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Science Advances Jun 2024The hierarchical chromatin organization begins with formation of nucleosomes, which fold into chromatin domains punctuated by boundaries and ultimately chromosomes. In a...
The hierarchical chromatin organization begins with formation of nucleosomes, which fold into chromatin domains punctuated by boundaries and ultimately chromosomes. In a hierarchal organization, lower levels shape higher levels. However, the dependence of higher-order 3D chromatin organization on the nucleosome-level organization has not been studied in cells. We investigated the relationship between nucleosome-level organization and higher-order chromatin organization by perturbing nucleosomes across the genome by deleting () and () chromatin remodeling factors in budding yeast. We find that changes in nucleosome-level properties are accompanied by changes in 3D chromatin organization. Short-range chromatin contacts up to a few kilo-base pairs decrease, chromatin domains weaken, and boundary strength decreases. Boundary strength scales with accessibility and moderately with width of nucleosome-depleted region. Change in nucleosome positioning seems to alter the stiffness of chromatin, which can affect formation of chromatin contacts. Our results suggest a biomechanical "bottom-up" mechanism by which nucleosome distribution across genome shapes 3D chromatin organization.
Topics: Nucleosomes; Chromatin Assembly and Disassembly; Chromatin; Genome, Fungal; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; DNA-Binding Proteins; Transcription Factors; Adenosine Triphosphatases
PubMed: 38848364
DOI: 10.1126/sciadv.adn2955 -
Renal Failure Dec 2024Fabry disease, a lysosomal storage disease, is an uncommon X-linked recessive genetic disorder stemming from abnormalities in the alpha-galactosidase gene () that codes...
Fabry disease, a lysosomal storage disease, is an uncommon X-linked recessive genetic disorder stemming from abnormalities in the alpha-galactosidase gene () that codes human alpha-Galactosidase A (α-Gal A). To date, over 800 mutations have been found to cause Fabry disease (FD). Continued enhancement of the mutation spectrum will contribute to a deeper recognition and underlying mechanisms of FD. In this study, a 27-year-old male proband exhibited a typical phenotype of Fabry disease. Subsequently, family screening for Fabry disease was conducted, and high-throughput sequencing was employed to identify the mutated gene. The three-level structure of the mutated protein was analyzed, and its subcellular localization and enzymatic activity were determined. Apoptosis was assessed in mutant cell lines to confirm the functional effects. As a result, a new mutation, c.777_778del (p. Gly261Leufs*3), in the gene was identified. The mutation caused a frameshift during translation and the premature appearance of a termination codon, which led to a partial deletion of the domain in C-terminal region and altered the protein's tertiary structure. experiments revealed a significant reduction of the enzymatic activity in mutant cells. The expression was noticeably decreased at the mRNA and protein levels in mutant cell lines. Additionally, the subcellular localization of α-Gal A changed from a homogeneous distribution to punctate aggregation in the cytoplasm. mutant cells exhibited significantly higher levels of apoptosis compared to wild-type cells.
Topics: Humans; Fabry Disease; alpha-Galactosidase; Male; Codon, Nonsense; Adult; Pedigree; China; Asian People; Apoptosis; East Asian People
PubMed: 38847497
DOI: 10.1080/0886022X.2024.2362391 -
AMB Express Jun 2024Bacterial contamination is the most prevalent infectious complication of blood transfusion in the developed world. To mitigate this, several ultraviolet light-based...
Bacterial contamination is the most prevalent infectious complication of blood transfusion in the developed world. To mitigate this, several ultraviolet light-based pathogen reduction technologies (PRTs), some of which require photo-chemicals, have been developed to minimize infection transmission. Relative to UV light, visible 405-nm light is safer and has shown potential to be developed as a PRT for the in situ treatment of ex vivo human plasma and platelet concentrates, without the need for photo-chemicals. This study investigates the effect of 405-nm light on human plasma, with focus on the compatibility of antimicrobial light doses with essential plasma clotting factors. To determine an effective antimicrobial dose that is compatible with plasma, prebagged human plasma (up to 300 mL) was seeded with common microbial contaminants and treated with increasing doses of 405-nm light (16 mW cm; ≤ 403 J cm). Post-exposure plasma protein integrity was investigated using an AOPP assay, in vitro coagulation tests, and ELISA-based measurement of fibrinogen and Protein S. Microbial contamination in 300 mL prebagged human plasma was significantly reduced (P ≤ 0.05) after exposure to ≤ 288 J cm, with microbial loads reduced by > 96.2%. This dose did not significantly affect the plasma protein quality parameters tested (P > 0.05). Increased doses (≥ 345 J cm) resulted in a 4.3% increase in clot times with no statistically significant change in protein activity or levels. Overall, this study has demonstrated that the effective microbicidal 405 light dose shows little to no negative effect on plasma quality.
PubMed: 38842656
DOI: 10.1186/s13568-024-01725-0 -
Scientific Reports Jun 2024Subcutaneous dirofilariasis, caused by the parasitic nematode Dirofilaria repens, is a growing concern in Europe, affecting both dogs and humans. This study focused on...
Subcutaneous dirofilariasis, caused by the parasitic nematode Dirofilaria repens, is a growing concern in Europe, affecting both dogs and humans. This study focused on D. repens Dr20/22, a protein encoded by an alt (abundant larval transcript) gene family. While well-documented in L3 larvae of other filariae species, this gene family had not been explored in dirofilariasis. The research involved cloning Dr20/22 cDNA, molecular characterization, and evaluating its potential application in the diagnosis of dirofilariasis. Although Real-Time analysis revealed mRNA expression in both adult worms and microfilariae, the native protein remained undetected in lysates from both developmental stages. This suggests the protein's specificity for L3 larvae and may be related to a process called SLTS (spliced leader trans-splicing), contributing to stage-specific gene expression. The specificity of the antigen for invasive larvae positions it as a promising early marker for dirofilariasis. However, ELISA tests using sera from infected and uninfected dogs indicated limited diagnostic utility. While further research is required, our findings contribute to a deeper understanding of the molecular and immunological aspects of host-parasite interactions and could offer insights into the parasite's strategies for evading the immune system.
Topics: Animals; Dogs; Dirofilariasis; Dirofilaria repens; Dog Diseases; Antibodies, Helminth; Helminth Proteins; Antigens, Helminth; Larva; Antibody Formation
PubMed: 38839868
DOI: 10.1038/s41598-024-63523-9 -
Proceedings of the National Academy of... Jun 2024Alcohol dehydrogenase 1B (ADH1B) is a primate-specific enzyme which, uniquely among the ADH class 1 family, is highly expressed both in adipose tissue and liver. Its...
Alcohol dehydrogenase 1B (ADH1B) is a primate-specific enzyme which, uniquely among the ADH class 1 family, is highly expressed both in adipose tissue and liver. Its expression in adipose tissue is reduced in obesity and increased by insulin stimulation. Interference with expression has also been reported to impair adipocyte function. To better understand the role of ADH1B in adipocytes, we used CRISPR/Cas9 to delete in human adipose stem cells (ASC). Cells lacking ADH1B failed to differentiate into mature adipocytes manifested by minimal triglyceride accumulation and a marked reduction in expression of established adipocyte markers. As ADH1B is capable of converting retinol to retinoic acid (RA), we conducted rescue experiments. Incubation of ADH1B-deficient preadipocytes with 9-cis-RA, but not with all-transretinol, significantly rescued their ability to accumulate lipids and express markers of adipocyte differentiation. A homozygous missense variant in (p.Arg313Cys) was found in a patient with congenital lipodystrophy of unknown cause. This variant significantly impaired the protein's dimerization, enzymatic activity, and its ability to rescue differentiation in ADH1B-deficient ASC. The allele frequency of this variant in the Middle Eastern population suggests that it is unlikely to be a fully penetrant cause of severe lipodystrophy. In conclusion, ADH1B appears to play an unexpected, crucial and cell-autonomous role in human adipocyte differentiation by serving as a necessary source of endogenous retinoic acid.
Topics: Humans; Alcohol Dehydrogenase; Adipogenesis; Adipocytes; Tretinoin; Cell Differentiation; CRISPR-Cas Systems; Mutation, Missense; Adipose Tissue
PubMed: 38838011
DOI: 10.1073/pnas.2319301121 -
Frontiers in Endocrinology 2024A prothrombotic state was demonstrated in patients with Cushing's syndrome and is involved in the development and progression of cardiovascular and renal damage in...
BACKGROUND AND AIMS
A prothrombotic state was demonstrated in patients with Cushing's syndrome and is involved in the development and progression of cardiovascular and renal damage in hypertensive patients. This study was designed to examine the relationships between cortisol secretion and the hemostatic and fibrinolytic systems in hypertension.
METHODS
In 149 middle-aged, nondiabetic, essential hypertensive patients free of cardiovascular and renal complications, we measured hemostatic markers that express the spontaneous activation of the coagulation and fibrinolytic systems and assessed daily cortisol levels (8 AM, 3 PM, 12 AM; area under the curve, AUC-cortisol) together with the cortisol response to dexamethasone overnight suppression (DST-cortisol).
RESULTS
Plasma levels of D-dimer (D-dim), prothrombin fragment 1 + 2 (F1 + 2), and von Willebrand factor (vWF) were progressively and significantly higher across tertiles of AUC-cortisol and DST-cortisol, whereas no differences were observed in fibrinogen, tissue plasminogen activator, plasminogen activator inhibitor-1, antithrombin III, protein C, and protein S. D-dim, F1 + 2, and vWF were significantly and directly correlated with age and both AUC-cortisol and DST-cortisol. Multivariate regression analysis showed that both AUC-cortisol and DST-cortisol were related to plasma D-dim, F1 + 2, and vWF independently of age, body mass index, blood pressure, and renal function.
CONCLUSION
Greater daily cortisol profile and cortisol response to overnight suppression are independently associated with a prothrombotic state in hypertensive patients and might contribute to the development of organ damage and higher risk of cardiovascular complications.
Topics: Humans; Male; Middle Aged; Female; Hydrocortisone; Dexamethasone; Hypertension; Adult; Thrombosis; von Willebrand Factor; Circadian Rhythm; Aged; Biomarkers
PubMed: 38836224
DOI: 10.3389/fendo.2024.1397062 -
Cellular & Molecular Biology Letters Jun 2024The molecular basis for bulk autophagy activation due to a deficiency in essential nutrients such as carbohydrates, amino acids, and nitrogen is well understood. Given...
The molecular basis for bulk autophagy activation due to a deficiency in essential nutrients such as carbohydrates, amino acids, and nitrogen is well understood. Given autophagy functions to reduce surplus to compensate for scarcity, it theoretically possesses the capability to selectively degrade specific substrates to meet distinct metabolic demands. However, direct evidence is still lacking that substantiates the idea that autophagy selectively targets specific substrates (known as selective autophagy) to address particular nutritional needs. Recently, Gross et al. found that during phosphate starvation (P-S), rather than nitrogen starvation (N-S), yeasts selectively eliminate peroxisomes by dynamically altering the composition of the Atg1/ULK kinase complex (AKC) to adapt to P-S. This study elucidates how the metabolite sensor Pho81 flexibly interacts with AKC and guides selective autophagic clearance of peroxisomes during P-S, providing novel insights into the metabolic contribution of autophagy to special nutritional needs.
Topics: Autophagy; Phosphates; Saccharomyces cerevisiae Proteins; Peroxisomes; Saccharomyces cerevisiae; Autophagy-Related Protein-1 Homolog; Autophagy-Related Proteins; Protein Serine-Threonine Kinases; Protein Kinases
PubMed: 38834954
DOI: 10.1186/s11658-024-00597-3 -
Nature Communications Jun 2024Persisting replication intermediates can confer mitotic catastrophe. Loss of the fission yeast telomere protein Taz1 (ortholog of mammalian TRF1/TRF2) causes telomeric...
Persisting replication intermediates can confer mitotic catastrophe. Loss of the fission yeast telomere protein Taz1 (ortholog of mammalian TRF1/TRF2) causes telomeric replication fork (RF) stalling and consequently, telomere entanglements that stretch between segregating mitotic chromosomes. At ≤20 °C, these entanglements fail to resolve, resulting in lethality. Rif1, a conserved DNA replication/repair protein, hinders the resolution of telomere entanglements without affecting their formation. At mitosis, local nuclear envelope (NE) breakdown occurs in the cell's midregion. Here we demonstrate that entanglement resolution occurs in the cytoplasm following this NE breakdown. However, in response to taz1Δ telomeric entanglements, Rif1 delays midregion NE breakdown at ≤20 °C, in turn disfavoring entanglement resolution. Moreover, Rif1 overexpression in an otherwise wild-type setting causes cold-specific NE defects and lethality, which are rescued by membrane fluidization. Hence, NE properties confer the cold-specificity of taz1Δ lethality, which stems from postponement of NE breakdown. We propose that such postponement promotes clearance of simple stalled RFs, but resolution of complex entanglements (involving strand invasion between nonsister telomeres) requires rapid exposure to the cytoplasm.
Topics: Nuclear Envelope; Schizosaccharomyces; Telomere; Schizosaccharomyces pombe Proteins; Telomere-Binding Proteins; Anaphase; DNA Replication
PubMed: 38830842
DOI: 10.1038/s41467-024-48382-2 -
MSystems Jun 2024Fast growth phenotypes are achieved through optimal transcriptomic allocation, in which cells must balance tradeoffs in resource allocation between diverse functions....
Fast growth phenotypes are achieved through optimal transcriptomic allocation, in which cells must balance tradeoffs in resource allocation between diverse functions. One such balance between stress readiness and unbridled growth in has been termed the fear versus greed (f/g) tradeoff. Two specific RNA polymerase (RNAP) mutations observed in adaptation to fast growth have been previously shown to affect the f/g tradeoff, suggesting that genetic adaptations may be primed to control f/g resource allocation. Here, we conduct a greatly expanded study of the genetic control of the f/g tradeoff across diverse conditions. We introduced 12 RNA polymerase (RNAP) mutations commonly acquired during adaptive laboratory evolution (ALE) and obtained expression profiles of each. We found that these single RNAP mutation strains resulted in large shifts in the f/g tradeoff primarily in the RpoS regulon and ribosomal genes, likely through modifying RNAP-DNA interactions. Two of these mutations additionally caused condition-specific transcriptional adaptations. While this tradeoff was previously characterized by the RpoS regulon and ribosomal expression, we find that the GAD regulon plays an important role in stress readiness and ppGpp in translation activity, expanding the scope of the tradeoff. A phylogenetic analysis found the greed-related genes of the tradeoff present in numerous bacterial species. The results suggest that the f/g tradeoff represents a general principle of transcriptome allocation in bacteria where small genetic changes can result in large phenotypic adaptations to growth conditions.IMPORTANCETo increase growth, must raise ribosomal content at the expense of non-growth functions. Previous studies have linked RNAP mutations to this transcriptional shift and increased growth but were focused on only two mutations found in the protein's central region. RNAP mutations, however, commonly occur over a large structural range. To explore RNAP mutations' impact, we have introduced 12 RNAP mutations found in laboratory evolution experiments and obtained expression profiles of each. The mutations nearly universally increased growth rates by adjusting said tradeoff away from non-growth functions. In addition to this shift, a few caused condition-specific adaptations. We explored the prevalence of this tradeoff across phylogeny and found it to be a widespread and conserved trend among bacteria.
PubMed: 38829048
DOI: 10.1128/msystems.00305-24 -
PeerJ 2024The method currently available to diagnose shigellosis is insensitive and has many limitations. Thus, this study was designed to identify specific antigenic protein(s)...
BACKGROUND
The method currently available to diagnose shigellosis is insensitive and has many limitations. Thus, this study was designed to identify specific antigenic protein(s) among the cell surface associated proteins (SAPs) of that would be valuable in the development of an alternative diagnostic assay for shigellosis, particularly one that could be run using a stool sample rather than serum.
METHODS
The SAPs of clinical isolates of S. , , and were extracted from an overnight culture grown at 37 °C using acidified-glycine extraction methods. Protein profiles were observed by SDS-PAGE. To determine if antibodies specific to certain SAPs were present in both sera and stool suspensions, Western blot analysis was used to detect the presence of IgA, IgG, and IgM.
RESULTS
Immunoblot analysis revealed that sera from patients infected with . recognized 31 proteins. These SAP antigens are recognized by the host humoral response during infection. Specific antibodies against these antigens were also observed in intestinal secretions of shigellosis patients. Of these 31 proteins, the 35 kDa protein specifically reacted against IgA present in patients' stool suspensions. Further study illustrated the immunoreactivity of this protein in , and . This is the first report that demonstrates the presence of immunoreactive SAPs in stool suspensions. The SAPSs could be very useful in developing a simple and rapid serodiagnostic assay for shigellosis directly from stool specimens.
Topics: Humans; Feces; Dysentery, Bacillary; Shigella flexneri; Bacterial Proteins; Antibodies, Bacterial; Antigens, Bacterial; Blotting, Western; Electrophoresis, Polyacrylamide Gel; Immunoglobulin A
PubMed: 38827305
DOI: 10.7717/peerj.17498