Hyposecretion of cervical MUC5B is related to preterm birth in pregnant women after cervical excisional surgery.

PROBLEM: Excisional surgery for cervical intraepithelial neoplasia is a risk factor for preterm birth in subsequent pregnancies. However, the underlying mechanisms of this association remain unclear. We previously showed that cervical MUC5B, a mucin protein, may be a barrier to ascending pathogens during pregnancy. We thus hypothesized that hyposecretion of cervical MUC5B is associated with preterm birth after cervical excisional surgery. METHOD OF STUDY: This prospective nested case-control study (Study 1) included pregnant women who had previously undergone cervical excisional surgery across 11 hospitals. We used proteomics to compare cervicovaginal fluid at 18-22 weeks of gestation between the preterm and term birth groups. In another case-control analysis (Study 2), we compared MUC5B expression in nonpregnant uterine tissues between 15 women with a history of cervical excisional surgery and 26 women without a history of cervical surgery. RESULTS: The abundance of MUC5B in cervicovaginal fluid was significantly decreased in the preterm birth group (fold change = 0.41, p = .035). Among the 480 quantified proteins, MUC5B had the second highest positive correlation with gestational age at delivery in the combined preterm and term groups. The cervicovaginal microbiome composition was not significantly different between the two groups. Cervical length was not correlated with gestational age at delivery (r = 0.18, p = .079). Histologically, the MUC5B-positive area in the nonpregnant cervix was significantly decreased in women with a history of cervical excisional surgery (0.85-fold, p = .048). The distribution of MUC5B-positive areas in the cervical tissues of 26 women without a history of cervical excisional surgery differed across individuals. CONCLUSIONS: This study suggests that the primary mechanism by which cervical excisional surgery causes preterm birth is the hyposecretion of MUC5B due to loss of the cervical glands.

Vaginal Microbiota and Pregnancy Outcomes of Patients with Conization Histories.

Background: One of the major risks of preterm birth is a history of conization. However, the risk of infection due to this procedure is still not well known. Using next-generation sequencing, we aimed to reveal the influence of conization on vaginal microbiota in the following pregnancy, and their relationship between spontaneous preterm birth (sPTB). Methods: We conducted a prospective cohort study including 133 pregnant patients, of whom 25 had conization histories and 108 did not. Vaginal microbiome samples were collected using swabs by an obstetrician upon inclusion in the first trimester and during delivery. V1-V2 of the 16S rRNA gene were amplified and analyzed to identify the bacteria. Results: The conization group had a significantly lower delivery week (34 weeks vs. 36 weeks, p = 0.003) and higher sPTB rate (64% vs. 8.3%, p ≤ 0.001) than the control group. In the conization group, alpha (Chao 1, p = 0.02; phylogenetic diversity whole tree, p = 0.04) and beta diversity (permutational multivariate analysis of variance test, p = 0.04) of the vaginal microbiota was significantly higher during delivery in patients who delivered preterm than in those who delivered term. Community-state type IV in the first trimester was significantly associated with sPTB (overall odds ratio 3.80, 95% confidence interval 1.33-10.8, p = 0.01). Conclusions: Conization is a risk factor for sPTB. Increased risk of sPTB in patients after conization may belong to the vulnerable defense mechanism, due to the shortened cervix and decreased cervical mucus.

Two target gene activation pathways for orphan ERR nuclear receptors.

Estrogen-related receptors (ERRα/ß/γ) are orphan nuclear receptors that function in energy-demanding physiological processes, as well as in development and stem cell maintenance, but mechanisms underlying target gene activation by ERRs are largely unknown. Here, reconstituted biochemical assays that manifest ERR-dependent transcription have revealed two complementary mechanisms. On DNA templates, ERRs activate transcription with just the normal complement of general initiation factors through an interaction of the ERR DNA-binding domain with the p52 subunit of initiation factor TFIIH. On chromatin templates, activation by ERRs is dependent on AF2 domain interactions with the cell-specific coactivator PGC-1α, which in turn recruits the ubiquitous p300 and MED1/Mediator coactivators. This role of PGC-1α may also be fulfilled by other AF2-interacting coactivators like NCOA3, which is shown to recruit Mediator selectively to ERRß and ERRγ. Importantly, combined genetic and RNA-seq analyses establish that both the TFIIH and the AF2 interaction-dependent pathways are essential for ERRß/γ-selective gene expression and pluripotency maintenance in embryonic stem cells in which NCOA3 is a critical coactivator.

The 3' Pol II pausing at replication-dependent histone genes is regulated by Mediator through Cajal bodies' association with histone locus bodies.

Non-polyadenylated mRNAs of replication-dependent histones (RDHs) are synthesized by RNA polymerase II (Pol II) at histone locus bodies (HLBs). HLBs frequently associate with Cajal bodies (CBs), in which 3'-end processing factors for RDH genes are enriched; however, this association's role in transcription termination of RDH genes remains unclear. Here, we show that Pol II pauses immediately upstream of transcript end sites of RDH genes and Mediator plays a role in this Pol II pausing through CBs' association with HLBs. Disruption of the Mediator docking site for Little elongation complex (LEC)-Cap binding complex (CBC)-Negative elongation factor (NELF), components of CBs, interferes with CBs' association with HLBs and 3' Pol II pausing, resulting in increased aberrant unprocessed RDH gene transcripts. Our findings suggest Mediator's involvement in CBs' association with HLBs to facilitate 3' Pol II pausing and subsequent 3'-end processing of RDH genes by supplying 3'-end processing factors.

Conceptual design of the Hybrid Ring with superconducting linac.

The Hybrid Ring with a superconducting-linac injector as a highly flexible synchrotron radiation source to enable new experimental techniques and enhance many existing ones is proposed. It is designed to be operated with the coexistence of the storage (SR) bunches characterized by the performance of the storage ring, and the single-pass (SP) bunches characterized by the performance of the superconducting linac. Unique experiments can be performed by simultaneous use of the SR and SP beams, in addition to research with various experimental techniques utilizing the versatile SR beam and research in the field of ultrafast dynamics utilizing the ultrashort pulse of the SP beam. The extendability of the Hybrid Ring will allow it to be developed into a synchrotron radiation complex.

Construction and commissioning of mid-infrared self-amplified spontaneous emission free-electron laser at compact energy recovery linac.

The mid-infrared range is an important spectrum range where materials exhibit a characteristic response corresponding to their molecular structure. A free-electron laser (FEL) is a promising candidate for a high-power light source with wavelength tunability to investigate the nonlinear response of materials. Although the self-amplification spontaneous emission (SASE) scheme is not usually adopted in the mid-infrared wavelength range, it may have advantages such as layout simplicity, the possibility of producing a single pulse, and scalability to a short-wavelength facility. To demonstrate the operation of a mid-infrared SASE FEL system in an energy recovery linac (ERL) layout, we constructed an SASE FEL setup in cERL, a test facility of the superconducting linac with the ERL configuration. Despite the adverse circumstance of space charge effects due to the given boundary condition of the facility, we successfully established the beam condition at the undulators and observed FEL emission at a wavelength of 20 µm. The results show that the layout of cERL has the potential for serving as a mid-infrared light source.

Sumoylation of the human histone H4 tail inhibits p300-mediated transcription by RNA polymerase II in cellular extracts.

The post-translational modification of histones by the small ubiquitin-like modifier (SUMO) protein has been associated with gene regulation, centromeric localization, and double-strand break repair in eukaryotes. Although sumoylation of histone H4 was specifically associated with gene repression, this could not be proven due to the challenge of site-specifically sumoylating H4 in cells. Biochemical crosstalk between SUMO and other histone modifications, such as H4 acetylation and H3 methylation, that are associated with active genes also remains unclear. We addressed these challenges in mechanistic studies using an H4 chemically modified at Lys12 by SUMO-3 (H4K12su) and incorporated into mononucleosomes and chromatinized plasmids for functional studies. Mononucleosome-based assays revealed that H4K12su inhibits transcription-activating H4 tail acetylation by the histone acetyltransferase p300, as well as transcription-associated H3K4 methylation by the extended catalytic module of the Set1/COMPASS (complex of proteins associated with Set1) histone methyltransferase complex. Activator- and p300-dependent in vitro transcription assays with chromatinized plasmids revealed that H4K12su inhibits both H4 tail acetylation and RNA polymerase II-mediated transcription. Finally, cell-based assays with a SUMO-H4 fusion that mimics H4 tail sumoylation confirmed the negative crosstalk between histone sumoylation and acetylation/methylation. Thus, our studies establish the key role for histone sumoylation in gene silencing and its negative biochemical crosstalk with active transcription-associated marks in human cells.

Immunoglobulin free light chains as an inflammatory biomarker of heart failure with myocarditis.

BACKGROUND: In this study, we measured immunoglobulin free light chains (FLC), a biomarker of inflammation in the sera of patients with heart failure due to myocarditis. METHODS: FLC kappa and FLC lambda were assayed in stored serum samples from patients with heart failure with myocarditis from the US myocarditis treatment trial by a competitive-inhibition multiplex Luminex® assay. RESULTS: The median concentration of circulating FLC kappa/lambda ratio was significantly lower in the sera from patients with heart failure with myocarditis than in healthy controls, and FLC kappa/lambda ratio had good diagnostic ability for identification of heart failure with myocarditis. Further, FLC kappa/lambda ratio was an independent prognostic factor for overall survival, and allowed creation of three prognostic groups by combining with N-terminal pro-B-type natriuretic peptide. CONCLUSIONS: This study suggests that FLC kappa/lambda ratio is a promising biomarker of heart failure with myocarditis.

Correction to: Immunoglobulin free light chains: an inflammatory biomarker of diabetes.

In the original publication of this paper contains some typographical errors in Table 1.

Immunoglobulin free light chains: an inflammatory biomarker of diabetes.

OBJECTIVE: Inflammation is increasingly understood as playing an important role in type 2 diabetes mellitus (T2D) development. A critical mechanism of the inflammatory cascade in developing T2D is nuclear factor-kappa B (NF-kB) activation. As immunoglobulin free light chains (FLC) could be a biomarker of activation of NF-kB, we measured FLC in patients with T2D. SUBJECTS: The age range of the 77 patients with T2D and the 75 healthy control participants were 45-87 years (median 60) and 25-72 years (median 51), respectively. METHODS: Serum FLC kappa and lambda were assayed by a competitive-inhibition multiplex Luminex assay. RESULTS: The concentration of circulating FLC the kappa/lambda ratio was lower in patients with T2D than in healthy volunteers. The area under the receiver operating curve (ROC-AUC) of the FLC kappa/lambda ratio showed the largest ROC-AUC compared with other FLC variables and hemoglobin A1c (HbA1c). The diagnostic performance for distinguishing between T2D and healthy control was a sensitivity of 0.96 and a specificity of 1. The odds ratio was 0.000018. CONCLUSIONS: These results suggest that FLC kappa/lambda may be more specific and sensitive for the diagnosis of T2D than HbA1c, and thus represents a potentially promising biomarker of inflammation.

Shank2 Binds to aPKC and Controls Tight Junction Formation with Rap1 Signaling during Establishment of Epithelial Cell Polarity.

Epithelial cells establish apicobasal polarity by forming tight junctions (TJs) at the apical-lateral boundary, which play fundamental roles in physiological functions. An evolutionarily conserved atypical protein kinase C (aPKC)-partitioning defective (PAR) complex functions as a platform for TJ assembly during cell polarity establishment. However, how this complex converts the spatial cues into a subsequent active unit is unclear. Here, we identify an epithelial isoform of Shank2 as a mediator of the aPKC-PAR complex. Shank2 binds to and colocalizes with aPKC at apical junctional regions of polarized epithelial cells. Shank2 knockdown results in defects in TJ formation. Mechanistically, we find that the N-terminal SPN domain is required for the junctional localization of Shank2 and binds to the active form of Rap1 small GTPase, which is involved in TJ formation. Our findings suggest that a close physical and functional relationship between aPKC and Shank2-active Rap1 signaling serves as the platform for TJ assembly to regulate epithelial cell polarity.

E2A-PBX1 functions as a coactivator for RUNX1 in acute lymphoblastic leukemia.

E2A, a basic helix-loop-helix transcription factor, plays a crucial role in determining tissue-specific cell fate, including differentiation of B-cell lineages. In 5% of childhood acute lymphoblastic leukemia (ALL), the t(1,19) chromosomal translocation specifically targets the E2A gene and produces an oncogenic E2A-PBX1 fusion protein. Although previous studies have shown the oncogenic functions of E2A-PBX1 in cell and animal models, the E2A-PBX1-enforced cistrome, the E2A-PBX1 interactome, and related mechanisms underlying leukemogenesis remain unclear. Here, by unbiased genomic profiling approaches, we identify the direct target sites of E2A-PBX1 in t(1,19)-positive pre-B ALL cells and show that, compared with normal E2A, E2A-PBX1 preferentially binds to a subset of gene loci cobound by RUNX1 and gene-activating machineries (p300, MED1, and H3K27 acetylation). Using biochemical analyses, we further document a direct interaction of E2A-PBX1, through a region spanning the PBX1 homeodomain, with RUNX1. Our results also show that E2A-PBX1 binding to gene enhancers is dependent on the RUNX1 interaction but not the DNA-binding activity harbored within the PBX1 homeodomain of E2A-PBX1. Transcriptome analyses and cell transformation assays further establish a significant RUNX1 requirement for E2A-PBX1-mediated target gene activation and leukemogenesis. Notably, the RUNX1 locus itself is also directly activated by E2A-PBX1, indicating a multilayered interplay between E2A-PBX1 and RUNX1. Collectively, our study provides the first unbiased profiling of the E2A-PBX1 cistrome in pre-B ALL cells and reveals a previously unappreciated pathway in which E2A-PBX1 acts in concert with RUNX1 to enforce transcriptome alterations for the development of pre-B ALL.

Gene-Specific H1 Eviction through a Transcriptional Activator→p300→NAP1→H1 Pathway.

Linker histone H1 has been correlated with transcriptional inhibition, but the mechanistic basis of the inhibition and its reversal during gene activation has remained enigmatic. We report that H1-compacted chromatin, reconstituted in vitro, blocks transcription by abrogating core histone modifications by p300 but not activator and p300 binding. Transcription from H1-bound chromatin is elicited by the H1 chaperone NAP1, which is recruited in a gene-specific manner through direct interactions with activator-bound p300 that facilitate core histone acetylation (by p300) and concomitant eviction of H1 and H2A-H2B. An analysis in B cells confirms the strong dependency on NAP1-mediated H1 eviction for induction of the silent CD40 gene and further demonstrates that H1 eviction, seeded by activator-p300-NAP1-H1 interactions, is propagated over a CCCTC-binding factor (CTCF)-demarcated region through a distinct mechanism that also involves NAP1. Our results confirm direct transcriptional inhibition by H1 and establish a gene-specific H1 eviction mechanism through an activator→p300→NAP1→H1 pathway.

Stimulated Excitation of an Optical Cavity by a Multibunch Electron Beam via Coherent-Diffraction-Radiation Process.

With a low emittance and short-bunch electron beam at a high repetition rate realized by a superconducting linac, stimulated excitation of an optical cavity at the terahertz spectrum range is shown. The electron beam passes through small holes in the cavity mirrors without being destroyed. A sharp resonance structure which indicates wideband stimulated emission via coherent diffraction radiation is observed while scanning the round-trip length of the cavity.

p300-Mediated Lysine 2-Hydroxyisobutyrylation Regulates Glycolysis.

Lysine 2-hydroxyisobutyrylation (Khib) is an evolutionarily conserved and widespread histone mark like lysine acetylation (Kac). Here we report that p300 functions as a lysine 2-hyroxyisobutyryltransferase to regulate glycolysis in response to nutritional cues. We discovered that p300 differentially regulates Khib and Kac on distinct lysine sites, with only 6 of the 149 p300-targeted Khib sites overlapping with the 693 p300-targeted Kac sites. We demonstrate that diverse cellular proteins, particularly glycolytic enzymes, are targeted by p300 for Khib, but not for Kac. Specifically, deletion of p300 significantly reduces Khib levels on several p300-dependent, Khib-specific sites on key glycolytic enzymes including ENO1, decreasing their catalytic activities. Consequently, p300-deficient cells have impaired glycolysis and are hypersensitive to glucose-depletion-induced cell death. Our study reveals an p300-catalyzed, Khib-specific molecular mechanism that regulates cellular glucose metabolism and further indicate that p300 has an intrinsic ability to select short-chain acyl-CoA-dependent protein substrates.

A UTX-MLL4-p300 Transcriptional Regulatory Network Coordinately Shapes Active Enhancer Landscapes for Eliciting Transcription.

Enhancer activation is a critical step for gene activation. Here we report an epigenetic crosstalk at enhancers between the UTX (H3K27 demethylase)-MLL4 (H3K4 methyltransferase) complex and the histone acetyltransferase p300. We demonstrate that UTX, in a demethylase activity-independent manner, facilitates conversion of inactive enhancers in embryonic stem cells to an active (H3K4me1+/H3K27ac+) state by recruiting and coupling the enzymatic functions of MLL4 and p300. Loss of UTX leads to attenuated enhancer activity, characterized by reduced levels of H3K4me1 and H3K27ac as well as impaired transcription. The UTX-MLL4 complex enhances p300-dependent H3K27 acetylation through UTX-dependent stimulation of p300 recruitment, while MLL4-mediated H3K4 monomethylation, reciprocally, requires p300 function. Importantly, MLL4-generated H3K4me1 further enhances p300-dependent transcription. This work reveals a previously unrecognized cooperativity among enhancer-associated chromatin modulators, including a unique function for UTX, in establishing an "active enhancer landscape" and defines a detailed mechanism for the joint deposition of H3K4me1 and H3K27ac.