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MBio Jun 2024Human adenoviruses (HAdVs) are small DNA viruses that generally cause mild disease. Certain strains, particularly those belonging to species B HAdVs, can cause severe...
Human adenoviruses (HAdVs) are small DNA viruses that generally cause mild disease. Certain strains, particularly those belonging to species B HAdVs, can cause severe pneumonia and have a relatively high mortality rate. Little is known about the molecular aspects of how these highly pathogenic species affect the infected cell and how they suppress innate immunity. The present study provides molecular insights into how species B adenoviruses suppress the interferon signaling pathway. Our study shows that these viruses, unlike HAdV-C2, are resistant to type I interferon. This resistance likely arises due to the highly efficient suppression of interferon-stimulated gene expression. Unlike in HAdV-C2, HAdV-B7 and B14 sequester STAT2 and RNA polymerase II from interferon-stimulated gene promoters in infected cells. This results in suppressed interferon- stimulated gene activation. In addition, we show that RuvBL1 and RuvBL2, cofactors important for RNA polymerase II recruitment to promoters and interferon-stimulated gene activation, are redirected to the cytoplasm forming high molecular weight complexes that, likely, are unable to associate with chromatin. Proteomic analysis also identified key differences in the way these viruses affect the host cell, providing insights into species B-associated high pathogenicity. Curiously, we observed that at the level of protein expression changes to the infected cell, HAdV-C2 and B7 were more similar than those of the same species, B7 and B14. Collectively, our study represents the first such study of innate immune suppression by the highly pathogenic HAdV-B7 and B14, laying an important foundation for future investigations.IMPORTANCEHuman adenoviruses form a large family of double-stranded DNA viruses known for a variety of usually mild diseases. Certain strains of human adenovirus cause severe pneumonia leading to much higher mortality and morbidity than most other strains. The reasons for this enhanced pathogenicity are unknown. Our study provides a molecular investigation of how these highly pathogenic strains might inactivate the interferon signaling pathway, highlighting the lack of sensitivity of these viruses to type I interferon in general while providing a global picture of how viral changes in cellular proteins drive worse disease outcomes.
PubMed: 38940561
DOI: 10.1128/mbio.01038-24 -
MBio Jun 2024Conjugative type 4 secretion systems (T4SSs) are the main driver for the spread of antibiotic resistance genes and virulence factors in bacteria. To deliver the DNA...
Conjugative type 4 secretion systems (T4SSs) are the main driver for the spread of antibiotic resistance genes and virulence factors in bacteria. To deliver the DNA substrate to recipient cells, it must cross the cell envelopes of both donor and recipient bacteria. In the T4SS from the enterococcal conjugative plasmid pCF10, PrgK is known to be the active cell wall degrading enzyme. It has three predicted extracellular hydrolase domains: metallo-peptidase (LytM), soluble lytic transglycosylase (SLT), and cysteine, histidine-dependent amidohydrolases/peptidases (CHAP). Here, we report the structure of the LytM domain and show that its active site is degenerate and lacks the active site metal. Furthermore, we show that only the predicted SLT domain is functional and that it unexpectedly has a muramidase instead of a lytic transglycosylase activity. While we did not observe any peptidoglycan hydrolytic activity for the LytM or CHAP domain, we found that these domains downregulated the SLT muramidase activity. The CHAP domain was also found to be involved in PrgK dimer formation. Furthermore, we show that PrgK interacts with PrgL, which likely targets PrgK to the rest of the T4SS. The presented data provides important information for understanding the function of Gram-positive T4SSs.IMPORTANCEAntibiotic resistance is a large threat to human health and is getting more prevalent. One of the major contributors to the spread of antibiotic resistance among different bacteria is type 4 secretion systems (T4SS). However, mainly T4SSs from Gram-negative bacteria have been studied in detail. T4SSs from Gram-positive bacteria, which stand for more than half of all hospital-acquired infections, are much less understood. The significance of our research is in identifying the function and regulation of a cell wall hydrolase, a key component of the pCF10 T4SS from . This system is one of the best-studied Gram-positive T4SSs, and this added knowledge aids in our understanding of horizontal gene transfer in as well as other medically relevant Gram-positive bacteria.
PubMed: 38940556
DOI: 10.1128/mbio.00488-24 -
MBio Jun 2024Merkel cell polyomavirus (MCPyV) is a double-stranded tumor virus that is the main causative agent of Merkel cell carcinoma (MCC). The MCPyV large T antigen (LT), an...
UNLABELLED
Merkel cell polyomavirus (MCPyV) is a double-stranded tumor virus that is the main causative agent of Merkel cell carcinoma (MCC). The MCPyV large T antigen (LT), an essential viral DNA replication protein, maintains viral persistence by interacting with host Skp1-Cullin 1-F-box (SCF) E3 ubiquitin ligase complexes, which subsequently induces LT's proteasomal degradation, restricting MCPyV DNA replication. SCF E3 ubiquitin ligases require their substrates to be phosphorylated to bind them, utilizing phosphorylated serine residues as docking sites. The MCPyV LT unique region (MUR) is highly phosphorylated and plays a role in multiple host protein interactions, including SCF E3 ubiquitin ligases. Therefore, this domain highly governs LT stability. Though much work has been conducted to identify host factors that restrict MCPyV LT protein expression, the kinase(s) that cooperates with the SCF E3 ligase remains unknown. Here, we demonstrate that casein kinase 1 alpha (CK1α) negatively regulates MCPyV LT stability and LT-mediated replication by modulating interactions with the SCF β-TrCP. Specifically, we show that numerous CK1 isoforms (α, δ, ε) localize in close proximity to MCPyV LT through proximity ligation assays (PLA) and CK1α overexpression mainly resulted in decreased MCPyV LT protein expression. Inhibition of CK1α using short hairpin RNA (shRNA) and treatment of a CK1α inhibitor or an mTOR inhibitor, TORKinib, resulted in decreased β-TrCP interaction with LT, increased LT expression, and enhanced MCPyV replication. The expression level of the gene transcripts is higher in MCPyV-positive MCC, suggesting a vital role of CK1α in limiting MCPyV replication required for establishing persistent infection.
IMPORTANCE
Merkel cell polyomavirus (MCPyV) large tumor antigen is a polyphosphoprotein and the phosphorylation event is required to modulate various functions of LT, including viral replication. Therefore, cellular kinase pathways are indispensable for governing MCPyV polyomavirus infection and life cycle in coordinating with the immunosuppression environment at disease onset. Understanding the regulation mechanisms of MCPyV replication by viral and cellular factors will guide proper prevention strategies with targeted inhibitors for MCPyV-associated Merkel cell carcinoma (MCC) patients, who currently lack therapies.
PubMed: 38940554
DOI: 10.1128/mbio.01117-24 -
Analytical Chemistry Jun 2024Low-mass soluble β-amyloid peptide oligomers (LSAβOs) play a crucial role in the pathogenesis of Alzheimer's disease. However, these oligomers exhibit heterogeneity in...
Low-mass soluble β-amyloid peptide oligomers (LSAβOs) play a crucial role in the pathogenesis of Alzheimer's disease. However, these oligomers exhibit heterogeneity in terms of structure, stability, and stoichiometry, and their abundance in biofluids is low, making accurate identification challenging. In this study, we developed a DNA nanocage-assisted method for selective sizing and sensitive quantification of LSAβOs in serum. Using LSAβO less than 10 kDa (LSAβO) and less than 30 kDa (LSAβO) as models, the size-matching rules between DNA nanocages and LSAβOs were investigated, and two appropriate nanocages were selected for the detection of two LSAβOs, respectively. Both nanocages were functionalized by encapsulating oligomer's aptamer and a complementary sequence within their cavities. Once the LSAβO entered the corresponding nanocage cavity, the complementary sequence was released, triggering a hybridization chain reaction on an electrochemical sensing platform. The system achieved size-selective discrimination of LSAβO with a linear range of 10-150 pM and LSAβO with a linear range of 15-150 pM. Real sample testing confirmed the applicability of the method for blood-based diagnosis. The DNA nanocage-assisted electrochemical analysis platform provides an accurate, highly selective, and sensitive approach for oligomer analysis, which is significant for amyloid protein research and related disease diagnosis.
PubMed: 38940533
DOI: 10.1021/acs.analchem.4c01465 -
Animal Biotechnology Jun 2024As a protein structurally similar to insulin, relaxin3 (RLN3) plays a role in promoting arousal, suppressing depressive or anxious behaviors. Two studies revealed the...
Characterization of chicken gene: mRNA expression and response to reproductive hormone treatment in ovarian granulosa cells, and single nucleotide polymorphisms associated with egg laying traits in hens.
As a protein structurally similar to insulin, relaxin3 (RLN3) plays a role in promoting arousal, suppressing depressive or anxious behaviors. Two studies revealed the increase of RLN3 expression during chicken follicle selection. In this study, by real-time quantitative PCR and luciferase assay, mRNA expression and single nucleotide polymorphisms (SNPs) of chicken were investigated. The mRNA expression of chicken was higher in the granulosa cell of hierarchal follicles (Post-GCs) than that of pre-hierarchal follicles (Pre-GCs). In Pre-GCs, the mRNA expression of chicken was stimulated by FSH and progesterone; in Post-GCs, it was stimulated by higher concentration of estrogen and FSH, however, was inhibited by progesterone. Four SNPs including g.-655G > C, g-592G > A, g.-372T > A and g.-282G > C were identified in the critical promoter region from -1291 bp to -207 bp of chicken , among which g.-655G > C, and g-592G > A were associated with age at first laying and clutch size, respectively, in Zaozhuang Sunzhi chickens. At g.-655G > C and g-592G > A, allele and allele had higher transcriptional activity, respectively. These data suggest that RLN3 plays an important role in chicken follicle development and SNPs in its promoter region are potential DNA markers for improving egg production traits.
PubMed: 38940516
DOI: 10.1080/10495398.2024.2370810 -
MSphere Jun 2024The gut microbiome has the potential to buffer temporal variations in resource availability and consumption, which may play a key role in the ability of animals to adapt...
UNLABELLED
The gut microbiome has the potential to buffer temporal variations in resource availability and consumption, which may play a key role in the ability of animals to adapt to a broad range of habitats. We investigated the temporal composition and function of the gut microbiomes of wild common marmosets () exploiting a hot, dry environment-Caatinga-in northeastern Brazil. We collected fecal samples during two time periods (July-August and February-March) for 2 years from marmosets belonging to eight social groups. We used 16S rRNA gene amplicon sequencing, metagenomic sequencing, and butyrate RT-qPCR to assess changes in the composition and potential function of their gut microbiomes. Additionally, we identified the plant, invertebrate, and vertebrate components of the marmosets' diet via DNA metabarcoding. Invertebrate, but not plant or vertebrate, consumption varied across the year. However, gut microbiome composition and potential function did not markedly vary across study periods or as a function of diet composition. Instead, the gut microbiome differed markedly in both composition and potential function across marmosets residing in different social groups. We highlight the likely role of factors, such as behavior, residence, and environmental heterogeneity, in modulating the structure of the gut microbiome.
IMPORTANCE
In a highly socially cohesive and cooperative primate, group membership more strongly predicts gut microbiome composition and function than diet.
PubMed: 38940510
DOI: 10.1128/msphere.00233-24 -
MSphere Jun 2024Bacterial conjugation systems pose a major threat to human health through their widespread dissemination of mobile genetic elements (MGEs) carrying cargoes of antibiotic...
UNLABELLED
Bacterial conjugation systems pose a major threat to human health through their widespread dissemination of mobile genetic elements (MGEs) carrying cargoes of antibiotic resistance genes. Using the Cre Recombinase Assay for Translocation (CRAfT), we recently reported that the IncFV pED208 conjugation system also translocates at least 16 plasmid-encoded proteins to recipient bacteria. Here, we deployed a high-throughput CRAfT screen to identify the repertoire of chromosomally encoded protein substrates of the pED208 system. We identified 32 substrates encoded by the W3110 genome with functions associated with (i) DNA/nucleotide metabolism, (ii) stress tolerance/physiology, (iii) transcriptional regulation, or (iv) toxin inhibition. The respective gene deletions did not impact pED208 transfer proficiencies, nor did Group 1 (DNA/nucleotide metabolism) mutations detectably alter the SOS response elicited in new transconjugants upon acquisition of pED208. However, MC4100(pED208) donor cells intrinsically exhibit significantly higher SOS activation than plasmid-free MC4100 cells, and this plasmid carriage-induced stress response is further elevated in donor cells deleted of several Group 1 genes. Among 10 characterized substrates, we gained evidence of C-terminal or internal translocation signals that could function independently or synergistically for optimal protein transfer. Remarkably, nearly all tested proteins were also translocated through the IncN pKM101 and IncP RP4 conjugation systems. This repertoire of protein substrates, here termed the F plasmid "conjutome," is thus characterized by functions of potential benefit to new transconjugants, diverse TSs, and the capacity for promiscuous transfer through heterologous conjugation systems.
IMPORTANCE
Conjugation systems comprise a major subfamily of the type IV secretion systems (T4SSs) and are the progenitors of a second large T4SS subfamily dedicated to translocation of protein effectors. This study examined the capacity of conjugation machines to function as protein translocators. Using a high-throughput reporter screen, we determined that 32 chromosomally encoded proteins are delivered through an F plasmid conjugation system. The translocated proteins potentially enhance the establishment of the co-transferred F plasmid or mitigate mating-induced stresses. Translocation signals located C-terminally or internally conferred substrate recognition by the F system and, remarkably, many substrates also were translocated through heterologous conjugation systems. Our findings highlight the plasticity of conjugation systems in their capacities to co-translocate DNA and many protein substrates.
PubMed: 38940509
DOI: 10.1128/msphere.00354-24 -
The Journal of Clinical Endocrinology... Jun 2024Atypical parathyroid tumor (APT) represents a neoplasm characterized by histological features typical of parathyroid carcinoma (PC) but lacking local infiltration and/or...
CONTEXT
Atypical parathyroid tumor (APT) represents a neoplasm characterized by histological features typical of parathyroid carcinoma (PC) but lacking local infiltration and/or distant metastasis, leading to uncertainty regarding its malignant potential.
OBJECTIVE
To characterize the molecular landscape and deregulated pathways in APT.
METHODS
Whole exome sequencing (WES) was conducted on 16 APTs. DNA from tumors and matched peripheral blood underwent WES using Illumina HiSeq3000.
RESULTS
A total of 192 nonsynonymous variants were identified. The median number of protein-altering mutations was 9. The most frequently mutated genes included BCOR, CLMN, EZH1, JAM2, KRTAP13-3, MUC16, MUC19, and OR1S1. Seventeen mutated genes belong to the Cancer Gene Census list. The most consistent hub genes identified through STRING network analysis were ATM, COL4A5, EZH2, MED12, MEN1, MTOR, PI3, PIK3CA, PIK3CB, and UBR5. Deregulated pathways included the PI3 K/AKT/mTOR pathway, Wnt signaling, and extracellular matrix organization. Variants in genes such as MEN1, CDC73, EZH2, PIK3CA, and MTOR, previously reported as established or putative/candidate driver genes in benign adenoma (PA) and/or PC, were also identified in APT.
CONCLUSIONS
APT does not appear to have a specific molecular signature but shares genomic alterations with both PA and PC. The incidence of CDC73 mutations is low, and it remains unclear whether these mutations are associated with a higher risk of recurrence. Our study confirms that PI3 K/AKT/mTOR and Wnt signaling represents the pivotal pathways in parathyroid tumorigenesis and also revealed mutations in key epigenetic modifier genes (BCOR, KDM2A, MBD4, and EZH2) involved in chromatin remodeling, DNA, and histone methylation.
PubMed: 38940486
DOI: 10.1210/clinem/dgae441 -
Bioanalysis Jun 2024Increased knowledge of biodistribution and pharmacokinetics of lipid nanoparticle (LNP)-encapsulated mRNA drug components may aid efficacy and safety evaluation. Mice...
Increased knowledge of biodistribution and pharmacokinetics of lipid nanoparticle (LNP)-encapsulated mRNA drug components may aid efficacy and safety evaluation. Mice were subcutaneously administrated LNP encapsulated enhanced green fluorescent protein mRNA and sampled up to 72 h after dosing. LNP, mRNA and translated protein were quantified by LC-MS, branched DNA and ELISA. Highest levels of LNP and mRNA were detected in skin, followed by spleen, but also rapidly distributed to circulation. Translated protein showed high concentration in skin and spleen, but also in liver and kidney across 24 h where the LNP was cleared at 4 h. Subcutaneously dosing LNP encapsulated mRNA in mice resulted in a nonlinear relationship of LNP, mRNA and protein concentration across multiple tissues.
PubMed: 38940441
DOI: 10.1080/17576180.2024.2360361 -
Chembiochem : a European Journal of... Jun 2024Water-soluble phthalocyanine (Pc) derivatives have been regarded as potential G-quadruplex (G4) nucleic acid-targeting ligands for anticancer therapy and have been...
Water-soluble phthalocyanine (Pc) derivatives have been regarded as potential G-quadruplex (G4) nucleic acid-targeting ligands for anticancer therapy and have been extensively studied as effective photosensitizers for photodynamic therapy (PDT). Understanding how photosensitizers interact with nucleic acids and the subsequent photolytic reactions is essential for deciphering the initial steps of PDT, thereby aiding in the development of new photosensitizing agents. In this study, we found that red-light irradiation of a mixture of a Zn(II) Pc derivative and an all-parallel G4 DNA leads to catalytic and selective photodegradation of the DNA by reactive oxygen species (ROS) generated from the Zn(II) Pc derivative bound to DNA through a reaction mechanism similar to that of an enzyme reaction. This finding provides a novel insight into the molecular design of a photosensitizer to enhance its PDT efficacy.
PubMed: 38940417
DOI: 10.1002/cbic.202400197