yng1 and Sas3.

Phosphatidylinositol 4-kinase IIIß is required for severe acute respiratory syndrome coronavirus spike-mediated cell entry.

Phosphatidylinositol kinases (PI kinases) play an important role in the life cycle of several viruses after infection. Using gene knockdown technology, we demonstrate that phosphatidylinositol 4-kinase IIIß (PI4KB) is required for cellular entry by pseudoviruses bearing the severe acute respiratory syndrome-coronavirus (SARS-CoV) spike protein and that the cell entry mediated by SARS-CoV spike protein is strongly inhibited by knockdown of PI4KB. Consistent with this observation, pharmacological inhibitors of PI4KB blocked entry of SARS pseudovirions. Further research suggested that PI4P plays an essential role in SARS-CoV spike-mediated entry, which is regulated by the PI4P lipid microenvironment. We further demonstrate that PI4KB does not affect virus entry at the SARS-CoV S-ACE2 binding interface or at the stage of virus internalization but rather at or before virus fusion. Taken together, these results indicate a new function for PI4KB and suggest a new drug target for preventing SARS-CoV infection.

Interaction of JMJD6 with single-stranded RNA.

JMJD6 is a Jumonji C domain-containing hydroxylase. JMJD6 binds alpha-ketoglutarate and iron and has been characterized as either a histone arginine demethylase or U2AF65 lysyl hydroxylase. Here, we describe the structures of JMJD6 with and without alpha-ketoglutarate, which revealed a novel substrate binding groove and two positively charged surfaces. The structures also contain a stack of aromatic residues located near the active center. The side chain of one residue within this stack assumed different conformations in the two structures. Interestingly, JMJD6 bound efficiently to single-stranded RNA, but not to single-stranded DNA, double-stranded RNA, or double-stranded DNA. These structural features and truncation analysis of JMJD6 suggest that JMJD6 may bind and modify single-stand RNA rather than the previously reported peptide substrates.

Factors influencing entrepreneurial intention of university students in Yemen: The mediating role of entrepreneurial self-efficacy.

While entrepreneurship is believed to play a crucial role in economic growth and job creation in various parts of the world, particularly in developed countries, the key factors enhancing entrepreneurship behavior and intention in developing countries still need to be discovered. Therefore, this study examines the influence of personality traits and environmental and situational factors on the development of entrepreneurial intention among young students in Yemen. Data were collected through a survey responded to by 487 final-year university students from two universities (public and private) in Yemen. The study's hypotheses were tested using structural equation modeling (SEM). The study reveals that personality traits of the need for achievement (nAch) and locus of control (LoC) positively correlate with entrepreneurial self-efficacy (ESE) and entrepreneurial intention. Instrumental readiness positively correlates with ESE but not with entrepreneurial intent. The situational factors show a positive association with entrepreneurial intention but not ESE and a positive relationship between ESE and entrepreneurial intention. Furthermore, the study's findings show that ESE partially mediates the relationship between the nAch, LoC, instrumental readiness, and entrepreneurial intention. However, ESE did not mediate the relationship between situational factors and entrepreneurial intention. The study suggests that situational factors can influence entrepreneurial intention among Yemeni students and provide several recommendations to academicians and policymakers.

Molecular insight into the PCNA-binding mode of FBH1.

F-box DNA helicase 1 (FBH1) is involved in the regulation of cell responses to replicative stress. FBH1 is recruited to stalled DNA replication fork by PCNA where it inhibits homologous recombination and catalyzes fork regression. Here, we report the structural basis for the molecular recognition of two distinctly different motifs of FBH1, FBH1PIP and FBH1APIM, by PCNA. The crystal structure of PCNA in complex with FBH1PIP and analysis of NMR perturbations reveal overlapped FBH1PIP and FBH1APIM binding sites of PCNA and the dominant contribution of FBH1PIP in this interaction.

SPLUNC1 deficiency enhances airway eosinophilic inflammation in mice.

Short palate, lung and nasal epithelium clone 1 (SPLUNC1) is enriched in normal airway lining fluid, but is significantly reduced in airway epithelium exposed to a Th2 cytokine milieu. The role of SPLUNC1 in modulating airway allergic inflammation (e.g., eosinophils) remains unknown. We used SPLUNC1 knockout (KO) and littermate wild-type (C57BL/6 background) mice and recombinant SPLUNC1 protein to determine the impact of SPLUNC1 on airway allergic/eosinophilic inflammation, and to investigate the underlying mechanisms. An acute ovalbumin (OVA) sensitization and challenge protocol was used to induce murine airway allergic inflammation (e.g., eosinophils, eotaxin-2, and Th2 cytokines). Our results showed that SPLUNC1 in the bronchoalveolar lavage fluid of OVA-challenged wild-type mice was significantly reduced (P < 0.05), which was negatively correlated with levels of lung eosinophilic inflammation. Moreover, SPLUNC1 KO mice demonstrated significantly higher numbers of eosinophils in the lung after OVA challenges than did wild-type mice. Alveolar macrophages isolated from OVA-challenged SPLUNC1 KO versus wild-type mice had higher concentrations of baseline eotaxin-2 that was amplified by LPS (a known risk factor for exacerbating asthma). Human recombinant SPLUNC1 protein was applied to alveolar macrophages to study the regulation of eotaxin-2 in the context of Th2 cytokine and LPS stimulation. Recombinant SPLUNC1 protein attenuated LPS-induced eotaxin-2 production in Th2 cytokine-pretreated murine macrophages. These findings demonstrate that SPLUNC1 inhibits airway eosinophilic inflammation in allergic mice, in part by reducing eotaxin-2 production in alveolar macrophages.