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Molecular Oncology Apr 2024Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with a limited number of known driver mutations but considerable cancer cell heterogeneity....
Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with a limited number of known driver mutations but considerable cancer cell heterogeneity. Phosphoproteomics provides a direct read-out of aberrant signaling and the resultant clinically relevant phenotype. Mass spectrometry (MS)-based proteomics and phosphoproteomics were applied to 42 PDAC tumors. Data encompassed over 19 936 phosphoserine or phosphothreonine (pS/T; in 5412 phosphoproteins) and 1208 phosphotyrosine (pY; in 501 phosphoproteins) sites and a total of 3756 proteins. Proteome data identified three distinct subtypes with tumor intrinsic and stromal features. Subsequently, three phospho-subtypes were apparent: two tumor intrinsic (Phos1/2) and one stromal (Phos3), resembling known PDAC molecular subtypes. Kinase activity was analyzed by the Integrative iNferred Kinase Activity (INKA) scoring. Phospho-subtypes displayed differential phosphorylation signals and kinase activity, such as FGR and GSK3 activation in Phos1, SRC kinase family and EPHA2 in Phos2, and EGFR, INSR, MET, ABL1, HIPK1, JAK, and PRKCD in Phos3. Kinase activity analysis of an external PDAC cohort supported our findings and underscored the importance of PI3K/AKT and ERK pathways, among others. Interestingly, unfavorable patient prognosis correlated with higher RTK, PAK2, STK10, and CDK7 activity and high proliferation, whereas long survival was associated with MYLK and PTK6 activity, which was previously unknown. Subtype-associated activity profiles can guide therapeutic combination approaches in tumor and stroma-enriched tissues, and emphasize the critical role of parallel signaling pathways. In addition, kinase activity profiling identifies potential disease markers with prognostic significance.
PubMed: 38650175
DOI: 10.1002/1878-0261.13625 -
Acta Pharmaceutica Sinica. B Apr 2024In the treatment of central nervous system disease, the blood-brain barrier (BBB) is a major obstruction to drug delivery that must be overcome. In this study, we...
In the treatment of central nervous system disease, the blood-brain barrier (BBB) is a major obstruction to drug delivery that must be overcome. In this study, we propose a brain-targeted delivery strategy based on selective opening of the BBB. This strategy allows some simple bare nanoparticles to enter the brain when mixed with special opening material; however, the BBB still maintains the ability to completely block molecules from passing through. Based on the screening of BBB opening and matrix delivery materials, we determined that phospholipase A2-catalyzed 1-palmitoyl-2-oleoyl--glycero-3-phosphoserine liposomes can efficiently carry drugs into the brain immediately. At an effective dose, this delivery system is safe, especially with its effect on the BBB being reversible. This mix & act delivery system has a simple structure and rapid preparation, making it a strong potential candidate for drug delivery across the BBB.
PubMed: 38572103
DOI: 10.1016/j.apsb.2023.11.015 -
Synthetic and Systems Biotechnology Jun 2024Vitamin B plays a crucial role in cellular metabolism and stress response, making it an essential component for growth in all known organisms. However, achieving...
Vitamin B plays a crucial role in cellular metabolism and stress response, making it an essential component for growth in all known organisms. However, achieving efficient biosynthesis of vitamin B faces the challenge of maintaining a balanced distribution of metabolic flux between growth and production. In this study, our focus is on addressing this challenge through the engineering of phosphoserine aminotransferase (SerC) to resolve its redundancy and promiscuity. The enzyme SerC was semi-designed and screened based on sequences and predicted values, respectively. Mutants and heterologous proteins showing potential were then fine-tuned to optimize the production of vitamin B. The resulting strain enhances the production of vitamin B, indicating that different fluxes are distributed to the biosynthesis pathway of serine and vitamin B. This study presents a promising strategy to address the challenge posed by multifunctional enzymes, with significant implications for enhancing biochemical production through engineering processes.
PubMed: 38572022
DOI: 10.1016/j.synbio.2024.03.005 -
MedRxiv : the Preprint Server For... Mar 2024In Alzheimer's disease (AD), the most common cause of dementia, females have higher prevalence and faster progression, but sex-specific molecular findings in AD are...
In Alzheimer's disease (AD), the most common cause of dementia, females have higher prevalence and faster progression, but sex-specific molecular findings in AD are limited. Here, we comprehensively examined and validated 7,006 aptamers targeting 6,162 proteins in cerebral spinal fluid (CSF) from 2,077 amyloid/tau positive cases and controls to identify sex-specific proteomic signatures of AD. In discovery (N=1,766), we identified 330 male-specific and 121 female-specific proteomic alternations in CSF (FDR <0.05). These sex-specific proteins strongly predicted amyloid/tau positivity (AUC=0.98 in males; 0.99 in females), significantly higher than those with age, sex, and APOE-ε4 (AUC=0.85). The identified sex-specific proteins were well validated (r≥0.5) in the Stanford study (N=108) and Emory study (N=148). Biological follow-up of these proteins led to sex differences in cell-type specificity, pathways, interaction networks, and drug targets. Male-specific proteins, enriched in astrocytes and oligodendrocytes, were involved in postsynaptic and axon-genesis. The male network exhibited direct connections among 152 proteins and highlighted PTEN, NOTCH1, FYN, and MAPK8 as hubs. Drug target suggested melatonin (used for sleep-wake cycle regulation), nabumetone (used for pain), daunorubicin, and verteporfin for treating AD males. In contrast, female-specific proteins, enriched in neurons, were involved in phosphoserine residue binding including cytokine activities. The female network exhibits strong connections among 51 proteins and highlighted JUN and 14-3-3 proteins (YWHAG and YWHAZ) as hubs. Drug target suggested biperiden (for muscle control of Parkinson's disease), nimodipine (for cerebral vasospasm), quinostatin and ethaverine for treating AD females. Together, our findings provide mechanistic understanding of sex differences for AD risk and insights into clinically translatable interventions.
PubMed: 38559166
DOI: 10.1101/2024.03.15.24304164 -
BioRxiv : the Preprint Server For... Mar 2024The self-assembly of Tau(297-391) into filaments, which mirror the structures observed in Alzheimer's disease (AD) brains, raises questions about the role of AD-specific...
The self-assembly of Tau(297-391) into filaments, which mirror the structures observed in Alzheimer's disease (AD) brains, raises questions about the role of AD-specific post-translational modifications (PTMs) in the formation of paired helical filaments (PHFs). To investigate this, we developed a synthetic approach to produce Tau(291-391) featuring N-acetyllysine, phosphoserine, phosphotyrosine, and N-glycosylation at positions commonly modified in post-mortem AD brains, thus facilitating the study of their roles in Tau pathology. Using transmission electron microscopy (TEM), cryo-electron microscopy (cryo-EM), and a range of optical microscopy techniques, we discovered that these modifications generally hinder the assembly of Tau into PHFs. Interestingly, while acetylation's effect on Tau assembly displayed variability, either promoting or inhibiting phase transitions in the context of cofactor free aggregation, heparin-induced aggregation, and RNA-mediated liquid-liquid phase separation (LLPS), phosphorylation uniformly mitigated these processes. Our observations suggest that PTMs, particularly those situated outside the fibril's rigid core are pivotal in the nucleation of PHFs. Moreover, in scenarios involving heparin-induced aggregation leading to the formation of heterogeneous aggregates, most AD-specific PTMs, except for K311, appeared to decelerate the aggregation process. The impact of acetylation on RNA-induced LLPS was notably site-dependent, exhibiting both facilitative and inhibitory effects, whereas phosphorylation consistently reduced LLPS across all proteoforms examined. These insights underscore the complex interplay between site-specific PTMs and environmental factors in modulating Tau aggregation kinetics, enhancing our understanding of the molecular underpinnings of Tau pathology in AD and highlighting the critical role of PTMs located outside the ordered filament core in driving the self-assembly of Tau into PHF structures.
PubMed: 38559065
DOI: 10.1101/2024.03.08.583040 -
Molecular Genetics & Genomic Medicine Apr 2024Phosphoserine aminotransferase deficiency (PSATD) is an autosomal recessive disorder associated with hypertonia, psychomotor retardation, and acquired microcephaly.... (Review)
Review
BACKGROUND
Phosphoserine aminotransferase deficiency (PSATD) is an autosomal recessive disorder associated with hypertonia, psychomotor retardation, and acquired microcephaly. Patients with PSATD have low concentrations of serine in plasma and cerebrospinal fluid.
METHODS
We reported a 2-year-old female child with developmental delay, dyskinesia, and microcephaly. LC-MS/MS was used to detect amino acid concentration in the blood and whole-exome sequencing (WES) was used to identify the variants. PolyPhen-2 web server and PyMol were used to predict the pathogenicity and changes in the 3D model molecular structure of protein caused by variants.
RESULTS
WES demonstrated compound heterozygous variants in PSAT1, which is associated with PSATD, with a paternal likely pathogenic variant (c.235G>A, Gly79Arg) and a maternal likely pathogenic variant (c.43G>C, Ala15Pro). Reduced serine concentration in LC-MS/MS further confirmed the diagnosis of PSATD in this patient.
CONCLUSIONS
Our findings demonstrate the importance of WES combined with LC-MS/MS reanalysis in the diagnosis of genetic diseases and expand the PSAT1 variant spectrum in PSATD. Moreover, we summarize all the cases caused by PSAT1 variants in the literature. This case provides a vital reference for the diagnosis of future cases.
Topics: Child, Preschool; Female; Humans; Chromatography, Liquid; Exome Sequencing; Liquid Chromatography-Mass Spectrometry; Microcephaly; Psychomotor Disorders; Seizures; Serine; Tandem Mass Spectrometry; Transaminases
PubMed: 38546032
DOI: 10.1002/mgg3.2400 -
Genes & Development Mar 2024Metabolic reprogramming of stem cells is a targetable pathway to control regeneration. Activation of stem cells results in down-regulation of oxidative phosphorylation... (Review)
Review
Metabolic reprogramming of stem cells is a targetable pathway to control regeneration. Activation of stem cells results in down-regulation of oxidative phosphorylation (OXPHOS) and fatty acid oxidation (FAO) and turns on glycolysis to provide fuel for proliferation and specific signaling events. How cell type-specific events are regulated is unknown. In this issue of Ciuffoli and colleagues (pp. 151-167) use metabolomic, gene inactivation, and functional approaches to show that phosphoserine aminotransferase (Psat1), an enzyme in serine biosynthesis, is activated in muscle stem cells and contributes to cell expansion and skeletal muscle regeneration via the production of α-ketoglutarate and glutamine.
Topics: Muscle, Skeletal; Oxidative Phosphorylation; Glycolysis; Stem Cells; Satellite Cells, Skeletal Muscle
PubMed: 38485266
DOI: 10.1101/gad.351666.124 -
Plants (Basel, Switzerland) Feb 2024Cadmium (Cd) hampers plant growth and harms photosynthesis. Glutamate (Glu) responds to Cd stress and activates the Ca signaling pathway in duckweed, emphasizing Glu's...
Cadmium (Cd) hampers plant growth and harms photosynthesis. Glutamate (Glu) responds to Cd stress and activates the Ca signaling pathway in duckweed, emphasizing Glu's significant role in Cd stress. In this study, we overexpressed phosphoserine aminotransferase (), a crucial enzyme in Glu metabolism, in duckweed. We investigated the response of -transgenic duckweed to Cd stress, including growth, Glu metabolism, photosynthesis, antioxidant enzyme activity, Cd flux, and gene expression. Remarkably, under Cd stress, -transgenic duckweed prevented root abscission, upregulated the expression of photosynthesis ability, and increased Chl a, Chl b, and Chl a + b levels by 13.9%, 7%, and 12.6%, respectively. Antioxidant enzyme activity (CAT and SOD) also improved under Cd stress, reducing cell membrane damage in -transgenic duckweeds. Transcriptomic analysis revealed an upregulation of Glu metabolism-related enzymes in -transgenic duckweed under Cd stress. Moreover, metabolomic analysis showed a 68.4% increase in Glu content in duckweed exposed to Cd. This study sheds novel insights into the role of in enhancing plant resistance to Cd stress, establishing a theoretical basis for the impact of Glu metabolism on heavy metal tolerance in plants.
PubMed: 38475473
DOI: 10.3390/plants13050627 -
BioRxiv : the Preprint Server For... Feb 2024Nucleophosmin (NPM1) is the 46th most abundant human protein with many functions whose dysregulation leads to various cancers. Pentameric NPM1 resides in the nucleolus...
Nucleophosmin (NPM1) is the 46th most abundant human protein with many functions whose dysregulation leads to various cancers. Pentameric NPM1 resides in the nucleolus but can also shuttle to the cytosol. NPM1 is regulated by multisite phosphorylation, yet molecular consequences of site-specific NPM1 phosphorylation remain elusive. Here we identify four 14-3-3 protein binding sites in NPM1 concealed within its oligomerization and α-helical C-terminal domains that are found phosphorylated . By combining mutagenesis, in-cell phosphorylation and PermaPhos technology for site-directed incorporation of a non-hydrolyzable phosphoserine mimic, we show how phosphorylation promotes NPM1 monomerization and partial unfolding, to recruit 14-3-3 dimers with low-micromolar affinity. Using fluorescence anisotropy we quantified pairwise interactions of all seven human 14-3-3 isoforms with four recombinant NPM1 phosphopeptides and assessed their druggability by fusicoccin. This revealed a complex hierarchy of 14-3-3 affinities toward the primary (S48, S293) and secondary (S106, S260) sites, differentially modulated by the small molecule. As three of these 14-3-3 binding phospho-sites in NPM1 reside within signal sequences, this work highlights a key mechanism of NPM1 regulation by which NPM1 phosphorylation promotes 14-3-3 binding to control nucleocytoplasmic shuttling. It also provides further evidence that phosphorylation-induced structural rearrangements of globular proteins serve to expose otherwise cryptic 14-3-3-binding sites that are important for cellular function.
PubMed: 38405961
DOI: 10.1101/2024.02.13.580064 -
Nucleic Acids Research Apr 2024Protein-protein and protein-rRNA interactions at the interface between ribosomal proteins uS4 and uS5 are thought to maintain the accuracy of protein synthesis by...
An evolutionarily conserved phosphoserine-arginine salt bridge in the interface between ribosomal proteins uS4 and uS5 regulates translational accuracy in Saccharomyces cerevisiae.
Protein-protein and protein-rRNA interactions at the interface between ribosomal proteins uS4 and uS5 are thought to maintain the accuracy of protein synthesis by increasing selection of cognate aminoacyl-tRNAs. Selection involves a major conformational change-domain closure-that stabilizes aminoacyl-tRNA in the ribosomal acceptor (A) site. This has been thought a constitutive function of the ribosome ensuring consistent accuracy. Recently, the Saccharomyces cerevisiae Ctk1 cyclin-dependent kinase was demonstrated to ensure translational accuracy and Ser238 of uS5 proposed as its target. Surprisingly, Ser238 is outside the uS4-uS5 interface and no obvious mechanism has been proposed to explain its role. We show that the true target of Ctk1 regulation is another uS5 residue, Ser176, which lies in the interface opposite to Arg57 of uS4. Based on site specific mutagenesis, we propose that phospho-Ser176 forms a salt bridge with Arg57, which should increase selectivity by strengthening the interface. Genetic data show that Ctk1 regulates accuracy indirectly; the data suggest that the kinase Ypk2 directly phosphorylates Ser176. A second kinase pathway involving TORC1 and Pkc1 can inhibit this effect. The level of accuracy appears to depend on competitive action of these two pathways to regulate the level of Ser176 phosphorylation.
Topics: Saccharomyces cerevisiae Proteins; Saccharomyces cerevisiae; Ribosomal Proteins; Arginine; Protein Biosynthesis; Phosphoserine; Cyclin-Dependent Kinases; Phosphorylation; Evolution, Molecular; Protein Kinases
PubMed: 38340338
DOI: 10.1093/nar/gkae053