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The identification of genes that confer either extension of life span or accelerate age-related decline was a step forward in understanding the mechanisms of aging and revealed that it is partially controlled by genetics and transcriptional programs. Here, we discovered that the human DNA sequence C16ORF70 encodes a protein, named MYTHO (macroautophagy and youth optimizer), which controls life span and health span. MYTHO protein is conserved from Caenorhabditis elegans to humans and its mRNA was upregulated in aged mice and elderly people. Deletion of the orthologous myt-1 gene in C. elegans dramatically shortened life span and decreased animal survival upon exposure to oxidative stress. Mechanistically, MYTHO is required for autophagy likely because it acts as a scaffold that binds WIPI2 and BCAS3 to recruit and assemble the conjugation system at the phagophore, the nascent autophagosome. We conclude that MYTHO is a transcriptionally regulated initiator of autophagy that is central in promoting stress resistance and healthy aging., Introduction In nature, organisms are continuously exposed to environmental stresses that challenge their survival. The species that quickly and efficiently adapt to hostile conditions are positively selected. This response is [...]

Molecular profiling of clear cell renal cell carcinoma (ccRCC) tumors of patients in a clinical trial has identified distinct transcriptomic signatures with predictive value, yet data in non-clear cell variants (nccRCC) are lacking. We examined the transcriptional profiles of RCC tumors representing key molecular pathways, from a multi-institutional, real- world patient cohort, including ccRCC and centrally reviewed nccRCC samples. ccRCC had increased angiogenesis signature scores compared with the heterogeneous group of nccRCC tumors, while cell cycle, fatty acid oxidation/AMPK signaling, and fatty acid synthesis/pentose phosphate signature scores were increased in one or more nccRCC subtypes. Among both ccRCC and nccRCC tumors, T effector scores statistically correlated with increased immune cell infiltration and were more commonly associated with immunotherapy-related markers ([PD- L1.sup.+]/[TMB.sup.hi]/[MSI.sup.hi]). In conclusion, this study provides evidence of differential gene transcriptional profiles among ccRCC versus nccRCC tumors, providing insights for optimizing personalized and histology-specific therapeutic strategies for patients with advanced RCC., Introduction Renal cell carcinoma (RCC) is a common cancer among men and women in the United States, with an estimated 81,800 new cases and 14,890 deaths expected in 2023 (1). [...]

One critical mechanism through which prostate cancer (PCa) adapts to treatments targeting androgen receptor (AR) signaling is the emergence of ligand-binding domain-truncated and constitutively active AR splice variants, particularly AR-V7. While AR-V7 has been intensively studied, its ability to activate distinct biological functions compared with the full- length AR (AR-FL), and its role in regulating the metastatic progression of castration- resistant PCa (CRPC), remain unclear. Our study found that, under castrated conditions, AR-V7 strongly induced osteoblastic bone lesions, a response not observed with ARFL overexpression. Through combined ChIP-seq, ATAC-seq, and RNA-seq analyses, we demonstrated that AR-V7 uniquely accesses the androgen-responsive elements in compact chromatin regions, activating a distinct transcription program. This program was highly enriched for genes involved in epithelial-mesenchymal transition and metastasis. Notably, we discovered that SOX9, a critical metastasis driver gene, was a direct target and downstream effector of AR-V7. Its protein expression was dramatically upregulated in AR-V7-induced bone lesions. Moreover, we found that Ser81 phosphorylation enhanced AR-V7's pro-metastasis function by selectively altering its specific transcription program. Blocking this phosphorylation with CDK9 inhibitors impaired the AR-V7-mediated metastasis program. Overall, our study has provided molecular insights into the role of AR splice variants in driving the metastatic progression of CRPC., Introduction The androgen receptor (AR) is critical in driving prostate cancer (PCa) development, with androgen deprivation therapy (ADT) being the standard treatment for PCa patients. Although initial responses are generally [...]

Nuclear receptor 4A1 (NR4A1), a member of the NR4A subfamily, acts as a gene regulator in a wide range of signaling pathways and responses to human diseases. Here, we provide a brief overview of the current functions of NR4A1 in human diseases and the factors involved in its function. A deeper understanding of these mechanisms can potentially improve drug development and disease therapy. Key words: NR4A1, function, human disease, molecular pathway, 1. Introduction The nuclear receptor 4A (NR4A) subfamily, which belongs to the nuclear receptor (NR) superfamily, contains three members: NR4A1 (mouse homologue Nur77, rat homologue NGFI-B, and human homologue TR3 [...]

Aberrant activation of Wnt/[beta]-catenin induces renal dysfunction by initiating pro-apoptotic cascades, fibrosis, oxidative and inflammatory burden. This study tested the therapeutic effects of Wnt/[beta]-catenin inhibitor pyrvinium against cisplatin-induced acute kidney injury (AKI) in rats. Cisplatin was administered at a single dose of 5 mg/kg (i.p.) and renal cisplatin accumulation and uptake in cortical slices were determined after the fifth day by atomic absorption spectroscopy. Levels of pro- inflammatory cytokines were checked by ELISA, and organic cation transporter-2 (OCT-2) transcription and expression in renal tissue were evaluated by RT-PCR and immunohistochemical technique. Cisplatin administration produced renal dysfunction manifested as increase in serum creatinine, blood urea nitrogen, proteinuria, reduced clearance and electrolyte imbalance. Oxidative stress indices, pro-inflammatory cytokines, fibronectin, and caspase-3 activity were elevated in cisplatin-challenged rats. Moreover, increased renal OCT-2 transcription and immunostaining were detected in cisplatin kidneys which resulted in platinum accumulation. Additional docking studies depicted strong interaction between the [beta]-catenin and OCT-2 protein. These manifestations induced mitochondrial dysfunction, histological damage and fibrosis. Notably, Wnt/[beta]-catenin inhibitor pyrvinium (60 [micro]g/kg; p.o.) treatment reduced the renal OCT-2 gene transcription causing a decline in platinum levels. Thus, the present study concludes that Wnt/[beta]-catenin inhibition attenuates cisplatin-induced AKI in rats, partly by down-regulating OCT-2 expression. Key words: cisplatin, acute kidney injury, fibrosis, Wnt/ [beta]-catenin pathway, oxidative stress, inflammation L'activation aberrante de la voie de signalisation Wnt/catenine [beta] entraine une insuffisance renale par l'amorce de cascades proapoptotiques, de fibrose et d'une charge oxydative et inflammatoire. Cette etude a porte sur les effets therapeutiques du pyrvinium, un inhibiteur de la Wnt/ catenine ft, contre l'insuffisance renale aigue (IRA) entrainee par le cisplatine chez le rat. Nous avons administre du cisplatine a une dose unique de 5 mg/kg par voie i.p. et, a l'aide de la spectroscopie d'absorption atomique, nous avons etabli l'accumulation et l'assimilation renales du cisplatine dans des coupes de cortex apres le 5e jour. A l'aide de la technique d'ELISA, nous avons verifie les taux de cytokines proinflammatoires, de meme que la transcription et l'expression du transporteur de cations organiques 2 (OCT-2) dans le tissu renal a l'aide des techniques de PCR en temps reel et d'immunohistochimie. L'administration de cisplatine entrainait une insuffisance renale qui se manifestait par une hausse de la creatinine et de l'azote ureique dans le serum avec une proteinurie, une diminution de la clairance et un desequilibre electrolytique. Nous avons observe une hausse des indices du stress oxydatif de meme que de l'activite des cytokines proinflammatoires, de la fibronectine et de la caspase 3 avec la provocation par le cisplatine chez le rat. En outre, nous avons observe une augmentation de la transcription d'OCT-2 et de l'immunocoloration dans les reins soumis au cisplatine, ce qui a mene a une accumulation de platine. Des etudes d'amarrages supplementaires ont montre la presence d'une forte interaction entre la catenine p et l'OCT-2 proteique. Ces manifestations entrainaient un dysfonctionnement mitochondrial, des dommages histologiques et de la fibrose. Il est interessant de noter que l'administration de pyrvinium a 60 [micro]g/kg par voie orale entrainait un abaissement de la transcription renale d'OCT-2 menant a une diminution des taux de platine. Par consequent, la presente etude permet de conclure que chez le rat, l'inhibition de la voie de signalisation Wnt/catenine [beta] permet d'attenuer l'IRA provoquee par le cisplatine, en partie par la regulation a la baisse de l'expression d'OCT-2. [Traduit par la Redaction] Mots-cles : cisplatine, insuffisance renale aigue, fibrose, voie de signalisation Wnt/ catenine [beta], stress oxydatif, inflammation, 1. Introduction Cisplatin (cis-diamminedichloroplatinum II) is an anticancer agent and has been extensively utilized in the treatment of metastatic testicular, lung, neck, and ovarian carcinoma (Pabla and Dong 2008). Dose-dependent [...]

* Context.--Diagnostic testing for SARS-CoV-2 in symptomatic and asymptomatic children remains integral to care, particularly for supporting return to and attendance in schools. The concordance of SARS-CoV-2 detection in children, using various specimen types, has not been widely studied. Objective.--To compare 3 sample types for SARS-CoV-2 polymerase chain reaction (PCR) testing in children, collected and tested at a single facility. Design.--We prospectively recruited 142 symptomatic and asymptomatic children/young adults into a sample comparison study performed in a single health care system. Each child provided self-collected saliva, and a trained health care provider collected a mid-turbinate nasal swab and nasopharyngeal (NP) swab. Specimens were assayed within 24 hours of collection by using reverse transcription-polymerase chain reaction (RT-PCR) to detect SARS-CoV-2 on a single testing platform. Results.--Concurrently collected saliva and mid-turbinate swabs had greater than 95% positive agreement with NP swabs when obtained within 10 days of symptom onset. Positive agreement of saliva and mid-turbinate samples collected from children with symptom onset >10 days prior, or without symptoms, was 82% compared to NP swab samples. Cycle threshold (Ct) values for mid-turbinate nasal samples more closely correlated with Ct values from NP samples than from saliva samples. Conclusions.--These findings suggest that all 3 sample types from children are useful for SARS-CoV-2 diagnostic testing by RT-PCR, and that concordance is greatest when the child has had symptoms of COVID-19 within the past 10 days. This study provides scientific justification for using sample types other than the NP swab for SARS-CoV-2 testing in pediatric populations. (Arch Pathol Lab Med. 2022;146:1056-1061; doi: 10.5858/arpa.2021-0625-SA), A pandemic provides a unique setting upon which the basic tenets of laboratory medicine are challenged. Specimen collection for SARS-CoV-2 is one such example, for which validation studies, which are [...]

Inorganic pyrophosphatase (iPPase) is an enzyme that cleaves pyrophosphate into two phosphate molecules. This enzyme is an essential component of in vitro transcription (IVT) reactions for RNA preparation as it prevents pyrophosphate from precipitating with magnesium, ultimately increasing the rate of the IVT reaction. Large-scale RNA production is often required for biochemical and biophysical characterization studies of RNA, therefore requiring large amounts of IVT reagents. Commercially purchased iPPase is often the most expensive component of any IVT reaction. In this paper, we demonstrate that iPPase can be produced in large quantities and high quality using a reasonably generic laboratory facility and that laboratory-purified iPPase is as effective as commercially available iPPase. Furthermore, using size exclusion chromatography coupled with multi-angle light scattering and dynamic light scattering, analytical ultracentrifugation, and small-angle X-ray scattering, we demonstrate that yeast iPPase can form tetramers and hexamers in solution as well as the enzymatically active dimer. Our work provides a robust protocol for laboratories involved with RNA in vitro transcription to efficiently produce active iPPase, significantly reducing the financial strain of large-scale RNA production. Key words: inorganic pyrophosphatase, RNA in vitro transcription, iPPase, analytical ultracentrifuge, small-angle X-ray scattering La pyrophosphatase inorganique (iPPase) est une enzyme qui clive le pyrophosphate en deux molécules de phosphate. Cette enzyme est un composant essentiel des réactions de transcription in vitro (TIV) pour la préparation d'ARN, car elle empêche le pyrophosphate de précipiter avec le magnésium, ce qui augmente finalement la vitesse de la réaction de TIV. La production d'ARN à grande échelle est souvent requise pour les études de caractérisation biochimique et biophysique de i'ARN, ce qui nécessite de grandes quantités de réactifs pour la TIV. L'iPPase achetée dans le commerce est souvent le composant le plus cher de toute réaction de TIV. Dans cet article, les auteurs démontrent que l'iPPase peut être produite en grandes quantités avec une haute qualité en utilisant une installation de laboratoire raisonnablement générique, et que l'iPPase purifiée en laboratoire est aussi efficace que l'iPPase disponible dans le commerce. De plus, en utilisant la chromatographie d'exclusion stérique couplée à la diffusion multiangle de la lumière et à la diffusion dynamique de la lumière, l'ultracentrifugation analytique et la diffusion de rayons X aux petits angles, ils démontrent que l'iPPase de levure peut former des tétramères et des hexamères en solution, ainsi que le dimère enzymatiquement actif. Ce travail offre un protocole robuste pour les laboratoires engagés dans la transcription in vitro d'ARN afin de produire efficacement l'iPPase active, réduisant ainsi de manière significative les contraintes financières de la production d'ARN à grande échelle. [Traduit par la Rédaction] Mots-clés : pyrophosphatase inorganique, transcription in vitro de l'ARN, iPPase, ultracentrifugeuse analytique, diffusion des rayons X aux petits angles, Introduction Inorganic pyrophosphatase (iPPase; EC 3.6.1.1) is primarily characterized by its function to catalyze the hydrolysis of pyrophosphate into two monophosphates (Harold 1966), as presented in eq. 1. Three different [...]

MYB transcription factor, which contains a conserved DNA binding domain, has been found in almost all eukaryotes. MYB genes have variable functions in plants and are involved in many pathways. We systemically analyzed the MYB gene family in three Ericaceae species, Rhododendron williamsianum Rehder & E.H. Wilson, Rhododendron delavayi Franch., and Vaccinium corymbosum L., and one outgroup, Actinidia chinensis Planch., with 99, 156, 480, and 185 MYB genes found, respectively. The MYB genes were classified into five types based on the number of conserved MYB motifs, and the two repeat (2R) types were dominant in all four species. The percentage of 2R type MYB ranged from 48.5% to 87.9% depending on the species. We further classified the conserved MYB motifs into M1, M2, and M3 types based on motif definition. We found an abundance of 3xM2 type in the 3R group, but found a species-specific type preference for 1R and 2R genes. In searching for Arabidopsis drought-resistant genes, we detected 34 potential candidates in four species. The expression profile of R. delavayi showed 11 candidate drought-resistant RdMYB genes, which provide a potential molecular design target for breeders. Our results describe the MYB gene family in these four species and could play an important role in future analyses. Key words: Ericaceae, Rhododendron, blueberry, kiwi fruit, conserved motif. Le facteur de transcription MYB, qui comporte un domaine de liaison a l'ADN conserve, a ete trouve chez presque tous les eucaryotes. Les genes MYB exercent des fonctions variables chez les plantes et sont impliques dans de nombreuses voies. Les auteurs ont analyse de maniere systematique la famille des genes MYB chez trois especes d'Ericaceae, Rhododendron williamsianum Rehder & E.H. Wilson, Rhododendron delavayi Franch. et Vaccinium corymbosum L., et chez un hors-groupe, Actinidia chinensis Planch., avec 99, 156, 480 et 185 genes MYB trouves, respectivement. Les genes MYB ont ete classes en cinq types en fonction du nombre de motifs MYB conserves, et les types a deux repetitions (2R) etaient dominants chez les quatre especes. Le pourcentage de MYB de type 2R variait de 48,5 % a 87,9 % chez les differentes especes. Ils ont ensuite classe les motifs MYB conserves en types M1, M2 et M3 en fonction de la definition du motif. Ils ont constate une abondance du type 3xM2 chez le groupe 3R, mais une preference specifique a l'espece quant aux types pour les genes 1R et 2R. En recherchant les genes de resistance a la secheresse chez Arabidopsis, ils ont detecte 34 candidats potentiels chez les quatre especes. Le profil d'expression de R. delavayi a montre la presence de 11 genes RdMYB candidats pour la resistance a la secheresse, qui constituent une cible moleculaire potentielle pour les selectionneurs. Ces resultats decrivent la famille de genes MYB chez ces quatre especes qui pourraient jouer un role important dans les analyses futures. [Traduit par la Redaction] Mots-cles : Ericaceae, Rhododendron, myrtille, kiwi, motif conserve., Introduction Transcription factors (TFs) play a central role in developmental and metabolic programsby regulating the transcription of downstream target genes (Cao et al. 2020). v-Myb avian myeloblastosis viral oncogene homolog [...]

A method for the quantitative determination of mRNA of TEM-type serine beta-lactamases in bacteria resistant to beta-lactam antibiotics has been developed. The method includes several stages: the isolation of a total RNA fraction from a bacterial culture and the production of the beta-lactamase target DNA in sequential reverse transcription and polymerase chain reaction with the incorporation of a biotin label, and the hybridization of the target DNA on colorimetric biochips. Sample preparation conditions were optimized to increase the yield of the analyzed target DNA. A calibration curve for determining the amount of mRNA was constructed using a standard sample of beta-lactamase TEM-1 mRNA obtained by transcription in vitro. The advantage of using a standard sample corresponding to the full-length bla.sub.TEM-1 gene is that it passes through all the stages of analysis in parallel with the test samples with the same efficiency. The detection limit for TEM-1 beta-lactamase mRNA was 0.40 ± 0.05 amol/mL; the range of determined concentrations was from 1.0 amol/mL to 2000 fmol/mL, and the relative standard deviation did not exceed 12%. The duration of analysis after the production of a bacterial culture was about 7 h. The developed method can be used to study the conditions for the expression of beta-lactamase genes in bacteria resistant to antimicrobial drugs., Author(s): M. Yu. Rubtsova [sup.1], A. A. Filippova [sup.1], N. K. Fursova [sup.2], V. G. Grigorenko [sup.1], G. V. Presnova [sup.1], M. M. Ulyashova [sup.1], A. M. Egorov [sup.1] Author [...]

Wood frogs are a few vertebrate species that can survive whole-body freezing. Multiple adaptations support this, including cryoprotectant production (glucose), metabolic rate depression, and selective changes in gene and protein expression to activate pro-survival pathways. The role of DNA methylation machinery (DNA methyltransferases, DNMTs) in regulating nuclear gene expression to support freezing survival has already been established. However, a comparable role for DNMTs in the mitochondria has not been explored in wood frogs. We examined the mitochondrial protein levels of DNMT-1, DNMT-3A, DNMT-3B, and DNMT-3L as well as mitochondrial DNMT activity in the liver and heart to assess the involvement of DNMT in the survival of freezing and dehydration stresses (cellular dehydration being a component of freezing). Our results showed stress- and tissue-specific responses to mitochondrial DNMT-1 in the liver and heart, respectively. During 24 h of freezing and whole-body dehydration, we observed an overall downregulation of mitochondrial DNMT-1, a major protein involved in maintaining methylation levels related to its role in the selective transcription of mitochondrial genes as well as antioxidant response. Tissue-specific responses of protein levels of DNMT-3A, DNMT-3B, DNMT-3L, and DNMT activity in the liver suggested a preference for a higher methylation state in the liver under both freezing and dehydration stress, but not in the heart. Key words: DNMT activity, metabolic rate depression, antioxidant response, mitochondria, mitochondrial proteins. Les grenouilles des bois sont l'une des rares especes de vertebres qui peuvent survivre a la congelation du corps entier. De multiples adaptations permettent cette survie, notamment la production de cryoprotecteurs (glucose), la reduction de la vitesse du metabolisme et des changements selectifs dans l'expression de genes et de proteines pour activer les voies favorisant la survie. Le role de la machinerie de methylation de l'ADN (ADN methyltransferases, DNMT) dans la regulation de l'expression des genes nucleaires favorisant la survie lors de la congelation a deja ete etabli. Cependant, un role comparable des DNMT dans les mitochondries n'a pas ete explore chez les grenouilles des bois. Les auteurs ont examine les niveaux des proteines DNMT-1, DNMT-3A, DNMT-3B et DNMT-3L dans les mitochondries ainsi que l'activite DNMT mitochondriale dans le foie et le coeur afin d'evaluer l'implication des DNMT dans la survie aux deux types de stress provoques par le gel et la deshydratation (la deshydratation cellulaire etant une composante du gel). Leurs resultats ont montre une reponse par la proteine DNMT-1 mitochondriale specifique au stress et au tissu dans le foie et le coeur respectivement. Au cours d'une congelation de 24 heures et d'une deshydratation du corps entier, ils ont observe une regulation globale a la baisse de la DNMT-1 mitochondriale, une proteine majeure impliquee dans le maintien des niveaux de methylation, liee a son role dans la transcription selective des genes mitochondriaux ainsi que dans la reponse antioxydante. La reponse specifique au tissu des niveaux des proteines DNMT-3A, DNMT-3B et DNMT-3L et l'activite DNMT dans le foie ont suggere une preference pour un etat de methylation plus eleve dans le foie sous les deux types de stress de congelation et de deshydratation, mais pas dans le coeur. [Traduit par la Redaction] Mots-cles : Activite DNMT, reduction de la vitesse du metabolisme, reponse antioxydante, mitochondries, proteines mitochondriales., Introduction Wood frogs, Rana sylvatica, are one of the best-studied models of natural freeze-tolerance; they can survive whole-body freezing during winter, with up to 65%--70% of their total body water [...]

This study aimed to reveal the gene transcriptional alterations, possible molecular mechanisms, and pathways involved in the synergy of 5-aza-2'-deoxycytidine (DAC) and Cisplatin (CDDP) in urothelial carcinoma (UC). Two UC cell lines, 5637 and T24, were used in the present study. A cDNA microarray was performed to identify critical genes involved in the synergistic mechanism of both agents against UC cells. The results showed that several key regulatory genes, such as interleukin 24 (IL24), fibroblast growth factor 1 (FGF1), and transforming growth factor beta induced (TGFBI), may play critical roles in the synergy of DAC and CDDP in UC. Pathway enrichment suggested that many carcinogenesis-related pathways, such as the ECM-receptor interaction and MAPK signaling pathways, may participate in the synergy of both agents. Our results suggest that TGF-[beta]1 stimulates the phosphorylation of ERK1/2 and p38 by increasing TGFBI expression, and that the TGFBI-MAPK signaling pathway plays an important role in the synergy of DAC and CDDP against UC. Therefore, we revealed the synergistic mechanism of DAC and CDDP in UC. Several key regulatory genes play critical roles in the synergy of combined treatment, and the TGFBI-MAPK signaling pathway may be an important potential target of these two agents. Key words: urothelial carcinoma, DAC, CDDP, TGFBI, MAPK. Cette etude vise a reveler les modifications transcriptionnelles de genes, le possible mecanisme moleculaire et les voies de signalisation qui sont impliques dans la synergie de la 5-aza-2'-desoxycytidine (DAC) et du cisplatine (CDDP) dans le cancer urothelial (CU). Deux lignees cellulaires de CU, les 5637 et T24, ont ete utilisees dans cette etude. Une biopuce d'ADNc a ete construite afin d'identifier les genes critiques impliques dans le mecanisme synergique des deux agents contre les cellules de CU. Les resultats ont montre que plusieurs genes regulateurs cles, tels que les genes de l'interleukine 24 (IL24), du facteur de croissance fibroblastique 1 (FGF1) et du facteur de croissance transformant beta induit (TGFBI), ont ete identifies comme pouvant jouer un role cle dans la synergie de la DAC et du CDDP dans le CU. L'analyse d'enrichissement en matiere de voies de signalisation suggerait que de nombreuses voies liees a la carcinogenese, comme l'interaction MEC-recepteur et les voies de signalisation MAPK, peuvent participer a la synergie des deux agents. Ces resultats suggerent que le TGF-[beta]1 stimule les niveaux de phosphorylation de ERK1/2 et p38 par l'augmentation de l'expression du TGFBI et que la voie de signalisation TGFBI-MAPK joue un role important dans la synergie de la DAC et du CDDP contre le CU. Ainsi, les auteurs ont revele le mecanisme synergique de la DAC et du CDDP dans le CU, determine que plusieurs genes regulateurs cles jouent des roles critiques dans la synergie du traitement combine, et que la voie de signalisation TGFBI-MAPK pourrait etre une cible potentielle importante de ces deux agents. [Traduit par la Redaction] Mots-cles : carcinome urothelial, DAC, CDDP, TGFBI, MAPK., Introduction Urothelial carcinoma (UC) is a common malignancy of the urinary tract and the ninth most common cancer (Ferlay et al. 2015). Approximately 70%--80% of UC cases are non-muscle-invasive, of [...]