VRK1 Regulates Sensitivity to Oxidative Stress by Altering Histone Epigenetic Modifications and the Nuclear Phosphoproteome in Tumor Cells.

The chromatin organization and its dynamic remodeling determine its accessibility and sensitivity to DNA damage oxidative stress, the main source of endogenous DNA damage. We studied the role of the VRK1 chromatin kinase in the response to oxidative stress. which alters the nuclear pattern of histone epigenetic modifications and phosphoproteome pathways. The early effect of oxidative stress on chromatin was studied by determining the levels of 8-oxoG lesions and the alteration of the epigenetic modification of histones. Oxidative stress caused an accumulation of 8-oxoG DNA lesions that were increased by VRK1 depletion, causing a significant accumulation of DNA strand breaks detected by labeling free 3'-DNA ends. In addition, oxidative stress altered the pattern of chromatin epigenetic marks and the nuclear phosphoproteome pathways that were impaired by VRK1 depletion. Oxidative stress induced the acetylation of H4K16ac and H3K9 and the loss of H3K4me3. The depletion of VRK1 altered all these modifications induced by oxidative stress and resulted in losses of H4K16ac and H3K9ac and increases in the H3K9me3 and H3K4me3 levels. All these changes were induced by the oxidative stress in the epigenetic pattern of histones and impaired by VRK1 depletion, indicating that VRK1 plays a major role in the functional reorganization of chromatin in the response to oxidative stress. The analysis of the nuclear phosphoproteome in response to oxidative stress detected an enrichment of the phosphorylated proteins associated with the chromosome organization and chromatin remodeling pathways, which were significantly decreased by VRK1 depletion. VRK1 depletion alters the histone epigenetic pattern and nuclear phosphoproteome pathways in response to oxidative stress. The enzymes performing post-translational epigenetic modifications are potential targets in synthetic lethality strategies for cancer therapies.

Pathogenic effects of Leu200Pro and Arg387His VRK1 protein variants on phosphorylation targets and H4K16 acetylation in distal hereditary motor neuropathy.

Rare recessive variants in the human VRK1 gene are associated with several motor neuron diseases (MND), such as amyotrophic lateral sclerosis, spinal muscular atrophy, or distal hereditary motor neuropathies (dHMN). A case with dHMN carrying two novel VRK1 gene variants, expressing Leu200Pro (L200P) and Arg387His (R387H) variant proteins, identified that these protein variants are functionally different. The Leu200Pro variant shares with several variants in the catalytic domain the loss of the kinase activity on different substrates, such as histones, p53, or coilin. However, the distal Arg387His variant and the distal Trp375* (W375X) chinese variant, both located at the end of the low complexity C-terminal region and proximal to the termination codon, retain their catalytic activity on some substrates, and mechanistically their functional impairment is different. The L200P variant, as well as most VRK1 pathogenic variants, impairs the phosphorylation of BAF and histone H4K16 acetylation, which are required for DNA attachment to the nuclear envelope and chromatin accessibility to DNA repair mechanisms, respectively. The R387H variant impairs phosphorylation of H2AX, an early step in different types of DNA damage responses. The functional variability of VRK1 protein variants and their different combinations are a likely contributor to the clinical phenotypic heterogeneity of motor neuron and neurological diseases associated with rare VRK1 pathogenic variants. KEY MESSAGES: VRK1 variants implicated in motor neuron diseases are functionally different. The L200P variant is kinase inactive, and the R387H variant is partially active. VRK1 variants alter H4K16 acetylation and loss of coilin and BAF phosphorylation. VRK1 variants alter Cajal bodies and DNA damage responses. VRK1 variant combination determines the neurological phenotype heterogeneity.

Loss of VRK1 alters the nuclear phosphoproteome in the DNA damage response to doxorubicin.

Dynamic chromatin remodeling requires regulatory mechanisms for its adaptation to different nuclear function, which are likely to be mediated by signaling proteins. In this context, VRK1 is a nuclear Ser-Thr kinase that regulates pathways associated with transcription, replication, recombination, and DNA repair. Therefore, VRK1 is a potential regulatory, or coordinator, molecule in these processes. In this work we studied the effect that VRK1 depletion has on the basal nuclear and chromatin phosphoproteome, and their associated pathways. VRK1 depletion caused an alteration in the pattern of the nuclear phosphoproteome, which is mainly associated with nucleoproteins, ribonucleoproteins, RNA splicing and processing. Next, it was determined the changes in proteins associated with DNA damage that was induced by doxorubicin treatment. Doxorubicin alters the nuclear phosphoproteome affecting proteins implicated in DDR, including DSB repair proteins NBN and 53BP1, cellular response to stress and chromatin organization proteins. In VRK1-depleted cells, the effect of doxorubicin on protein phosphorylation was reverted to basal levels. The nuclear phosphoproteome patterns induced by doxorubicin are altered by VRK1 depletion, and is enriched in histone modification proteins and chromatin associated proteins. These results indicate that VRK1 plays a major role in processes requiring chromatin remodeling in its adaptation to different biological contexts.

The pattern of histone H3 epigenetic posttranslational modifications is regulated by the VRK1 chromatin kinase.

BACKGROUND: Dynamic chromatin remodeling is associated with changes in the epigenetic pattern of histone acetylations and methylations required for processes based on dynamic chromatin remodeling and implicated in different nuclear functions. These histone epigenetic modifications need to be coordinated, a role that may be mediated by chromatin kinases such as VRK1, which phosphorylates histones H3 and H2A. METHODS: The effect of VRK1 depletion and VRK1 inhibitor, VRK-IN-1, on the acetylation and methylation of histone H3 in K4, K9 and K27 was determined under different conditions, arrested or proliferating cells, in A549 lung adenocarcinoma and U2OS osteosarcoma cells. RESULTS: Chromatin organization is determined by the phosphorylation pattern of histones mediated by different types of enzymes. We have studied how the VRK1 chromatin kinase can alter the epigenetic posttranslational modifications of histones by using siRNA, a specific inhibitor of this kinase (VRK-IN-1), and of histone acetyl and methyl transferases, as well as histone deacetylase and demethylase. Loss of VRK1 implicated a switch in the state of H3K9 posttranslational modifications. VRK1 depletion/inhibition causes a loss of H3K9 acetylation and facilitates its methylation. This effect is similar to that of the KAT inhibitor C646, and to KDM inhibitors as iadademstat (ORY-1001) or JMJD2 inhibitor. Alternatively, HDAC inhibitors (selisistat, panobinostat, vorinostat) and KMT inhibitors (tazemetostat, chaetocin) have the opposite effect of VRK1 depletion or inhibition, and cause increase of H3K9ac and a decrease of H3K9me3. VRK1 stably interacts with members of these four enzyme families. However, VRK1 can only play a role on these epigenetic modifications by indirect mechanisms in which these epigenetic enzymes are likely targets to be regulated and coordinated by VRK1. CONCLUSIONS: The chromatin kinase VRK1 regulates the epigenetic patterns of histone H3 acetylation and methylation in lysines 4, 9 and 27. VRK1 is a master regulator of chromatin organization associated with its specific functions, such as transcription or DNA repair.

Evaluation of Low-Toxic Hybrid Sol-Gel Coatings with Organic pH-Sensitive Inhibitors for Corrosion Protection of AA2024 Aluminium Alloy.

Today's environmental needs require the reduction of the weight of vehicles, thus reducing fuel consumption and associated emissions. For this reason, the use of light alloys is being studied, which, due to their reactivity, must be protected before use. In this work, the effectiveness of a hybrid sol-gel coating doped with various organic environmentally friendly corrosion inhibitors applied to an AA2024 lightweight aluminium alloy is evaluated. Some of the inhibitors tested are pH indicators, acting as both corrosion inhibitors and optical sensors for the surface of the alloy. Samples are subjected to a corrosion test in a simulated saline environment and characterised before and after the test. The experimental results regarding their best inhibitor performance for their potential application in the transport industry are evaluated.

VRK1 Kinase Activity Modulating Histone H4K16 Acetylation Inhibited by SIRT2 and VRK-IN-1.

The accessibility of DNA to different cellular functions requires a dynamic regulation of chromatin organization that is mediated by different epigenetic modifications, which regulate chromatin accessibility and degree of compaction. These epigenetic modifications, particularly the acetylation of histone H4 in lysine 14 (H4K16ac), determine the degree of chromatin accessibility to different nuclear functions, as well as to DNA damage drugs. H4K16ac is regulated by the balance between two alternative histone modifications, acetylation and deacetylation, which are mediated by acetylases and deacetylases. Tip60/KAT5 acetylates, and SIRT2 deacetylates histone H4K16. However, the balance between these two epigenetic enzymes is unknown. VRK1 regulates the level of H4K16 acetylation by activating Tip60. We have shown that the VRK1 and SIRT2 are able to form a stable protein complex. For this work, we used in vitro interaction, pull-down and in vitro kinase assays. In cells, their interaction and colocalization were detected by immunoprecipitation and immunofluorescence. The kinase activity of VRK1 is inhibited by a direct interaction of its N-terminal kinase domain with SIRT2 in vitro. This interaction causes a loss of H4K16ac similarly to the effect of a novel VRK1 inhibitor (VRK-IN-1) or VRK1 depletion. The use of specific SIRT2 inhibitors in lung adenocarcinoma cells induces H4K16ac, contrary to the novel VRK-IN-1 inhibitor, which prevents H4K16ac and a correct DNA damage response. Therefore, the inhibition of SIRT2 can cooperate with VRK1 in the accessibility of drugs to chromatin in response to DNA damage caused by doxorubicin.

Duodenal pyogenic granuloma: An unusual cause of chronic anemia.

We present the endoscopic finding of a juxtapapillary duodenal pyogenic granuloma in a patient under study for chronic recurrent anemia. The images show the presence of an enlarged papilla, with an exophytic growth of erythematous and friable tissue at its lower border. Although this is a very rare entity, it is necessary to take it into account in the differential diagnosis of refractory anemia because there is an effective endoscopic treatment for it.

Diagnosis of a cystic pancreatic neuroendocrine tumor: is cystic fluid chromogranin A useful?

Cystic pancreatic neuroendocrine tumors (cP-NET) are a diagnostic challenge for clinic, since sometimes nor imaging features, cytology, or the study of biological markers in pancreatic cyst fluid (PCF), are able to provide the nature of the lesion, and therefore the definitive diagnostic is often made in the surgical piece. Is not infrequent that the lesions are wrongly defined as benign, in special when the cytology is negative for malignancy and the PCL is not mucinous and has a high glucose and a low CEA. We could incorporate new markers to improve the diagnostic performance of PCF samples, like chromogranin A, as in our case may be the only specific finding to detect a cP-TNE.

The VRK1 chromatin kinase regulates the acetyltransferase activity of Tip60/KAT5 by sequential phosphorylations in response to DNA damage.

The regulation of histone epigenetic modifications mediates the adaptation of chromatin to different biological processes. DNA damage causes a local relaxation of chromatin associated to histone H4 acetylation in K16, mediated by Tip60/KAT5. In this work, we have studied the role that the VRK1 chromatin kinase plays on the activation of Tip60 during this process. In the DNA damage response induced by doxorubicin, VRK1 directly phosphorylates Tip60. However, the phosphorylated Tip60 residues and their functional roles are unknown. In DDR, we have identified these two Tip60 phosphorylated residues and the cooperation of the participating kinases. The T158 phosphorylation, mediated by VRK1, is early and transient, preceding that of S199, which is more sustained in time, and mediated by DNA-PK. The role of each phosphorylated residues was determined by using phosphomimetic and phosphonull mutants and their combination. T158 phosphorylation protects Tip60 from ubiquitin-mediated degradation, promotes its recruitment to chromatin from the nucleoplasm, and is necessary for its full trans-acetylase activity. The phosphorylation in S199 by DNA-PK directly facilitates Tip60 autoacetylation, but it is not enough for trans-acetylation of two of its targets, histone H4 and ATM, which requires a double phosphorylation of Tip60 in T158 and S199. DNA-PK inhibitors block the phosphorylation of S199. We propose a model in which the cooperation between VRK1 and DNA-PK mediates the sequential phosphorylation of Tip60/KAT5, and contributes to the recruitment of this protein to initiate the sequential remodeling of chromatin in DDR. Both proteins are candidates for novel synthetic lethality strategies in cancer treatment.

Mycoplasma pneumoniae: características clínicas diferenciales y dificultades diagnósticas de las neumonías atípicas en niños / Mycoplasma pneumoniae: differential clinical features and diagnostic challenges in atypical pneumonias in children

Introducción: la neumonía por Mycoplasma pneumoniae continúa estando infradiagnosticada en las consultas de Pediatría de Atención Primaria, especialmente en los niños más pequeños. Material y métodos: estudio prospectivo en 9 cupos pediátricos, sobre niños desde 1 mes a 14 años con neumonía diagnosticada mediante radiografía y clínica compatible. Diagnóstico etiológico mediante serología en la fase aguda. Se analiza la relación de diferentes variables con la etiología atípica y no atípica. Resultados: de los 92 pacientes incluidos, un 30,4% fueron neumonías atípicas que, a pesar de no ser raras en menores de 2 años (26% del total a esta edad), predominaron en los mayores de 5 años (80,9%). La edad media en meses fue significativamente mayor en atípicas (74,2 ± 42,2), que en las no atípicas (35,9 ± 33,8; p <0,0001). La congestión nasal (42,8%; OR: 1,8; p <0,01) y la taquipnea (64,2%; OR: 2,4; p <0,014) fueron significativamente más frecuentes en las no atípicas. El patrón alveolar se observó en el 53,6% de las neumonías atípicas, sin diferencias con las no atípicas. Solamente un 25% de las neumonías atípicas fueron correctamente tratadas inicialmente con macrólidos en monoterapia sin existir diferencias en cuanto a su evolución con respecto a la elección de un tipo u otro de terapia antibiótica. Las neumonías no atípicas precisaron antibioterapia intravenosa con una mayor frecuencia (15,6%) de forma no estadísticamente significativa. Conclusión: las neumonías por gérmenes atípicos parecen más prevalentes en niños pequeños de lo previamente descrito, en ocasiones en coexistencia con virus. Su mejor diagnóstico y tratamiento continúan siendo un reto por resolver (AU)

Introduction: pneumonia caused by Mycoplasma pneumoniae continues to be underdiagnosed in paediatric primary care, especially in younger children.Material and methods: prospective study conducted in 9 primary care paediatric caseloads, including children aged 1 month to 14 years with pneumonia diagnosed based on compatible radiographic findings and clinical features. The aetiological diagnosis was made by acute-phase serological testing. We analysed the association of different variables with atypical and typical aetiologies.Results: of the 92 patients in the sample, 30.4% had atypical pneumonias which, while not rare in children under 2 years (26%) predominated in children over 5 years (80.9%). The mean age in months was significantly higher in cases with an atypical (74.2±42.2) versus typical (35.9±33.8) aetiology (p<0.0001). Nasal congestion (42.8%; OR 1.8; p<0.01) and tachypnoea (64.2%; OR 2.4; p<0.014) were significantly more frequent in typical pneumonia. The alveolar pattern was observed in 53.6% of atypical pneumonias, with no differences compared to typical pneumonias. Only 25% of atypical pneumonia cases were treated correctly with first-line macrolide monotherapy, with no differences in outcomes based on the choice of antibiotherapy. Patients with typical pneumonia required intravenous antibiotic therapy more frequently (15.6%), but the difference was not statistically significant.Conclusion: atypical germs were more frequent at younger ages than previously described, in some cases with concomitant detection of viruses. Improving the diagnosis and treatment of atypical pneumonia remains a challenge. (AU)

Lysine Methyltransferase Inhibitors Impair H4K20me2 and 53BP1 Foci in Response to DNA Damage in Sarcomas, a Synthetic Lethality Strategy.

BACKGROUND: Chromatin is dynamically remodeled to adapt to all DNA-related processes, including DNA damage responses (DDR). This adaptation requires DNA and histone epigenetic modifications, which are mediated by several types of enzymes; among them are lysine methyltransferases (KMTs). METHODS: KMT inhibitors, chaetocin and tazemetostat (TZM), were used to study their role in the DDR induced by ionizing radiation or doxorubicin in two human sarcoma cells lines. The effect of these KMT inhibitors was tested by the analysis of chromatin epigenetic modifications, H4K16ac and H4K20me2. DDR was monitored by the formation of γH2AX, MDC1, NBS1 and 53BP1 foci, and the induction of apoptosis. RESULTS: Chaetocin and tazemetostat treatments caused a significant increase of H4K16 acetylation, associated with chromatin relaxation, and increased DNA damage, detected by the labeling of free DNA-ends. These inhibitors significantly reduced H4K20 dimethylation levels in response to DNA damage and impaired the recruitment of 53BP1, but not of MDC1 and NBS1, at DNA damaged sites. This modification of epigenetic marks prevents DNA repair by the NHEJ pathway and leads to cell death. CONCLUSION: KMT inhibitors could function as sensitizers to DNA damage-based therapies and be used in novel synthetic lethality strategies for sarcoma treatment.

Effects of Somatic Methylation in Colonic Polyps on Risk of Developing Metachronous Advanced Colorectal Lesions.

The utility of molecular markers for predicting the risk of metachronous advanced colorectal lesions (MACLs) remains poorly investigated. We examined the relationship between somatic hypermethylation in polyps at baseline and the risk of developing MACL. This retrospective cohort study included 281 consecutive patients with colonic polyps who were enrolled between 2007 and 2009 and followed-up until 2014. MACLs were defined as adenomas of ≥10 mm, high-grade dysplasia, or with a villous component; and serrated lesions of ≥10 mm or with dysplasia. In total, 595 polyps were removed at baseline colonoscopy and analyzed for pathological characteristics and CpG island methylator phenotype (CIMP) using the MS-MLPA (Methylation-Specific -- Multiplex Ligation-dependent Probe Amplification) technique. Forty-five patients (16.0%) showed at least one CIMP+ polyp. MACL risk was higher in patients with CIMP+ polyps (odds ratio (OR), 4.50; 95% CI, 1.78-11.4; p = 0.002). Patients with CIMP+ polyps also exhibited shorter time to MACL development (33.8 months vs. 50.1 months; p < 0.001), even with adjustment for polyp size and number (OR, 2.40; 95% CI, 1.33-4.34). Adding CIMP analysis improved the sensitivity (57.0% to 70.9%), negative predictive value (71.1% to 77.3%), and overall accuracy (49.8% to 52.0%) for MACL risk estimation. These results highlight that CIMP may be a useful marker for endoscopic surveillance.

A case report of a drug-induced liver injury (DILI) caused by multiple antidepressants with causality established by the updated Roussel Uclaf causality assessment method (RUCAM) and in vitro testing.

A 56-year-old female patient was hospitalized because of a lack of response and poor tolerance to multiple antidepressants, which included an episode of DILI. During hospitalization, the patient suffered another episode of DILI. Causality was assessed both by RUCAM and Lymphocyte Transformation Test, allowing to identify a safer medication.

Reinicio de la actividad ecográfica tras la fase aguda de la pandemia por COVID-19 / No disponible

No disponible

Resuming ultrasound activity after the acute phase of the COVID-19 pandemic.

Recently, Dr. Crespo et al. published in your Journal a paper recommending the use of ultrasonography during the current phase of the COVID-19 pandemic whilst wearing only a facial mask for protection, which we deem inadequate. Prevention is key when performing an ultrasound test, since this virus is highly contagious. During the pandemic, every patient should be considered as potentially infected and the procedure requires a close proximity. Therefore, extreme hygiene and a sonographer equipped with the appropriate personal protection (mask, cap, gown, gloves, shoe covers and goggles, with a facial screen and high-efficacy mask for confirmed or highly suspect cases) are of the utmost importance to prevent viral transmission.