Maintenance Of Integrity Research Articles

Enhanced targeted treatment of cervical cancer using nanoparticle-based doxycycline delivery system

This study investigates a nanoparticle-based doxycycline (DOX) delivery system targeting cervical cancer cells via the CD44 receptor. Molecular docking revealed a strong binding affinity between hyaluronic acid (HA) and CD44 (binding energy: -7.2 kJ/mol). Characterization of the HA-Chitosan nanoparticles showed a particle size of 284.6 nm, a zeta potential of 16.9 mV, and a polydispersity index of 0.314, with SEM confirming smooth surface morphology. The encapsulation efficiency of DOX-loaded nanoparticles was 89.32%, exhibiting a sustained release profile, with 67.45% released over 72 h in acidic conditions (pH 5.5). Cytotoxicity assays demonstrated a significant reduction in HeLa cell viability to 22% at 72 h, compared to 67% in normal HEK cells. Stability tests confirmed the maintenance of nanoparticle integrity and a consistent drug release profile over three months. Cell migration was reduced by 45%, and RT-PCR analysis revealed a 53% downregulation of TNF-α expression, suggesting effective targeting of inflammatory pathways. These results underscore the potential of HA-Chitosan-based DOX nanoparticles in improving cervical cancer treatment through enhanced targeted delivery and inhibition of tumor-promoting mechanisms.

Gut Microbiota-Based Interventions for Parkinson's Disease: Neuroprotective Mechanisms and Current Perspective.

Recent evidence links gut microbiota alterations to neurodegenerative disorders, including Parkinson's disease (PD). Replenishing the abnormal composition of gut microbiota through gut microbiota-based interventions "prebiotics, probiotics, synbiotics, postbiotics, and fecal microbiota transplantation (FMT)" has shown beneficial effects in PD. These interventions increase gut metabolites like short-chain fatty acids (SCFAs) and glucagon-like peptide-1 (GLP-1), which may protect dopaminergic neurons via the gut-brain axis. Neuroprotective effects of these interventions are mediated by several mechanisms, including the enhancement of neurotrophin and activation of the PI3K/AKT/mTOR signaling pathway, GLP-1-mediated gut-brain axis signaling, Nrf2/ARE pathway, and autophagy. Other pathways, such as free fatty acid receptor activation, synaptic plasticity improvement, and blood-brain and gut barrier integrity maintenance, also contribute to neuroprotection. Furthermore, the inhibition of the TLR4/NF-кB pathway, MAPK pathway, GSK-3β signaling pathway, miR-155-5p-mediated neuroinflammation, and ferroptosis could account for their protective effects. Clinical studies involving gut microbiota-based interventions have shown therapeutic benefits in PD patients, particularly in improving gastrointestinal dysfunction and some neurological symptoms. However, the effectiveness in alleviating motor symptoms remains mild. Large-scale clinical trials are still needed to confirm these findings. This review emphasizes the neuroprotective mechanisms of gut microbiota-based interventions in PD as supported by both preclinical and clinical studies.

Extensive homologous recombinationsafeguards oocyte genome integrity in mammals.

Meiosis in mammalian oocytes is interrupted by a prolonged arrest at the germinal vesiclestage, during which oocytes have to repair DNA lesions to ensure genome integrityor otherwise undergo apoptosis. The FIRRM/FLIP-FIGNL1 complex dissociates RAD51 from the joint DNA molecules in both homologous recombination (HR) and DNA replication. However, as a type of non-meiotic, non-replicative cells, whether this RAD51-dismantling mechanism regulates genome integrity in oocytes remains elusive. Here, we show that FIRRM/FLIP is required for disassembly of RAD51-filaments and maintenance of genome integrity in oocytes. Deletion of FIRRM in oocytes leads to formation of massive nuclear RAD51 foci in oocytes of primordial follicles and activated follicles in mice. These RAD51 foci colocalize with the sites of DNA damage repair, as indicated by RPA2 and EdU, suggesting substantial DNA damage and extensive HR in oocytes. Especially in fully-grown FIRRM-deleted oocytes, RAD51 forms a net-like structure. As a consequence, FIRRM-deleted females are infertile due to aberrant homologous chromosome segregation at metaphase I and primordial follicle insufficiency at young adulthood. Hence, our study demonstrates the physiological importance of HR in maintaining genome integrity in oocytes.

Linear ubiquitination at damaged lysosomes induces local NFKB activation and controls cell survival

ABSTRACT Lysosomes are the major cellular organelles responsible for nutrient recycling and degradation of cellular material. Maintenance of lysosomal integrity is essential for cellular homeostasis and lysosomal membrane permeabilization (LMP) sensitizes toward cell death. Damaged lysosomes are repaired or degraded via lysophagy, during which glycans, exposed on ruptured lysosomal membranes, are recognized by galectins leading to K48- and K63-linked poly-ubiquitination (poly-Ub) of lysosomal proteins followed by recruitment of the macroautophagic/autophagic machinery and degradation. Linear (M1) poly-Ub, catalyzed by the linear ubiquitin chain assembly complex (LUBAC) E3 ligase and removed by OTULIN (OTU deubiquitinase with linear linkage specificity) exerts important functions in immune signaling and cell survival, but the role of M1 poly-Ub in lysosomal homeostasis remains unexplored. Here, we demonstrate that L-leucyl-leucine methyl ester (LLOMe)-damaged lysosomes accumulate M1 poly-Ub in an OTULIN- and K63 Ub-dependent manner. LMP-induced M1 poly-Ub at damaged lysosomes contributes to lysosome degradation, recruits the NFKB (nuclear factor kappa B) modulator IKBKG/NEMO and locally activates the inhibitor of NFKB kinase (IKK) complex to trigger NFKB activation. Inhibition of lysosomal degradation enhances LMP- and OTULIN-regulated cell death, indicating pro-survival functions of M1 poly-Ub during LMP and potentially lysophagy. Finally, we demonstrate that M1 poly-Ub also occurs at damaged lysosomes in primary mouse neurons and induced pluripotent stem cell-derived primary human dopaminergic neurons. Our results reveal novel functions of M1 poly-Ub during lysosomal homeostasis, LMP and degradation of damaged lysosomes, with important implications for NFKB signaling, inflammation and cell death. Abbreviation: ATG: autophagy related; BafA1: bafilomycin A1; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CRISPR: clustered regularly interspaced short palindromic repeats; CHUK/IKKA: component of inhibitor of nuclear factor kappa B kinase complex; CUL4A-DDB1-WDFY1: cullin 4A-damage specific DNA binding protein 1-WD repeat and FYVE domain containing 1; DGCs: degradative compartments; DIV: days in vitro; DUB: deubiquitinase/deubiquitinating enzyme; ELDR: endo-lysosomal damage response; ESCRT: endosomal sorting complex required for transport; FBXO27: F-box protein 27; GBM: glioblastoma multiforme; IKBKB/IKKB: inhibitor of nuclear factor kappa B kinase subunit beta; IKBKG/NEMO: inhibitor of nuclear factor kappa B kinase regulatory subunit gamma; IKK: inhibitor of NFKB kinase; iPSC: induced pluripotent stem cell; KBTBD7: kelch repeat and BTB domain containing 7; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LCD: lysosomal cell death; LGALS: galectin; LMP: lysosomal membrane permeabilization; LLOMe: L-leucyl-leucine methyl ester; LOP: loperamide; LUBAC: linear ubiquitin chain assembly complex; LRSAM1: leucine rich repeat and sterile alpha motif containing 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; MTORC1: MTOR complex 1; NBR1: NBR1 autophagy cargo receptor; NFKB/NF-κB: nuclear factor kappa B; NFKBIA/IĸBα: nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor alpha; OPTN: optineurin; ORAS: OTULIN-related autoinflammatory syndrome; OTULIN: OTU deubiquitinase with linear linkage specificity; RING: really interesting new gene; RBR: RING-in-between-RING; PLAA: phospholipase A2 activating protein; RBCK1/HOIL-1: RANBP2-type and C3HC4-type zinc finger containing 1; RNF31/HOIP: ring finger protein 31; SHARPIN: SHANK associated RH domain interactor; SQSTM1/p62: sequestosome 1; SR-SIM: super-resolution-structured illumination microscopy; TAX1BP1: Tax1 binding protein 1; TBK1: TANK binding kinase 1; TH: tyrosine hydroxylase; TNF/TNFα: tumor necrosis factor; TNFRSF1A/TNFR1-SC: TNF receptor superfamily member 1A signaling complex; TRIM16: tripartite motif containing 16; Ub: ubiquitin; UBE2QL1: ubiquitin conjugating enzyme E2 QL1; UBXN6/UBXD1: UBX domain protein 6; VCP/p97: valosin containing protein; WIPI2: WD repeat domain, phosphoinositide interacting 2; YOD1: YOD1 deubiquitinase.

Odorant production surges induced by exogenous oxidative stress: An overlooked risk in PAA-based moderate preoxidation for algal-laden water.

Moderate preoxidation is feasible for odor-producing algae treatments, usually requiring trade-offs in algal removal and integrity maintenance. However, dosing oxidants may cause internal oxidative homeostasis imbalances and secondary odorous metabolite responses, adding new trade-offs for moderate treatments. Peracetic acid (PAA)/Fe processes are promising strategies in moderate treatments and thus were applied to examine how to achieve the following three trade-offs: good algal removal, no odorant increases and no releases. For algal removal, the highest removal efficiency was 71.1 %, 87.0 % and 98.1 %, respectively, in PAA/Fe(Ⅲ) separate, PAA/Fe(Ⅱ) separate and PAA/Fe(Ⅱ) simultaneous process. Odorant responses always followed a pattern of "promotion-low dosages, inhibition-high dosages", with the highest production reaching 3.91-fold of the control without PAA addition, well above odorant threshold concentrations (500 ng/L). Instant releases did not occur, yet with delayed releases observed during sludge retention in sedimentation tanks (5, 16 h). It was verified that exogenous oxidative stress stimulated odorant generation with multiple reactive species. Among them, PAA oxidant was crucial in algal removal and odorant promotion; R-O· and ·OH caused odorant generation and degradation, respectively. Overall, PAA/Fe(Ⅱ) processes were more suitable for moderate treatments, simultaneously avoiding odorant increases and allowing sludge retention. This work provides a novel perspective for moderate preoxidation, highlighting instant odorant generation and delayed releases triggered by exogenous PAA stressors.

Ki-67 and CDK1 control the dynamic association of nuclear lipids with mitotic chromosomes.

Nuclear lipids play roles in regulatory processes, such as signaling, transcriptional regulation, and DNA repair. In this report, we demonstrate that nuclear lipids may contribute to Ki-67-regulated chromosome integrity during mitosis. In COS-7cells, nuclear lipids are enriched at the perichromosomal layer and excluded from intrachromosomal regions during early mitosis but are then detected in intrachromosomal regions during late mitosis, as revealed by TT-ExM (expansion microscopy with trypsin digestion and tyramide signal amplification), an improved expansion microscopy technique that enables high-sensitivity and super-resolution imaging of proteins, lipids, and nuclear DNA. The nuclear nonhistone protein Ki-67 acts as a surfactant to form a repulsive molecular brush around fully condensed sister chromatids in early mitosis, preventing the diffusion or penetration of nuclear lipids into intrachromosomal regions. Ki-67 is phosphorylated during mitosis by cyclin-dependent kinase 1 (CDK1), the best-known master regulator of the cell cycle. Both Ki-67 knockdown and reduced Ki-67 phosphorylation by CDK1 inhibitors allow nuclear lipids to penetrate chromosomal regions. Thus, both Ki-67 protein level and phosphorylation status during mitosis appear to influence the perichromosomal distribution of nuclear lipids. Ki-67 knockdown and CDK1 inhibition also lead to uneven chromosome disjunction between daughter cells, highlighting the critical role of this regulatory mechanism in ensuring accurate chromosome segregation. Given that Ki-67 has been proposed to promote chromosome individualization and establish chromosome-cytoplasmic compartmentalization during open mitosis in vertebrates, our results reveal that nuclear lipid enrichment at the perichromosomal layer enhances the ability of Ki-67 to form a protective perichromosomal barrier (chromosome envelope), which is critical for correct chromosome segregation and maintenance of genome integrity during mitosis.

Vitamin D receptor regulates methyltransferase like 14 to mitigate colitis-associated colorectal cancer.

Colitis-associated colorectal cancer (CAC), a serious complication of ulcerative colitis (UC), is associated with a poor prognosis. The vitamin D receptor (VDR) is recognized for its protective role in UC and CAC through the maintenance of intestinal barrier integrity and the regulation of inflammation. This study demonstrates a significant reduction in m6A-related genes, particularly methyltransferase like 14 (METTL14), in UC and CAC patients and identifies an association between METTL14 and VDR. In the azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced mouse model, vitamin D treatment increases METTL14 expression and reduces tumor burden, while Vdr-knockout mice exhibit lower METTL14 levels and increased tumorigenesis. In vitro, the VDR agonist calcipotriol upregulates METTL14 in NCM460 cells, with this effect attenuated by VDR knockdown. VDR knockdown in DLD-1 colon cancer cells decreases METTL14 expression and promotes proliferation, which is reversed by METTL14 overexpression. Mechanistic studies reveal that VDR regulates METTL14 expression via promoter binding, modulating key target genes such as SOX4, DROSH, and PHLPP2. This study highlights the role of the VDR-METTL14 axis as a protective mechanism in CAC and suggests its potential as a therapeutic target for preventing and treating CAC.

Exploring Student Perspectives on ChatGPT: Knowledge, Attitudes, Concerns, and Usage Patterns at Universiti Teknikal Malaysia Melaka

This study explores the prevalence and perspectives of ChatGPT usage among students at Universiti Teknikal Malaysia Melaka, encompassing their knowledge, attitudes, concerns, perceived ethics, and usage patterns. A total of 367 students from five faculties participated in the survey, which was conducted via Google Forms. The findings reveal a high level of awareness and familiarity with ChatGPT, with 87.2% of respondents having heard about it and 79.6% understanding its workings. However, significant concerns were identified, including the potential undermining of educational goals (mean = 3.90, SD = 0.95) and negative impacts on creative writing skills (mean = 3.46, SD = 0.91). Ethical apprehensions were also prominent, with 49.1% of students recognizing ethical or legal considerations and 64.8% expressing comfort in interacting with ChatGPT. Despite these concerns, the utility of ChatGPT in enhancing productivity and aiding academic tasks is acknowledged, evidenced by high mean scores for assistance in drafting essays (mean = 3.87, SD = 0.87) and resolving academic doubts (mean = 3.83, SD = 0.85). This study underscores the need for ethical guidelines and controlled integration of ChatGPT into educational practices to balance its benefits with the maintenance of academic integrity and critical thinking skills.

Clear Cell Adenocarcinoma of the Urinary Tract Primary to the Renal Pelvis: A Multi-institutional Clinicopathologic and Molecular Study of Five Patients.

Clear cell adenocarcinoma (CCA) of the urinary tract is a rare malignancy and tumors involving the renal pelvis are notably sparse in the literature, with only 5 other patients reported. We present 5 patients, 4 women, and 1 man, with CCA of the renal pelvis. The age at presentation ranged from 29 to 81 years. The tumor size ranged from 4.5 to 8.0 cm. Tumors exhibited shared morphologic and immunohistochemical features with CCA of the female genital tract and those originating in the bladder and urethra, including cells with large nuclei, prominent nucleoli, nuclear hobnailing, and scant clear cytoplasm. Common immunohistochemical findings included reactivity for PAX8, CK7, HNF1β, and Napsin-A. One of the tumors arose in the background of a mixed epithelial and stromal tumor. Another tumor occurred in a renal allograft and tumor cells were positive for the BK virus, demonstrated by SV40 immunohistochemistry. All tumors were negative for TFE3 and TFEB rearrangement and lacked TERT alterations. Follow-up was limited with no recurrence in 4 patients at a maximum of 20 months follow-up and 1 patient died of an unrelated cause at 25 months of follow-up. Next-generation sequencing analysis of all 5 CCAs revealed mutations within genes implicated in DNA damage repair and chromatin remodeling pathways, including ATM , BRCA1 , BRCA2, ARID1A, DICER1, SMAD4, NOTCH1 , and MYC amplification. These molecular findings underscore the dysregulation of fundamental cellular processes essential for genomic integrity maintenance.

Mitogen-activated protein (MAP) kinase signalling pathway VmMkh1-VmMkk1-VmSpm1 regulates cell wall integrity in Valsa mali

Apple Valsa canker disease, caused by Valsa mali Miyabe et Yamada, seriously endangers the healthy growth of apple trees. Mitogen-Activated Protein Kinase (MAPK) signaling pathway is an important pathway to transmit signals stimulated by environmental stress. In this study, we identify and functionally characterize MAPKKK VmMkh1, MAPKK VmMkk1 and MAPK VmSpm1. VmMkh1 and VmMkk1 positively regulate the phosphorylation of VmSpm1. The radial growth rate of the VmSpm1 deletion mutant was reduced by approximately 31%. There was no significant difference in growth rate between the VmMkh1 and VmMkk1 mutant and the wild-type. VmMkh1 hyphe branches into a curved shape. The VmMkh1, VmMkk1, and VmSpm1 deletion mutant produced fewer conidia than the wild-type strain at 20 days post inoculation. Moreover, the VmMkh1, VmMkk1, and VmSpm1 deletion mutant slows conidial germination. The hyphal growth of VmMkh1, VmMkk1, and VmSpm1 deletion mutants are significantly inhibited on media containing NaCl, KCl, sorbitol (high osmotic stresses). The hyphal growth of VmMkh1, VmMkk1, and VmSpm1 deletion mutants are significantly inhibited on media containing Congo red, CFW, SDS, and Lysing encymes (Cell wall stress agents). A looser distribution of spacers in VmMkh1, VmMkk1, and VmSpm1 deletion mutants compared with the wild-type strain. The size of lesions on apple fruits and branches inoculated with VmSpm1 deletion mutant showed a reduction of approximately 46% and 43%, respectively, after 9 dpi. Overall, our findings demonstrate that VmMkh1, VmMkk1, and VmSpm1 are involved in regulating the growth and development, colony surface hydrophobicity, osmotic stress, cell wall integrity maintenance, carbon and nitrogen source utilization, septa formation, and pathogenicity of Valsa mali.

Potent inhibitors of the human RNA ligase Rlig1 highlights its role in RNA integrity maintenance under oxidative cellular stress

Potent inhibitors of the human RNA ligase Rlig1 highlights its role in RNA integrity maintenance under oxidative cellular stress

Comprehensive Review of Biological Functions and Therapeutic Potential of Perilla Seed Meal Proteins and Peptides

This comprehensive review explores the biological functions of Perilla frutescens seed proteins and peptides, highlighting their significant potential for health and therapeutic applications. This review delves into the mechanisms through which perilla peptides combat oxidative stress and protect cells from oxidative damage, encompassing free radical scavenging, metal chelating, in vivo antioxidant, and cytoprotective activities. Perilla peptides exhibit robust anti-aging properties by activating the Nrf2 pathway, enhancing cellular antioxidant capacity, and supporting skin health through the promotion of keratinocyte growth, maintenance of collagen integrity, and reduction in senescent cells. Additionally, they demonstrate antidiabetic activity by inhibiting α-amylase and α-glucosidase. The cardioprotective effects of perilla peptides are underscored by ACE-inhibitory activities and combat oxidative stress through enhanced antioxidant defenses. Further, perilla peptides contribute to improved gut health by enhancing beneficial gut flora and reinforcing intestinal barriers. In liver, kidney, and testicular health, they reduce oxidative stress and apoptotic damage while normalizing electrolyte levels and protecting against cyclophosphamide-induced reproductive and endocrine disruptions by restoring hormone synthesis. Promising anticancer potential is also demonstrated by perilla peptides through the inhibition of key cancer cell lines, alongside their anti-inflammatory and immunomodulating activities. Their anti-fatigue effects enhance exercise performance and muscle function, while perilla seed peptide nanoparticles show potential for targeted drug delivery. The diverse applications of perilla peptides support their potential as functional food additives and therapeutic agents.

Overdue and Overdrawn: Internal Services, Administrative Debt, and the Government of Canada's Real Property File

AbstractAdministrative debt is when successive waves of reform result in an accumulation of rusting‐out systems that are not optimized. This occurs when the drivers for change prioritize expediency, offering quick fixes rather than a focus on long‐term sustainable re‐design. This is the case for the federal government's Internal Services and is illustrated through a look at the Real Property function over time. This function has been routinely discounted when priorities and budgets are set. Resulting capability deficits are exposed periodically in audits and as part of higher profile reviews revealing gaps in the maintenance of asset integrity that put public‐facing program delivery at risk. We make the case that Internal Services are inseparable from how governments deliver public‐facing external programs. We suggest ways to close the information gaps contributing to this overall administrative debt.

MRE11-independent effects of Mirin on mitochondrial DNA integrity and cellular immune responses.

Mirin, a chemical inhibitor of MRE11, has been recently reported to suppress immune response triggered by mitochondrial DNA (mtDNA) breakage and release during replication stalling. We show that while Mirin reduces mitochondrial replication fork breakage in mitochondrial 3´-exonuclease MGME1 deficient cells, this effect occurs independently of MRE11. We also discovered that Mirin directly inhibits cellular immune responses, as shown by its suppression of STAT1 phosphorylation in Poly(I:C)-treated cells. Furthermore, Mirin also altered mtDNA supercoiling and accumulation of hemicatenated replication termination intermediates - hallmarks of topoisomerase dysfunction - while mitigating topological changes induced by the overexpression of mitochondrial TOP3A, including TOP3A-dependent strand breakage at the non-coding region of mtDNA. Although Mirin does not seem to inhibit TOP3A activity in vitro, our findings demonstrate its MRE11-independent effects in cells and give insight into the mechanisms of the maintenance of mtDNA integrity.

The survival grip-how cell adhesion promotes tumor maintenance within the microenvironment.

Cell adhesion is warranted by proteins that are crucial for the maintenance of tissue integrity and homeostasis. Most of these proteins behave as receptors to link adhesion to the control of cell survival and their expression or regulation are often altered in cancers. B-cell malignancies do not evade this principle as they are sustained in relapsed niches by interacting with the microenvironment that includes cells and their secreted factors. Focusing on chronic lymphocytic leukemia and mantle cell lymphoma, this Review delves with the molecules involved in the dialog between the adhesion platforms and signaling pathways known to regulate both cell adhesion and survival. Current therapeutic strategies disrupt adhesive structures and compromise the microenvironment support to tumor cells, rendering them sensitive to immune recognition. The development of organ-on-chip and 3D culture systems, such as spheroids, have revealed the importance of mechanical cues in regulating signaling pathways to organize cell adhesion and survival. All these elements contribute to the elaboration of the crosstalk of lymphoma cells with the microenvironment and the education processes that allow the establishment of the supportive niche.

The Formyl Peptid Receptor Ligand Ac2-26 Improves the Integrity of the Blood−Brain Barrier in the Course of Pneumococcal Meningitis

Background: The brain is protected from invading pathogens by the blood−brain barrier (BBB) and the innate immune system. Pattern recognition receptors play a crucial role in detecting bacteria and initiating the innate immune response. Among these are G-protein-coupled formyl peptide receptors (FPR), which are expressed by immune cells in the central nervous system. In this study, we investigated the influence of the FPR ligand Ac2-26 on the integrity of the BBB during pneumococcal meningitis. Methods: Wild-type (WT) and Fpr1- and Fpr2-deficient mice were intrathecally infected with Streptococcus pneumoniae. Subsequently, different groups of mice were treated with intraperitoneal injections of Ac2-26. The integrity of the BBB was analyzed using various markers through immunohistochemistry and immunofluorescence. Results: The results showed reduced BBB integrity during the course of bacterial meningitis. Treatment with Ac2-26 in WT mice significantly prolonged the maintenance of BBB integrity. However, this effect was not observed in Fpr2-deficient mice. Conclusions: This study extends previous findings on the anti-inflammatory properties of Ac2-26 by demonstrating that Ac2-26 positively affects BBB integrity via FPR2 during pneumococcal meningitis. These findings suggest that further investigation of Ac2-26 and other FPR modulators as potential therapies for Streptococcus pneumoniae-induced meningitis is warranted.

It's all in the gut: the central role of the gut and microbiome in preventing disease progression in simian immunodeficiency viruses infected African nonhuman primates.

Typically, both HIV-infected humans and simian immunodeficiency virus (SIV)-infected Asian nonhuman primates (NHPs) eventually progress to AIDS, while African NHPs that are natural hosts of SIV do not, in spite of life-long, high levels of viral replication. Lack of disease progression in African NHPs is not due to some adaptation by the virus, but rather to host adaptations to the virus. Central to these adaptations is maintenance of the gut integrity during acute viral replication and inflammation, which allows natural hosts to avoid the chronic inflammation characteristic to pathogenic HIV/SIV infection. It has been recently shown that natural hosts of SIVs, such as the African green monkey (AGM), avoid damage to the mucosal epithelium through wound healing mechanisms, possibly with the contribution of a unique anti-inflammatory microbiome. Furthermore, these mechanisms are independent of viral replication, and CD4+ T-cell activation or depletion. Future SIV research on natural hosts should focus on further elucidating the anti-inflammatory state of their gut, and the role of microbiome/dysbiosis in the pathogenesis of SIV infection, with the goal of development new regiments or treatments to reduce or even halt the vicious cycle of gut damage and inflammation triggered by pathogenic HIV/SIV infection.

Disruption of Epithelial Barrier Integrity via Altered GILZ/c-Rel/RACK1 Signaling in Inflammatory Bowel Disease.

Given the role of Receptor for Activated C Kinase 1 (RACK1) in both immune cell activation and in the maintenance of the intestinal epithelial barrier integrity, we investigated whether it was involved in inflammatory bowel disease (IBD). RACK1 expression was analyzed in intestinal mucosal samples of healthy and IBD patients, in mice with chemically induced colitis, and in diseased in vitro 2D and 3D coculture models by luciferase assay, reverse transcription-quantitative PCR, Western blotting, immunofluorescence, and immunohistochemistry. Based on our finding that glucocorticoid-induced leucine zipper (GILZ or tsc22d3) positively correlates with RACK1 expression in IBD patients, GILZ knockout mice and cell silencing experiments were performed. RACK1 was significantly decreased in IBD, especially in ulcerative colitis. This was associated with an NF-κB/c-Rel-related mechanism, correlating with decreased GILZ protein expression. GILZ depletion confirmed a decrease in RACK1 expression, which favored SRC activation and led to a significant reduction in E-cadherin, resulting in impaired epithelial barrier integrity. Finally, our data highlighted that this novel mechanism could be considered to develop new therapies since dexamethasone, the first line of treatment in IBD, restored RACK1 expression through the glucocorticoid receptor in a c-Rel/GILZ-independent manner. We provide the first evidence that an alteration of RACK1/SRC/E-cadherin regulatory mechanism, correlating with decreased GILZ protein expression, is involved in epithelial barrier disruption. The clinical relevance is based on the fact that this mechanism involving GILZ/c-Rel-related RACK1 expression could be considered to improve IBD therapies, particularly in patients with low or no response to glucocorticoid treatment.

Site directed mutagenesis reveals functional importance of conserved amino acid residues within the N-terminal domain of Dpb2 in budding yeast.

In spite of being dispensable for catalysis, Dpb2, the second largest subunit of leading strand DNA polymerase (Polymerase ε) is essential for cell survival in budding yeast. Dpb2 physically connects polymerase epsilon with the replicative helicase (CMG,Cdc45-Mcm-GINS) by interacting with its Psf1 subunit. Dpb2-Psf1 interaction has been shown to be critical for incorporating polymerase ε into the replisome. Site-directed mutagenesis studies on conserved amino acid residues within the N-terminal domain of Dpb2 led to identification of key amino acid residues involved in interaction with Psf1 subunit of GINS. These amino acid residues are found to be well conserved among Dpb2 orthologues in higher eukaryotes thereby indicating the protein-protein interaction to be evolutionarily conserved. Replicating cells are known to mount a strong replicative stress response and DNA damage response upon exposure to diverse range of stressors. Here, we show that the absence of the N-terminal domain of Dpb2 increases the vulnerability of the budding yeast cells towards the cytotoxic effects of hydroxyurea (HU) and methyl methane sulphonate (MMS). Our results illustrate the importance of N-terminal domain of Dpb2 not only during replisome assembly but also in coordinating stress response in budding yeast. Considering high degree of sequence conservation across eukaryotes, Dpb2 subunit of leading-strand DNA polymerase appears to have important implications in maintenance of genome integrity.

Relative frequency dynamics and loading of beet necrotic yellow vein virus genomic RNAs during the acquisition by its vector Polymyxa betae.

Our understanding of the transmission of plant viruses by protozoan vectors remains poor and fragmented. The fate of viral elements in the living stages of the vector is unknown. Here, we first established a protocol allowing the purification of two forms of the vector free of cellular contaminants. This permitted the examination of the relative frequencies of beet necrotic yellow vein virus RNAs in the roots of its natural host and in two forms of its protozoan vector, Polymyxa betae, responsible for virus transmission. Our findings provide new insights into virus behavior during vector transmission, allowing us to analyze how the virus regulates its RNA frequencies and load within the vector. By focusing on the early stages of viral transmission and separating virus acquisition from transmission to new hosts, we pave the way for experiments aimed at elucidating the molecular mechanisms behind viral acquisition and the maintenance of viral genome integrity by P. betae.