Recent studies have highlighted the significant role of 5-hydroxymethylcytosine (5hmC) in carcinogenesis. However, the specific role of 5hmC in osteosarcoma (OS) remains largely unexplored. The-re-fore, this study aimed to investigate the function of 5hmC and TET3 in OS. In this study, we found a decreased total level of 5hmC in OS tissues. The expression of the TET3 protein was also decreased in OS. Importantly, the decreased levels of TET3 were associated with a decreased disease-free survival (DFS) rate in patients. To investigate the role of TET3 and 5hmC in OS, we manipulated the levels of TET3 in MG-63 cells. Silencing TET3 in these cells resulted in a twofold increase in proliferation. Additio-nally, the level of 5hmC decreased in these cells. Con-versely, over-expression of TET3 in MG-63 cells led to the expected inhibition of proliferation and invasion, accompanied by an increase in 5hmC levels. In conclusion, both 5hmC and TET3 protein levels were decreased in OS. Additionally, the over-expression of TET3 inhibited the proliferation of MG-63 cells, while the suppression of TET3 had the opposite effect. These findings suggest that decreased levels of 5hmC and TET3 may serve as potential markers for OS.

• MeSH
• 5-methylcytosin * analogy a deriváty   metabolismus   MeSH
• demetylace DNA * MeSH
• dioxygenasy * metabolismus   MeSH
• epigeneze genetická * MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádory kostí genetika   metabolismus   patologie   MeSH
• osteosarkom genetika   metabolismus   patologie   MeSH
• proliferace buněk * MeSH
• protoonkogenní proteiny metabolismus   genetika   MeSH
• regulace genové exprese u nádorů MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Defective mitophagy is consistently found in postmortem brain and iPSC-derived neurons from Alzheimer disease (AD) patients. However, there is a lack of extensive examination of mitophagy status in serum or cerebrospinal fluid (CSF), and the clinical potential of mitophagy biomarkers has not been tested. We quantified biomarkers of mitophagy/autophagy and lysosomal degradation (PINK1, BNIP3L and TFEB) in CSF and serum from 246 individuals, covering mild cognitive impairment due to AD (MCI-AD, n = 100), dementia due to AD (AD-dementia, n = 100), and cognitively unimpaired individuals (CU, n = 46), recruited from the Czech Brain Aging Study. Cognitive function and brain atrophy were also assessed. Our data show that serum and CSF PINK1 and serum BNIP3L were higher, and serum TFEB was lower in individuals with AD than in corresponding CU individuals. Additionally, the magnitude of mitophagy impairment correlated with the severity of clinical indicators in AD patients. Specifically, levels of PINK1 positively correlated with phosphorylated (p)-MAPT/tau (181), total (t)-MAPT/tau, NEFL (neurofilament light chain), and NRGN (neurogranin) levels in CSF and negatively with memory, executive function, and language domain. Serum TFEB levels negatively correlated with NEFL and positively with executive function and language. This study reveals mitophagy impairment reflected in biofluid biomarkers of individuals with AD and associated with more advanced AD pathology.Abbreviation: Aβ: amyloid beta; AD: Alzheimer disease; AVs: autophagic vacuoles; BNIP3L: BCL2 interacting protein 3 like; CU: cognitively unimpaired; CSF: cerebrospinal fluid; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCI: mild cognitive impairment; NRGN: neurogranin; NEFL: neurofilament light chain; p-MAPT/tau: phosphorylated microtubule associated protein tau; PINK1: PTEN induced kinase 1; t-MAPT/tau: total microtubule associated protein tau; TFEB: transcription factor EB; TMT: Trail Making Test.

• MeSH
• Alzheimerova nemoc * mozkomíšní mok   krev   diagnóza   MeSH
• biologické markery * mozkomíšní mok   krev   metabolismus   MeSH
• kognitivní dysfunkce mozkomíšní mok   krev   diagnóza   MeSH
• lidé středního věku MeSH
• lidé MeSH
• membránové proteiny mozkomíšní mok   metabolismus   krev   MeSH
• mitofagie * MeSH
• mozek metabolismus   patologie   MeSH
• nádorové supresorové proteiny MeSH
• proteinkinasy metabolismus   MeSH
• proteiny tau mozkomíšní mok   metabolismus   MeSH
• protoonkogenní proteiny mozkomíšní mok   krev   metabolismus   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• transkripční faktory BHLH-Zip metabolismus   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Diagnosis of soft tissue tumors is often challenging, given the large number of entities, often with non-specific or overlapping morphology. Although morphology still plays an important part in diagnostic process, additional studies including immunohistochemistry and molecular genetics are often needed to arrive at correct diagnosis. We report a case of 61-year-old male with subcutaneous tumor in right hip area, that was surgically removed. The tumor was composed of uniform bland spindle cells in mild to moderately cellular myxoid nodules, with limited areas of collagenization and the diagnosis of low grade fibromyxoid sarcoma was made. The tumor recurred 3 years after the initial diagnosis and the new sample showed a high-grade round cell sarcoma with limited residual low-grade areas and non-specific immunoprofile after extended immunohistochemical work-up. Molecular analysis demonstrated ZC3H7B::BCOR fusion. Sarcomas with ZC3H7B::BCOR fusion occurring outside of uterus are exceedingly rare. A comprehensive review of previously published cases and a short discussion about classification of the entity is provided, together with data about morphology and immunoprofile of the lesions. The case also underscores the necessity of extended work up of soft tissue tumors with unusual immunohistochemical or morphological features in order to accurately assess their biological potential.

• MeSH
• fibrosarkom * diagnóza   MeSH
• lidé středního věku MeSH
• lidé MeSH
• lokální recidiva nádoru MeSH
• nádorové biomarkery analýza   MeSH
• nádory měkkých tkání * diagnóza   patologie   MeSH
• proteiny vázající RNA MeSH
• protoonkogenní proteiny metabolismus   MeSH
• represorové proteiny metabolismus   MeSH
• sarkom * patologie   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• kazuistiky MeSH
• přehledy MeSH

The transcription factor PU.1 (Purine-rich DNA binding, SPI1) is a key regulator of hematopoiesis, whose level is influenced by transcription through its enhancers and its post-transcriptional degradation via microRNA-155 (miR-155). The degree of transcriptional regulation of the PU.1 gene is influenced by repression via DNA methylation, as well as other epigenetic factors, such as those related to progenitor maturation status, which is modulated by the transcription factor Myeloblastosis oncogene (MYB). In this work, we show that combinatorial treatment of acute myeloid leukemia (AML) cells with DNA methylation inhibitors (5-Azacytidine), MYB inhibitors (Celastrol), and anti-miR-155 (AM155) ideally leads to overproduction of PU.1. We also show that PU.1 reactivation can be compensated by miR-155 and that only a combined approach leads to sustained PU.1 derepression, even at the protein level. The triple effect on increasing PU.1 levels in myeloblasts stimulates the myeloid transcriptional program while inhibiting cell survival and proliferation, leading to partial leukemic differentiation.

• MeSH
• akutní myeloidní leukemie * farmakoterapie   genetika   MeSH
• buněčná diferenciace genetika   MeSH
• lidé MeSH
• mikro RNA * genetika   metabolismus   MeSH
• protoonkogenní proteiny genetika   metabolismus   MeSH
• regulace genové exprese u leukemie MeSH
• trans-aktivátory metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Although human nucleoporin Tpr is frequently deregulated in cancer, its roles are poorly understood. Here we show that Tpr depletion generates transcription-dependent replication stress, DNA breaks, and genomic instability. DNA fiber assays and electron microscopy visualization of replication intermediates show that Tpr deficient cells exhibit slow and asymmetric replication forks under replication stress. Tpr deficiency evokes enhanced levels of DNA-RNA hybrids. Additionally, complementary proteomic strategies identify a network of Tpr-interacting proteins mediating RNA processing, such as MATR3 and SUGP2, and functional experiments confirm that their depletion trigger cellular phenotypes shared with Tpr deficiency. Mechanistic studies reveal the interplay of Tpr with GANP, a component of the TREX-2 complex. The Tpr-GANP interaction is supported by their shared protein level alterations in a cohort of ovarian carcinomas. Our results reveal links between nucleoporins, DNA transcription and replication, and the existence of a network physically connecting replication forks with transcription, splicing, and mRNA export machinery.

• MeSH
• acetyltransferasy genetika   metabolismus   MeSH
• HeLa buňky MeSH
• intracelulární signální peptidy a proteiny genetika   metabolismus   MeSH
• komplex proteinů jaderného póru genetika   metabolismus   MeSH
• lidé MeSH
• mapy interakcí proteinů MeSH
• nádory genetika   MeSH
• nestabilita genomu MeSH
• poškození DNA MeSH
• protoonkogenní proteiny genetika   metabolismus   MeSH
• replikace DNA * MeSH
• transport RNA MeSH
• viabilita buněk MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The nuclear pore complex (NPC) has emerged as a hub for the transcriptional regulation of a subset of genes, and this type of regulation plays an important role during differentiation. Nucleoporin TPR forms the nuclear basket of the NPC and is crucial for the enrichment of open chromatin around NPCs. TPR has been implicated in the regulation of transcription; however, the role of TPR in gene expression and cell differentiation has not been described. Here we show that depletion of TPR results in an aberrant morphology of murine proliferating C2C12 myoblasts (MBs) and differentiated C2C12 myotubes (MTs). The ChIP-Seq data revealed that TPR binds to genes linked to muscle formation and function, such as myosin heavy chain (Myh4), myocyte enhancer factor 2C (Mef2C) and a majority of olfactory receptor (Olfr) genes. We further show that TPR, possibly via lysine-specific demethylase 1 (LSD1), promotes the expression of Myh4 and Olfr376, but not Mef2C. This provides a novel insight into the mechanism of myogenesis; however, more evidence is needed to fully elucidate the mechanism by which TPR affects specific myogenic genes.

• MeSH
• buněčná diferenciace MeSH
• buněčné linie MeSH
• exprese genu MeSH
• komplex proteinů jaderného póru metabolismus   MeSH
• kosterní svalová vlákna * cytologie   metabolismus   MeSH
• myoblasty kosterní * cytologie   metabolismus   MeSH
• myši MeSH
• protoonkogenní proteiny metabolismus   MeSH
• regulace genové exprese MeSH
• těžké řetězce myosinu metabolismus   MeSH
• vývoj svalů MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Cyclin A2 is a key regulator of the cell cycle, implicated both in DNA replication and mitotic entry. Cyclin A2 participates in feedback loops that activate mitotic kinases in G2 phase, but why active Cyclin A2-CDK2 during the S phase does not trigger mitotic kinase activation remains unclear. Here, we describe a change in localisation of Cyclin A2 from being only nuclear to both nuclear and cytoplasmic at the S/G2 border. We find that Cyclin A2-CDK2 can activate the mitotic kinase PLK1 through phosphorylation of Bora, and that only cytoplasmic Cyclin A2 interacts with Bora and PLK1. Expression of predominately cytoplasmic Cyclin A2 or phospho-mimicking PLK1 T210D can partially rescue a G2 arrest caused by Cyclin A2 depletion. Cytoplasmic presence of Cyclin A2 is restricted by p21, in particular after DNA damage. Cyclin A2 chromatin association during DNA replication and additional mechanisms contribute to Cyclin A2 localisation change in the G2 phase. We find no evidence that such mechanisms involve G2 feedback loops and suggest that cytoplasmic appearance of Cyclin A2 at the S/G2 transition functions as a trigger for mitotic kinase activation.

• MeSH
• aktivace enzymů genetika   MeSH
• buněčné jádro metabolismus   MeSH
• chromatin metabolismus   MeSH
• cyklin A2 genetika   metabolismus   MeSH
• cyklin-dependentní kinasa 2 nedostatek   genetika   MeSH
• cytoplazma metabolismus   MeSH
• fosforylace genetika   MeSH
• G2 fáze genetika   MeSH
• HeLa buňky MeSH
• lidé MeSH
• mitóza genetika   MeSH
• poškození DNA genetika   MeSH
• protein-serin-threoninkinasy metabolismus   MeSH
• proteinkinasa CDC2 nedostatek   genetika   MeSH
• proteiny buněčného cyklu metabolismus   MeSH
• protoonkogenní proteiny metabolismus   MeSH
• S fáze genetika   MeSH
• signální transdukce genetika   MeSH
• transfekce MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Selection of non-small-cell lung cancer patients for treatment relies on the detection of expression of anaplastic lymphoma kinase (ALK) and ROS proto-oncogene 1 (ROS1) protein by immunohistochemistry (IHC). We evaluated staining performance for different IHC protocols and laboratory characteristics, and their influence on ALK and ROS1 interpretation during external quality assessment schemes between 2015 and 2018. Participants received five formalin-fixed, paraffin-embedded cases for staining by their routine protocol, whereafter at least two pathologists scored them simultaneously under a multihead microscope and awarded a graded expert staining score (ESS) from 1 to 5 points based on staining quality. European Conformity in Vitro Diagnostic kits (such as D5F3) revealed a better ALK ESS compared with laboratory-developed tests. ESS was indifferent to the applied antibody dilution or a recent protocol change. Lower ESSs were observed for higher antibody incubation times and temperatures. ESS for various ROS1 protocols were largely similar. Overall, for both markers, ESS improved over time and for repeated external quality assessment participation but was independent of laboratory setting or experience. Except for ROS1, ESS positively correlated with laboratory accreditation. IHC stains with lower ESS correlated with increased error rates in ALK and ROS1 interpretation and analysis failures. Laboratory characteristics differently affected staining quality and interpretation, and laboratories should assess both aspects, and less common protocols need improvement in staining performance.

• MeSH
• anaplastická lymfomová kináza metabolismus   MeSH
• hybridizace in situ fluorescenční metody   MeSH
• imunohistochemie metody   MeSH
• laboratoře nemocniční MeSH
• lidé MeSH
• nádory plic metabolismus   MeSH
• nemalobuněčný karcinom plic metabolismus   MeSH
• patologové MeSH
• protoonkogen Mas MeSH
• protoonkogenní proteiny metabolismus   MeSH
• senzitivita a specificita MeSH
• tyrosinkinasy metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH

The identification of the essential role of cyclin-dependent kinases (CDKs) in the control of cell division has prompted the development of small-molecule CDK inhibitors as anticancer drugs. For many of these compounds, the precise mechanism of action in individual tumor types remains unclear as they simultaneously target different classes of CDKs - enzymes controlling the cell cycle progression as well as CDKs involved in the regulation of transcription. CDK inhibitors are also capable of activating p53 tumor suppressor in tumor cells retaining wild-type p53 gene by modulating MDM2 levels and activity. In the current study, we link, for the first time, CDK activity to the overexpression of the MDM4 (MDMX) oncogene in cancer cells. Small-molecule drugs targeting the CDK9 kinase, dinaciclib, flavopiridol, roscovitine, AT-7519, SNS-032, and DRB, diminished MDM4 levels and activated p53 in A375 melanoma and MCF7 breast carcinoma cells with only a limited effect on MDM2. These results suggest that MDM4, rather than MDM2, could be the primary transcriptional target of pharmacological CDK inhibitors in the p53 pathway. CDK9 inhibitor atuveciclib downregulated MDM4 and enhanced p53 activity induced by nutlin-3a, an inhibitor of p53-MDM2 interaction, and synergized with nutlin-3a in killing A375 melanoma cells. Furthermore, we found that human pluripotent stem cell lines express significant levels of MDM4, which are also maintained by CDK9 activity. In summary, we show that CDK9 activity is essential for the maintenance of high levels of MDM4 in human cells, and drugs targeting CDK9 might restore p53 tumor suppressor function in malignancies overexpressing MDM4.

• MeSH
• cyklin-dependentní kinasa 9 antagonisté a inhibitory   metabolismus   MeSH
• genetická transkripce MeSH
• imidazoly farmakologie   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• lidé MeSH
• melanom genetika   metabolismus   patologie   MeSH
• MFC-7 buňky MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádory prsu genetika   metabolismus   patologie   MeSH
• piperaziny farmakologie   MeSH
• pluripotentní kmenové buňky metabolismus   MeSH
• proteiny buněčného cyklu biosyntéza   genetika   metabolismus   MeSH
• protoonkogenní proteiny c-mdm2 biosyntéza   genetika   metabolismus   MeSH
• protoonkogenní proteiny biosyntéza   genetika   metabolismus   MeSH
• roskovitin farmakologie   MeSH
• sulfonamidy farmakologie   MeSH
• synergismus léků MeSH
• transfekce MeSH
• triaziny farmakologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Spliceosomal small nuclear ribonucleoprotein particles (snRNPs) undergo a complex maturation pathway containing multiple steps in the nucleus and in the cytoplasm. snRNP biogenesis is strictly proofread and several quality control checkpoints are placed along the pathway. Here, we analyzed the fate of small nuclear RNAs (snRNAs) that are unable to acquire a ring of Sm proteins. We showed that snRNAs lacking the Sm ring are unstable and accumulate in P-bodies in an LSm1-dependent manner. We further provide evidence that defective snRNAs without the Sm binding site are uridylated at the 3' end and associate with DIS3L2 3'→5' exoribonuclease and LSm proteins. Finally, inhibition of 5'→3' exoribonuclease XRN1 increases association of ΔSm snRNAs with DIS3L2, which indicates competition and compensation between these two degradation enzymes. Together, we provide evidence that defective snRNAs without the Sm ring are uridylated and degraded by alternative pathways involving either DIS3L2 or LSm proteins and XRN1.

• MeSH
• exoribonukleasy metabolismus   MeSH
• HeLa buňky MeSH
• konformace nukleové kyseliny * MeSH
• lidé MeSH
• organely metabolismus   MeSH
• proteinový komplex SMN metabolismus   MeSH
• proteiny vázající RNA metabolismus   MeSH
• protoonkogenní proteiny metabolismus   MeSH
• RNA malá jaderná chemie   metabolismus   MeSH
• sekvence nukleotidů MeSH
• stabilita RNA MeSH
• transport RNA * MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH