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Biology May 2024has emerged as a promising model organism for basic studies in Decapod. However, the current transcriptome information on this species is based on next-generation...
has emerged as a promising model organism for basic studies in Decapod. However, the current transcriptome information on this species is based on next-generation sequencing technologies, which are limited by a short read length. Therefore, the present study aimed to generate a full-length transcriptome assembly of utilizing the PacBio Sequel Ⅱ platform. The resulting transcriptome assembly comprised 5831 transcripts with an N50 value of 3697 bp. Remarkably, 90.5% of these transcripts represented novel isoforms of known genes. The transcripts were further searched against the NR, SwissProt, KEGG, KOG, GO, NT, and Pfam databases. A total of 24.8% of the transcripts can be annotated across all seven databases. Additionally, 1236 alternative splicing events, 344 transcription factors, and 124 long non-coding RNAs (LncRNAs) were predicted. Based on the alternative splicing annotation results, a RING finger protein NHL-1 gene from () was identified. There are 15 transcripts in . The longest transcript is 4995 bp in length and encodes a putative protein of 1665 amino acids. A phylogenetic analysis showed its close relationship with NHL-1 from other crustacean species. This report represents the full-length transcriptome of and will facilitate research on functional genomics and environmental adaptation in this species.
PubMed: 38927246
DOI: 10.3390/biology13060366 -
Biomolecules May 2024The Actinopterygian and specifically the Teleostean peroxisome proliferator-activated receptors (PPARs) present an impressive variability and complexity in their... (Review)
Review
The Actinopterygian and specifically the Teleostean peroxisome proliferator-activated receptors (PPARs) present an impressive variability and complexity in their structures, both at the gene and protein levels. These structural differences may also reflect functional divergence from their mammalian homologs, or even between fish species. This review, taking advantage of the data generated from the whole-genome sequencing of several fish species, highlights the differences in the primary structure of the receptors, while discussing results from the literature pertaining to the functions of fish PPARs and their activation by natural and synthetic compounds.
Topics: Animals; Peroxisome Proliferator-Activated Receptors; Fishes
PubMed: 38927038
DOI: 10.3390/biom14060634 -
Scientific Reports Jun 2024Opioids are the gold standard for the treatment of chronic pain but are limited by adverse side effects. In our earlier work, we showed that Heat shock protein 90...
Opioids are the gold standard for the treatment of chronic pain but are limited by adverse side effects. In our earlier work, we showed that Heat shock protein 90 (Hsp90) has a crucial role in regulating opioid signaling in spinal cord; Hsp90 inhibition in spinal cord enhances opioid anti-nociception. Building on these findings, we injected the non-selective Hsp90 inhibitor KU-32 by the intrathecal route into male and female CD-1 mice, showing that morphine anti-nociceptive potency was boosted by 1.9-3.5-fold in acute and chronic pain models. At the same time, tolerance was reduced from 21-fold to 2.9 fold and established tolerance was rescued, while the potency of constipation and reward was unchanged. These results demonstrate that spinal Hsp90 inhibition can improve the therapeutic index of morphine. However, we also found that systemic non-selective Hsp90 inhibition blocked opioid pain relief. To avoid this effect, we used selective small molecule inhibitors and CRISPR gene editing to identify 3 Hsp90 isoforms active in spinal cord (Hsp90α, Hsp90β, and Grp94) while only Hsp90α was active in brain. We thus hypothesized that a systemically delivered selective inhibitor to Hsp90β or Grp94 could selectively inhibit spinal cord Hsp90 activity, resulting in enhanced opioid therapy. We tested this hypothesis using intravenous delivery of KUNB106 (Hsp90β) and KUNG65 (Grp94), showing that both drugs enhanced morphine anti-nociceptive potency while rescuing tolerance. Together, these results suggest that selective inhibition of spinal cord Hsp90 isoforms is a novel, translationally feasible strategy to improve the therapeutic index of opioids.
Topics: Animals; HSP90 Heat-Shock Proteins; Spinal Cord; Mice; Analgesics, Opioid; Male; Female; Morphine; Protein Isoforms; Drug Tolerance; Chronic Pain; Disease Models, Animal; Injections, Spinal
PubMed: 38926482
DOI: 10.1038/s41598-024-65637-6 -
The Journal of Biological Chemistry Jun 2024Transthyretin (TTR) is a homotetrameric protein involved in the transport of thyroxine. More than 150 different mutations have been described in the TTR gene, several of...
Transthyretin (TTR) is a homotetrameric protein involved in the transport of thyroxine. More than 150 different mutations have been described in the TTR gene, several of them associated with familial amyloid cardiomyopathy (FAC). Recently, our group described a new variant of TTR in Brazil, namely A39D-TTR, which causes a severe cardiac condition. Position 39 is in the AB loop, a region of the protein that is located within the thyroxine-binding channels and is involved in tetramer formation. In the present study we solved the structure and characterize the thermodynamic stability of this new variant of TTR using urea and high hydrostatic pressure (HHP). Interestingly, during the process of purification, A39D-TTR turned out to be a dimer and not a tetramer, a variation that might be explained by the close contact of the four aspartic acids at position 39, where they face each other inside the thyroxine channel. In the presence of sub-denaturing concentrations of urea, bis-ANS binding and dynamic light scattering revealed A39D-TTR in the form of a molten-globule dimer. Co-expression of A39D and WT isoforms in the same bacterial cell did not produce heterodimers or heterotetramers, suggesting that somehow a negative charge at the AB loop precludes tetramer formation. A39D-TTR proved to be highly amyloidogenic, even at mildly acidic pH values where WT-TTR does not aggregate. Interestingly, despite being a dimer, aggregation of A39D-TTR was inhibited by diclofenac, which binds to the thyroxine channel in the tetramer, suggesting the existence of other pockets in A39D-TTR able to accommodate this molecule.
PubMed: 38925327
DOI: 10.1016/j.jbc.2024.107495 -
European Journal of Medicinal Chemistry Jun 2024Lactate dehydrogenase-A (LDHA) is the major isoform of lactate dehydrogenases (LDH) that is overexpressed and linked to poor survival in pancreatic ductal adenocarcinoma...
Lactate dehydrogenase-A (LDHA) is the major isoform of lactate dehydrogenases (LDH) that is overexpressed and linked to poor survival in pancreatic ductal adenocarcinoma (PDAC). Despite some progress, current LDH inhibitors have poor structural and physicochemical properties or exhibit unfavorable pharmacokinetics that have hampered their development. The present study reports the synthesis and biological evaluation of a novel class of LDHA inhibitors comprising a succinic acid monoamide motif. Compounds 6 and 21 are structurally related analogs that demonstrated potent inhibition of LDHA with ICs of 46 nM and 72 nM, respectively. We solved cocrystal structures of compound 21-bound to LDHA that showed that the compound binds to a distinct allosteric site between the two subunits of the LDHA tetramer. Inhibition of LDHA correlated with reduced lactate production and reduction of glycolysis in MIA PaCa-2 pancreatic cancer cells. The lead compounds inhibit the proliferation of human pancreatic cancer cell lines and patient-derived 3D organoids and exhibit a synergistic cytotoxic effect with the OXPHOS inhibitor phenformin. Unlike current LDHA inhibitors, 6 and 21 have appropriate pharmacokinetics and ligand efficiency metrics, exhibit up to 73% oral bioavailability, and a cumulative half-life greater than 4 h in mice.
PubMed: 38925013
DOI: 10.1016/j.ejmech.2024.116598 -
Annals of Clinical and Translational... Jun 2024To define tauopathy-associated changes in the human gray and white matter proteome.
OBJECTIVE
To define tauopathy-associated changes in the human gray and white matter proteome.
METHOD
We applied tandem mass tagged labeling and mass spectrometry, consensus, and ratio weighted gene correlation network analysis (WGCNA) to gray and white matter sampled from postmortem human dorsolateral prefrontal cortex. The sampled tissues included control as well as Alzheimer's disease, corticobasal degeneration, progressive supranuclear palsy, frontotemporal degeneration with tau pathology, and chronic traumatic encephalopathy.
RESULTS
Only eight proteins were unique to gray matter while six were unique to white matter. Comparison of the gray and white matter proteome revealed an enrichment of microglial proteins in the white matter. Consensus WGCNA sorted over 6700 protein isoforms into 46 consensus modules across the gray and white matter proteomic networks. Consensus network modules demonstrated unique and shared disease-associated microglial and endothelial protein changes. Ratio WGCNA sorted over 6500 protein ratios (white:gray) into 33 modules. Modules associated with mitochondrial proteins and processes demonstrated higher white:gray ratios in diseased tissues relative to control, driven by mitochondrial protein downregulation in gray and upregulation in white.
INTERPRETATION
The dataset is a valuable resource for understanding proteomic changes in human tauopathy gray and white matter. The identification of unique and shared disease-associated changes across gray and white matter emphasizes the utility of examining both tissue types. Future studies of microglial, endothelial, and mitochondrial changes in white matter may provide novel insights into tauopathy-associated changes in human brain.
PubMed: 38924699
DOI: 10.1002/acn3.52134 -
Toxins May 2024A certified reference material of ricin (CRM-LS-1) was produced by the EuroBioTox consortium to standardise the analysis of this biotoxin. This study established the...
A certified reference material of ricin (CRM-LS-1) was produced by the EuroBioTox consortium to standardise the analysis of this biotoxin. This study established the -glycan structures and proportions including their loci and occupancy of ricin CRM-LS-1. The glycan profile was compared with ricin from different preparations and other cultivars and isoforms. A total of 15 different oligomannosidic or paucimannosidic structures were identified in CRM-LS-1. Paucimannose was mainly found within the A-chain and oligomannose constituted the major glycan type of the B-chain. Furthermore, the novel primary structure variants E138 and D138 and four different C-termini of the A-chain as well as two B-chain variants V250 and F250 were elucidated. While the glycan proportions and loci were similar among all variants in CRM-LS-1 and ricin isoforms D and E of all cultivars analysed, a different stoichiometry for isoforms D and E and the amino acid variants were found. This detailed physicochemical characterization of ricin regarding the glycan profile and amino acid sequence variations yields unprecedented insight into the molecular features of this protein toxin. The variable attributes discovered within different cultivars present signature motifs and may allow discrimination of the biotoxin's origin that are important in molecular forensic profiling. In conclusion, our data of in-depth CRM-LS-1 characterization combined with the analysis of other cultivars is representative for known ricin variants.
Topics: Ricin; Polysaccharides; Reference Standards; Protein Isoforms
PubMed: 38922138
DOI: 10.3390/toxins16060243 -
Cells Jun 2024Serine/threonine kinase AKT isoforms play a well-established role in cell metabolism and growth. Most pancreatic adenocarcinomas (PDACs) harbor activation mutations of...
Serine/threonine kinase AKT isoforms play a well-established role in cell metabolism and growth. Most pancreatic adenocarcinomas (PDACs) harbor activation mutations of KRAS, which activates the PI3K/AKT signaling pathway. However, AKT inhibitors are not effective in the treatment of pancreatic cancer. To better understand the role of AKT signaling in mutant-KRAS pancreatic tumors, this study utilized proteolysis-targeting chimeras (PROTACs) and CRISPR-Cas9-genome editing to investigate AKT proteins. The PROTAC down-regulation of AKT proteins markedly slowed the growth of three pancreatic tumor cell lines harboring mutant KRAS. In contrast, the inhibition of AKT kinase activity alone had very little effect on the growth of these cell lines. The concurrent genetic deletion of all AKT isoforms (AKT1, AKT2, and AKT3) in the KPC (; ; ) pancreatic cancer cell line also dramatically slowed its growth in vitro and when orthotopically implanted in syngeneic mice. Surprisingly, insulin-like growth factor-1 (IGF-1), but not epidermal growth factor (EGF), restored KPC cell growth in serum-deprived conditions, and the IGF-1 growth stimulation effect was AKT-dependent. The RNA-seq analysis of AKT1/2/3-deficient KPC cells suggested that reduced cholesterol synthesis may be responsible for the decreased response to IGF-1 stimulation. These results indicate that the presence of all three AKT isoforms supports pancreatic tumor cell growth, and the pharmacological degradation of AKT proteins may be more effective than AKT catalytic inhibitors for treating pancreatic cancer.
Topics: Proto-Oncogene Proteins c-akt; Pancreatic Neoplasms; Animals; Cell Line, Tumor; Mice; Humans; Down-Regulation; Proto-Oncogene Proteins p21(ras); Mutation; Cell Proliferation; Signal Transduction; Gene Expression Regulation, Neoplastic
PubMed: 38920688
DOI: 10.3390/cells13121061 -
Cells Jun 2024Inflammasomes comprise a group of protein complexes with fundamental roles in the induction of inflammation. Upon sensing stress factors, their assembly induces the... (Review)
Review
Inflammasomes comprise a group of protein complexes with fundamental roles in the induction of inflammation. Upon sensing stress factors, their assembly induces the activation and release of the pro-inflammatory cytokines interleukin (IL)-1β and -18 and a lytic type of cell death, termed pyroptosis. Recently, CARD8 has joined the group of inflammasome sensors. The carboxy-terminal part of CARD8, consisting of a function-to-find-domain (FIIND) and a caspase activation and recruitment domain (CARD), resembles that of NLR family pyrin domain containing 1 (NLRP1), which is recognized as the main inflammasome sensor in human keratinocytes. The interaction with dipeptidyl peptidases 8 and 9 (DPP8/9) represents an activation checkpoint for both sensors. CARD8 and NLRP1 are activated by viral protease activity targeting their amino-terminal region. However, CARD8 also has some unique features compared to the established inflammasome sensors. Activation of CARD8 occurs independently of the inflammasome adaptor protein apoptosis-associated speck-like protein containing a CARD (ASC), leading mainly to pyroptosis rather than the activation and secretion of pro-inflammatory cytokines. CARD8 was also shown to have anti-inflammatory and anti-apoptotic activity. It interacts with, and inhibits, several proteins involved in inflammation and cell death, such as the inflammasome sensor NLRP3, CARD-containing proteins caspase-1 and -9, nucleotide-binding oligomerization domain containing 2 (NOD2), or nuclear factor kappa B (NF-κB). Single nucleotide polymorphisms (SNPs) of , some of them occurring at high frequencies, are associated with various inflammatory diseases. The molecular mechanisms underlying the different pro- and anti-inflammatory activities of CARD8 are incompletely understood. Alternative splicing leads to the generation of multiple CARD8 protein isoforms. Although the functional properties of these isoforms are poorly characterized, there is evidence that suggests isoform-specific roles. The characterization of the functions of these isoforms, together with their cell- and disease-specific expression, might be the key to a better understanding of CARD8's different roles in inflammation and inflammatory diseases.
Topics: Humans; Inflammasomes; CARD Signaling Adaptor Proteins; Apoptosis; Inflammation; Animals; Pyroptosis; Anti-Inflammatory Agents; Neoplasm Proteins
PubMed: 38920661
DOI: 10.3390/cells13121032 -
Cells Jun 2024Bone formation is a complex process regulated by a variety of pathways that are not yet fully understood. One of the proteins involved in multiple osteogenic pathways is...
Bone formation is a complex process regulated by a variety of pathways that are not yet fully understood. One of the proteins involved in multiple osteogenic pathways is TID (DNAJA3). The aim of this work was to study the association of TID with osteogenesis. Therefore, the expression profiles of the splice variants (, ) and their protein products were analyzed during the proliferation and differentiation of bone marrow mesenchymal stromal cells (B-MSCs) into osteoblasts. As the reference, the hFOB1.19 cell line was used. The phenotype of B-MSCs was confirmed by the presence of CD73, CD90, and CD105 surface antigens on ~97% of cells. The osteoblast phenotype was confirmed by increased alkaline phosphatase activity, calcium deposition, and expression of ALPL and SPP1. The effect of silencing the gene on the expression of and was also investigated. The TID proteins and the expression of splice variants were detected. After differentiation, the expression of and increased 5-fold and 3.7-fold, respectively, while their silencing resulted in increased expression of . Three days after transfection, the expression of increased 7.6-fold and 5.6-fold in B-MSCs and differentiating cells, respectively. Our preliminary study demonstrated that the expression of and changes under differentiation of B-MSCs into osteoblasts and may influence the expression of . However, for better understanding the functional association of these results with the relevant osteogenic pathways, further studies are needed.
Topics: Humans; Osteoblasts; Mesenchymal Stem Cells; Cell Differentiation; Osteogenesis; Protein Isoforms; Alkaline Phosphatase; Bone Marrow Cells; Cell Proliferation
PubMed: 38920651
DOI: 10.3390/cells13121021