However, detrimental actions of interleukin-1 and other pro-inflammatory cytokines in infected placentas are involved in adverse neonatal outcomes, suggesting that tightly regulated immune responses are crucial to sustain placenta homeostasis.208 Besides, human trophoblasts-associated antiviral microRNAs, such as chromosome 19 miRNA cluster that are packaged into placental exosomes, were also systemically isolated from CENPA pregnant women, which operate in a paracrine or autocrine manner to resist infection.209,210 Mechanistically, primary human trophoblast-derived chromosome 19 miRNA cluster family members drastically 5(6)-Carboxyfluorescein limited both RNA and DNA viral infections in non-placental cells by inducing autophagy, demonstrating a unique placenta-secreted effector for shielding virus-sensitive cells in placenta from infections.198,211 The neonatal fragment crystallizable receptor (FcRn) mediated antibodies transplacental transfer Transplacental passage of IgG begins in the first trimester of pregnancy and lasts until labor, which sets up another layer of fetal protection from viral infections.212 Generally, endocytosis of IgG from maternal blood was initiated by binding to a canonical IgG shuttle receptor, FcRn, around the apical side of STBs. fight the next emerging and re-emerging infectious disease in the pregnancy populace. family, is the most common blood-borne pathogen globally, which could lead to acute and chronic 5(6)-Carboxyfluorescein hepatitis in humans. The transmission routes of HBV are predominantly through blood and bodily fluids, vertical transmission, as well as sexual and parenteral contacts.9 Perinatal transmission from the mother to fetus or newborns is still responsible for the most chronic HBV infections in adults who are more prone to severe liver diseases and poor responses to antiviral therapies.10,11 The risk of perinatal transmission in case of mothers with positive HBV e antigen or high viral loads has been estimated to be as high as 90% if the newborns accept no immunoprophylaxis treatment (includes HBV vaccine and immune globulin).12 Therefore, to prevent the vertical transmission of HBV in advance, universal maternal screenings for the HBV surface antigen, HBV e antigen, viral load, and alanine aminotransferase level during pregnancy are priorities to be adopted.12 Although immunoprophylaxis at birth together with antiviral treatments for mothers in endemic areas currently are the common and effective strategies for global elimination and preventive interventions of HBV, vertical transmission of HBV occurs with high prevalence and should be taken seriously due to uneven coverage of vaccine globally and/or prophylaxis failure.13C15 Although pregnancy complications related to HBV infection if any are minimal, clinical evidence has indicated that chronic HBV infection may be vaguely associated with gestational diabetes, preterm labor, antepartum hemorrhage, and preeclampsia.16C18 For preterm birth, several meta-analyses have confirmed that seropositivity for HBV surface antigen in pregnant women could increase the risk of preterm labor, while another study involving 6781 prematurity cases inconsistently revealed no association with HBV contamination in the preterm birth group.17,19C22 Worthy of note, in the above-mentioned studies, half of the enrolled pregnant women exhibited abnormal liver functions such as nonalcoholic fatty liver disease, which may be an independent risk factor for preterm labor rather than the computer virus per se. Interestingly, women with HBV contamination were observed to develop 2.18-fold higher antepartum hemorrhage, probably due to placenta previa and placental abruption, which is attributed to coinfection of HBV with other viruses.21 Unexpectedly, a negative association or protective effect of HBV contamination on preeclampsia was demonstrated in a meta-analysis involving 11,566 cases.23 Nonetheless, the explicit causes underlying the above adverse pregnancy outcomes have not been extensively evaluated until now, placental inflammation, insulin resistance, increased 5(6)-Carboxyfluorescein immunotolerance, or impaired immune response upon HBV infection were proposed as suspected mechanisms.18,24 Furthermore, the hint of fetal and neonatal anomalies was also observed in pregnant women with chronic infection. It was noticed that 60% increased non-reassuring fetal heart rate patterns and 80% increase in asphyxia referring to 7600 pregnant HBV carriers from 18 studies by meta-analysis suggested fetal distress conditions related to HBV contamination.25,26 Additionally, 25.8% increased low birth weight and small infants were reported to be associated with HBV infection, while abnormally enhanced fetal growth and macrosomia were also found in a series of researches.26C28 Viral genotypes, co-existing hepatic disorders, coinfections with other pathogenic organisms, synergism with pregnancy complications, and the phase of chronic HBV infection probably led to contradictory phenotypes of fetal growth.29,30 The path of HBV vertical transmission includes intrauterine transmission, labor, and delivery, as well as breastfeeding. The primary risk period for infant HBV contamination is the peripartum period. Most cases of contamination occur during delivery when the mucosa of newborns is usually easily contaminated by maternal blood and secretions that contain.

Notably, in a KO, S263A had no additional reduction in NHEJ repair efficiency, but S263E recovered the KO NHEJ defect (Figure ?(Figure6D).6D). NHEJ efficiency, and similarly, PRKACB was found to phosphorylate XLF (a Nej1 human homolog) at S263, a corresponding residue of the yeast Nej1 S298. Together, our results uncover a new and conserved mechanism for Tpk1 and PRKACB in phosphorylating Nej1 (or XLF), which is critically required for NHEJ repair. Graphical Abstract Open in a separate windows Graphical Abstract Cellular model of yeast Tpk1 role with Nej1 and PRKACBwith XLF on NHEJ, as well as the loss of tpk1 in DNA resection and MMEJ. INTRODUCTION DNA double-stranded breaks (DSBs) are the most severe forms of DNA damage, causing genomic instability and chromosomal rearrangements (1), leading to serious human diseases including cancer and immunodeficiency response (2,3). In response to DSBs, eukaryotic cells recruit three repair pathways: homologous recombination (HR), non-homologous end joining (NHEJ), and microhomology-mediated end joining (MMEJ) to rescue genomic instability. NHEJ is the main DSB repair pathway in mammalian cells, and highly conserved in eukaryotes ranging from the budding yeast, genes, and one regulatory subunit, encoded by gene (15,16). Components of the PKA catalytic subunit trimeric complex (Tpk1-3) exhibit varied substrate specificities (17), indicating the potential for distinct functional profiles. A yeast proteome chip screening approach (17) has uncovered Nej1 as a phosphorylation substrate of Tpk1, but not Tpk2 SJG-136 or Tpk3, implying a likely SJG-136 role for Tpk1 in DNA repair. Our recent work has shown deletion impaired NHEJ (18), and that it is epistatic with a yeast uncharacterized gene, (HU resistance 1) in NHEJ, though the molecular mechanism of this connection in DNA repair remains unclear (19). While this and other evidence suggests the role of yeast SJG-136 PKA in DNA damage checkpoint pathways (13), and Tpk1 in NHEJ (18,19), how PKA subunits impact these processes, and whether there is a conserved role between yeast and human in DSB repair pathway, warrants further investigation. In the present study, we show that deletion of and (or the checkpoint kinase, kinase activity dependent on Nej1 serine residue at S298, a phosphosite of Dun1, suggesting a crosstalk between the two DNA damage checkpoint kinases (Tpk1, Dun1) through SJG-136 shared phosphorylation of the Nej1 S298. As well, the repair defect of deletion can be recovered by mutation of Nej1 S298E (phosphomimetic), but not Nej1 S298A (non-phosphorylatable), consistent with the phosphorylation of S298 being a requisite for Tpk1s role in NHEJ repair. As in yeast, PRKACB, a human homolog of Tpk1, showed comparable NHEJ defect in DSB repair by phosphorylating the Nej1 human homolog, XLF at S263, suggesting a conserved NHEJ role for Tpk1 in eukaryotes. Overall, our findings suggest that Nej1 phosphorylation by Tpk1 is usually intrinsic to NHEJ break repair and resolution, and spotlight a conserved model for NHEJ regulation. MATERIALS AND METHODS Besides the procedures described below, site-directed mutagenesis, kinase and phosphorylation-induced mobility shift, sensitivity to DNA damage, clonogenic survival, cell proliferation, phosphoproteomics/affinity purification coupled with mass spectrometry (MS), immunoblotting, purification of SJG-136 yeast Tpk1 and Nej1 recombinant proteins, generation of stable CRISPR gene knockouts (KOs), immunofluorescence staining and quantification of DNA damage foci, and alkaline comet assay, among others are detailed in Supplementary Methods. Mutant strains or CRISPR gene KOs, cell cultures and plasmids All yeast gene deletions (or mammalian gene KOs), plasmids, primers, and antibodies used are listed in Supplementary Table S1. Yeast Mouse monoclonal to CD54.CT12 reacts withCD54, the 90 kDa intercellular adhesion molecule-1 (ICAM-1). CD54 is expressed at high levels on activated endothelial cells and at moderate levels on activated T lymphocytes, activated B lymphocytes and monocytes. ATL, and some solid tumor cells, also express CD54 rather strongly. CD54 is inducible on epithelial, fibroblastic and endothelial cells and is enhanced by cytokines such as TNF, IL-1 and IFN-g. CD54 acts as a receptor for Rhinovirus or RBCs infected with malarial parasite. CD11a/CD18 or CD11b/CD18 bind to CD54, resulting in an immune reaction and subsequent inflammation FLAG or HA-tagged fusion proteins and mutant strains were created in BY4741 or JKM139 background by lithium acetate transformation (20). Deletions were confirmed by PCR and/or DNA sequencing across the deletion site. Yeast strains were produced in YPD (1% yeast extract, 2% bacto-peptone, 2% glucose) at 30C, unless otherwise noted. Human osteosarcoma U2OS and HEK293T cells were cultured in Dulbecco’s altered Eagle’s medium supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin mix at 37C and 5% CO2. Stable human gene KOs were created using a CRISPR-Cas9 system with LentiCRISPR-v2 blasticidin or puromycin, following established procedure (21). NHEJ.