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Genes May 2024Hereditary sensory and autonomic neuropathy type 1 is an autosomal dominant neuropathy caused by the or variants. These variants modify the preferred substrate of...
Hereditary sensory and autonomic neuropathy type 1 is an autosomal dominant neuropathy caused by the or variants. These variants modify the preferred substrate of serine palmitoyl transferase, responsible for the first step of sphingolipids synthesis, leading to accumulation of cytotoxic deoxysphingolipids. Diagnosis of HSAN1 is based on clinical symptoms, mainly progressive loss of distal sensory keep, and genetic analysis. Identifying new or "" variants raises the question as to their pathogenicity. This work focused on characterizing six new variants using in silico prediction tools, new meta-scores, 3D modeling, and functional testing to establish their pathogenicity. Variants from six patients with HSAN1 were studied. , CADD and REVEL scores and the 3D modeling software MITZLI were used to characterize the pathogenic effect of the variants. Functional tests based on plasma sphingolipids quantification (total deoxysphinganine, ceramides, and dihydroceramides) were performed by tandem mass spectrometry. In silico predictors did not provide very contrasting results when functional tests discriminated the different variants according to their impact on deoxysphinganine level or canonical sphingolipids synthesis. Two variants were newly described as pathogenic: NM_006415.4:c.998A>G and NM_006415.4:c.1015G>A. The combination of the different tools provides arguments to establish the pathogenicity of these new variants. When available, functional testing remains the best option to establish the in vivo impact of a variant. Moreover, the comprehension of metabolic dysregulation offers opportunities to develop new therapeutic strategies for these genetic disorders.
Topics: Humans; Serine C-Palmitoyltransferase; Hereditary Sensory and Autonomic Neuropathies; Mutation, Missense; Male; Female; Sphingolipids; Adult; Middle Aged
PubMed: 38927628
DOI: 10.3390/genes15060692 -
Genes May 2024The gene encodes a transcription factor in which pathogenic variants have been associated with both isolated and syndromic congenital cataracts. We aim to review the... (Review)
Review
The gene encodes a transcription factor in which pathogenic variants have been associated with both isolated and syndromic congenital cataracts. We aim to review the variants in the C-terminal DNA-binding domain associated with non-syndromic congenital cataracts and describe a patient with a novel, disease-causing de novo missense variant. Published reports of C-terminal variants and their associated congenital cataracts and ophthalmic findings were reviewed. The patient we present and his biological parents had genetic testing via a targeted gene panel followed by trio-based whole exome sequencing. A 4-year-old patient with a history of bilateral nuclear and cortical cataracts was found to have a novel, likely pathogenic de novo variant in , NM_005360.5:c.922A>G (p.Lys308Glu). No syndromic findings or anterior segment abnormalities were identified. We report the novel missense variant, c.922A>G (p.Lys308Glu), in the C-terminal DNA-binding domain of classified as likely pathogenic and associated with non-syndromic bilateral congenital cataracts.
Topics: Humans; Cataract; Proto-Oncogene Proteins c-maf; Mutation, Missense; Male; Child, Preschool; Protein Domains; Exome Sequencing
PubMed: 38927621
DOI: 10.3390/genes15060686 -
Genes May 2024In , is the primary gene involved in sex determination: haploid hemizygous eggs develop as drones, while females develop from eggs heterozygous for the gene. If...
In , is the primary gene involved in sex determination: haploid hemizygous eggs develop as drones, while females develop from eggs heterozygous for the gene. If diploid eggs are homozygous for the gene, diploid drones will develop, but will be eaten by worker bees before they are born. Therefore, high allelic diversity is a priority for colony survival and breeding. This study aims to investigate the variability of the hypervariable region (HVR) of the gene in bees sampled in an apiary under a selection scheme. To this end, an existing dataset of 100 whole-genome sequences was analyzed with a validated pipeline based on de novo assembly of sequences within the HVR region. In total, 102 allelic sequences were reconstructed and translated into amino acid sequences. Among these, 47 different alleles were identified, 44 of which had previously been observed, while 3 are novel alleles. The results show a high variability in the region in this breeding population of honeybees.
Topics: Animals; Bees; Alleles; Female; Sex Determination Processes; Male; Breeding; Italy; Insect Proteins; Genetic Variation
PubMed: 38927588
DOI: 10.3390/genes15060652 -
Biomedicines Jun 2024Chronic pancreatitis is often secondary to alcohol abuse, but pancreatitis with no other aetiology is frequently associated with variants in genes encoding proteins...
Chronic pancreatitis is often secondary to alcohol abuse, but pancreatitis with no other aetiology is frequently associated with variants in genes encoding proteins related to zymogen granule activation. Our goal was to identify genomic variants in a patient by analyzing an extended panel of genes associated with the intra-pancreatic activation of the trypsin pathway. A 23-year-old woman was addressed at our institution because of chronic pancreatitis of unknown aetiology presenting recurrent episodes since she was the age of four. Next Generation Sequencing was performed to analyze a panel of nine genes associated with pancreatitis (, , , , , , , and ). Three missense variants were found: p.Leu997Phe, maternally inherited, in the gene; p.Ile73Phe, paternally inherited, in the gene; and p.Phe790Ser, a de novo variant, in the gene. They were classified, respectively as probably benign, a Variant of Uncertain Significance, and the last one, which has never been described in the literature, as likely being pathogenic following American College of Medical Genetics and Genomics standard guidelines. Extensive intra-pancreatic activation of trypsin pathway gene sequencing detected rare variants that were not found with other gene screening and showed that variants in different genes may interact in contributing to the onset of the pancreatitis phenotype.
PubMed: 38927485
DOI: 10.3390/biomedicines12061278 -
Biomedicines May 2024The development of anticancer drugs based on zinc-dependent histone deacetylase inhibitors (HDACi) has acquired great practical significance over the past decade. The...
The development of anticancer drugs based on zinc-dependent histone deacetylase inhibitors (HDACi) has acquired great practical significance over the past decade. The most important HDACi characteristics are selectivity and strength of inhibition since they determine the mechanisms of therapeutic action. For in-cell testing of the selectivity of de novo-synthesized HDACi, Western blot analysis of the level of acetylation of bona fide protein substrates of HDACs of each class is usually used. However, the high labor intensity of this method prevents its widespread use in inhibitor screening. We developed an in-cell high-throughput screening method based on the use of three subtype-selective fluorogenic substrates of the general structure Boc-Lys(Acyl)-AMC, which in many cases makes it possible to determine the selectivity of HDACi at the class level. However, we found that the additional inhibitory activity of HDACi against metallo-β-lactamase domain-containing protein 2 (MBLAC2) leads to testing errors.
PubMed: 38927410
DOI: 10.3390/biomedicines12061203 -
Biomedicines May 2024Differentiation between acute liver failure (ALF) and acute-on-chronic liver failure (ACLF) can be challenging in patients with de novo liver disease but is important to...
Differentiation between acute liver failure (ALF) and acute-on-chronic liver failure (ACLF) can be challenging in patients with de novo liver disease but is important to indicate the referral to a transplant center and urgency of organ allocation. Leptin, an adipocyte-derived cytokine that regulates energy storage and satiety, has multiple regulatory functions in the liver. We enrolled 160 critically ill patients with liver disease and 20 healthy individuals to measure serum leptin concentrations as a potential biomarker for diagnostic and prognostic purposes. Notably, patients with ALF had higher concentrations of serum leptin compared to patients with decompensated advanced chronic liver disease (dACLD) or ACLF (110 vs. 50 vs. 29 pg/mL, < 0.001). Levels of serum leptin below 56 pg/mL excluded ALF in patients with acute hepatic disease, with a negative predictive value (NPV) of 98.8% in our cohort. Lastly, serum leptin did not show any dynamic changes within the first 48 h of ICU treatment, especially not in comparison with patients with ALF vs. ACLF or survivors vs. non-survivors. In conclusion, serum leptin may represent a helpful biomarker to exclude ALF in critically ill patients who present with acute liver dysfunction.
PubMed: 38927377
DOI: 10.3390/biomedicines12061170 -
Antibiotics (Basel, Switzerland) Jun 2024can acquire antimicrobial resistance (AMR) through horizontal gene transfer (HGT) from other spp. such as commensals like . Low doses of antimicrobials in food could...
can acquire antimicrobial resistance (AMR) through horizontal gene transfer (HGT) from other spp. such as commensals like . Low doses of antimicrobials in food could select for AMR in which could then be transferred to . In this study, we aimed to determine the lowest concentration of ciprofloxacin that can induce ciprofloxacin resistance (minimum selection concentration-MSC) in a isolate (ID-Co000790/2, a clinical isolate collected from a previous community study conducted at ITM). In this study, was serially passaged on gonococcal (GC) medium agar plates containing ciprofloxacin concentrations ranging from 1:100 to 1:10,000 below its ciprofloxacin MIC (0.006 µg/mL) for 6 days. After 6 days of serial passaging at ciprofloxacin concentrations of 1/100th of the MIC, 24 colonies emerged on the plate containing 0.06 µg/mL ciprofloxacin, which corresponds to the EUCAST breakpoint for . Their ciprofloxacin MICs were between 0.19 to 0.25 µg/mL, and whole genome sequencing revealed a missense mutation T91I in the gene, which has previously been found to cause reduced susceptibility to fluoroquinolones. The MSC was determined to be 0.06 ng/mL (0.00006 µg/mL), which is 100×-fold lower than the ciprofloxacin MIC. The implications of this finding are that the low concentrations of fluoroquinolones found in certain environmental samples, such as soil, river water, and even the food we eat, may be able to select for ciprofloxacin resistance in
PubMed: 38927226
DOI: 10.3390/antibiotics13060560 -
Biomolecules Jun 2024Resveratrol, a phenylpropanoid compound, exhibits diverse pharmacological properties, making it a valuable candidate for health and disease management. However, the...
Resveratrol, a phenylpropanoid compound, exhibits diverse pharmacological properties, making it a valuable candidate for health and disease management. However, the demand for resveratrol exceeds the capacity of plant extraction methods, necessitating alternative production strategies. Microbial synthesis offers several advantages over plant-based approaches and presents a promising alternative. stands out among microbial hosts due to its safe nature, abundant acetyl-CoA and malonyl-CoA availability, and robust pentose phosphate pathway. This study aimed to engineer for resveratrol production. The resveratrol biosynthetic pathway was integrated into by adding genes encoding tyrosine ammonia lyase from , 4-coumarate CoA ligase from , and stilbene synthase from . This resulted in the production of 14.3 mg/L resveratrol. A combination of endogenous and exogenous malonyl-CoA biosynthetic modules was introduced to enhance malonyl-CoA availability. This included genes encoding acetyl-CoA carboxylase 2 from , malonyl-CoA synthase, and a malonate transporter protein from . These strategies increased resveratrol production to 51.8 mg/L. The further optimization of fermentation conditions and the utilization of sucrose as an effective carbon source in YP media enhanced the resveratrol concentration to 141 mg/L in flask fermentation. By combining these strategies, we achieved a titer of 400 mg/L resveratrol in a controlled fed-batch bioreactor. These findings demonstrate the efficacy of as a platform for the de novo production of resveratrol and highlight the importance of metabolic engineering, enhancing malonyl-CoA availability, and media optimization for improved resveratrol production.
Topics: Resveratrol; Yarrowia; Metabolic Engineering; Sucrose; Acyltransferases; Vitis; Coenzyme A Ligases; Malonyl Coenzyme A; Nicotiana; Rhodotorula; Fermentation; Arabidopsis; Ammonia-Lyases; Bacterial Proteins
PubMed: 38927115
DOI: 10.3390/biom14060712 -
Mobile DNA Jun 2024Transposable Elements (TEs) are segments of DNA, typically a few hundred base pairs up to several tens of thousands bases long, that have the ability to generate new...
BACKGROUND
Transposable Elements (TEs) are segments of DNA, typically a few hundred base pairs up to several tens of thousands bases long, that have the ability to generate new copies of themselves in the genome. Most existing methods used to identify TEs in a newly sequenced genome are based on their repetitive character, together with detection based on homology and structural features. As new high quality assemblies become more common, including the availability of multiple independent assemblies from the same species, an alternative strategy for identification of TE families becomes possible in which we focus on the polymorphism at insertion sites caused by TE mobility.
RESULTS
We develop the idea of using the structural polymorphisms found in pangenomes to create a library of the TE families recently active in a species, or in a closely related group of species. We present a tool, pantera, that achieves this task, and illustrate its use both on species with well-curated libraries, and on new assemblies.
CONCLUSIONS
Our results show that pantera is sensitive and accurate, tending to correctly identify complete elements with precise boundaries, and is particularly well suited to detect larger, low copy number TEs that are often undetected with existing de novo methods.
PubMed: 38926873
DOI: 10.1186/s13100-024-00323-y -
Scientific Reports Jun 2024Heterozygous de novo mutations in the Activity-Dependent Neuroprotective Homeobox (ADNP) gene underlie Helsmoortel-Van der Aa syndrome (HVDAS). Most of these mutations...
Heterozygous de novo mutations in the Activity-Dependent Neuroprotective Homeobox (ADNP) gene underlie Helsmoortel-Van der Aa syndrome (HVDAS). Most of these mutations are situated in the last exon and we previously demonstrated escape from nonsense-mediated decay by detecting mutant ADNP mRNA in patient blood. In this study, wild-type and ADNP mutants are investigated at the protein level and therefore optimal detection of the protein is required. Detection of ADNP by means of western blotting has been ambiguous with reported antibodies resulting in non-specific bands without unique ADNP signal. Validation of an N-terminal ADNP antibody (Aviva Systems) using a blocking peptide competition assay allowed to differentiate between specific and non-specific signals in different sample materials, resulting in a unique band signal around 150 kDa for ADNP, above its theoretical molecular weight of 124 kDa. Detection with different C-terminal antibodies confirmed the signals at an observed molecular weight of 150 kDa. Our antibody panel was subsequently tested by immunoblotting, comparing parental and homozygous CRISPR/Cas9 endonuclease-mediated Adnp knockout cell lines and showed disappearance of the 150 kDa signal, indicative for intact ADNP. By means of both a GFPSpark and Flag-tag N-terminally fused to a human ADNP expression vector, we detected wild-type ADNP together with mutant forms after introduction of patient mutations in E. coli expression systems by site-directed mutagenesis. Furthermore, we were also able to visualize endogenous ADNP with our C-terminal antibody panel in heterozygous cell lines carrying ADNP patient mutations, while the truncated ADNP mutants could only be detected with epitope-tag-specific antibodies, suggesting that addition of an epitope-tag possibly helps stabilizing the protein. However, western blotting of patient-derived hiPSCs, immortalized lymphoblastoid cell lines and post-mortem patient brain material failed to detect a native mutant ADNP protein. In addition, an N-terminal immunoprecipitation-competent ADNP antibody enriched truncating mutants in overexpression lysates, whereas implementation of the same method failed to enrich a possible native mutant protein in immortalized patient-derived lymphoblastoid cell lines. This study aims to shape awareness for critical assessment of mutant ADNP protein analysis in Helsmoortel-Van der Aa syndrome.
Topics: Humans; Homeodomain Proteins; Nerve Tissue Proteins; Mutation; HEK293 Cells; Autism Spectrum Disorder; Heart Diseases; Facies; Neurodevelopmental Disorders
PubMed: 38926592
DOI: 10.1038/s41598-024-65608-x