Biology subjects

Shtil, A. A.

Publications and source records attributed to Shtil, A. A..

4 recordsLinked to original sources

bioRxiv2025

NONO as a Sensor of Intracellular Oxidation: Relevance to Neuroblastoma Cell Death

Neuroblastoma, a transcriptionally driven pediatric malignancy, exhibits a remarkable clinical and biological heterogeneity. Two major subtypes, the adrenergic and mesenchymal, are differentially governed by a subset of transcription factors that comprise the core regulatory circuit (CRC). The former subtype is often associated with MYCN amplification and is particularly aggressive and therapy-resistant, underscoring the need for novel targets. Here, we identify the multifunctional non-POU domain-containing octamer-binding (NONO) protein as a guardian of individual CRC genes, thereby contributing to survival of neuroblastoma cells with different MYCN copy numbers. Intracellular oxidation in response to auranofin, an inhibitor of thioredoxin reductase 1, rapidly down-regulated the amounts of NONO mRNA and protein in MYCN-amplified Kelly cell line. Conversely, NONO knockdown with RNA interference (siNONO) also triggered intracellular oxidation. These effects were less pronounced in the SK-N-AS cell line carrying a single MYCN copy, as well as in non-malignant HS5 fibroblasts. In Kelly cells, siNONO attenuated auranofin-induced activation of CRC genes HAND2 and PHOX2B. In line with preferential effects on NONO abundance, the Kelly cells were more sensitive than single MYCN copy counterparts to combinations of a sublethal concentration of auranofin with siNONO. Importantly, MYCN-amplified cells demonstrated a significantly suppressed clonogenic survival 14 days after transient exposure to these combinations compared with each agent alone; HS5 fibroblasts were largely spared. Our findings 1) establish NONO as a redox sensor, a non-trivial role for transcriptional proteins, and 2) justify the strategy of therapeutic targeting of MYCN-amplified tumors vulnerable to oxidative stress. Key pointsO_LINONO, a master regulator of the core regulatory circuit (CRC) in MYCN-amplified neuroblastoma, is rapidly down-regulated by auranofin-induced intracellular oxidation. C_LIO_LINONO knockdown synergizes with auranofin in triggering individual CRC gene deregulation and lethal oxidative stress preferentially in MYCN-amplified cells. C_LI

cancer biology↗

bioRxiv2024

Kolmogorov-Smirnov and Shapiro-Wilk single distribution analysis methods in examining sample structure, strength of genetic background, and effects of interventions in lifespan and aging studies

The analysis of lifespan and ageing involves a variety of calculations based on survival data. One of these is the comparison of survival curves. However, inspection of survival curves provides little information about the frequency of phenotypes within samples. Here we propose to divide the distribution into intervals to obtain information about the sample structure. Then, we apply the normality criterion to the distributions using the Kolmogorov-Smirnov and Shapiro-Wilk tests. Using this approach, we reveal the destabilising effect of mutations on ontogenesis and lifespan, and estimate the strength of the genetic background of the lines. The proposed methodology allows effective estimation of sample structure and adds a new layer of information to longevity studies.

genetics↗

bioRxiv2023

Knockout of cyclin dependent kinases 8 and 19 leads to depletion of cyclin C and suppresses spermatogenesis and male fertility in mice

Paralogs CDK8 and CDK19 are regulatory kinases associated with the transcriptional Mediator complex. We have e generated mice with the systemic inducible Cdk8 knockout on the background of Cdk19 constitutive knockout. Cdk8/19 double knockout (iDKO) males, but not single Cdk8 and Cdk19 KO, had an atrophic reproductive system and were infertile. The iDKO males lacked postmeiotic spermatids and spermatocytes after meiosis I pachytene. Testosterone levels were decreased whereas the amounts of the luteinizing hormone were unchanged. Single cell RNA sequencing showed marked differences in the expression of steroidogenic genes (such as Cyp17a1, Star and Fads) in Leydig cells concomitant with alterations in Sertoli cells and spermatocytes likely associated with impaired synthesis of steroids. Star and Fads were also downregulated in cultivated Leydig cells after iDKO. The treatment of primary Leydig cells culture with a CDK8/19 inhibitor did not induce the same changes in gene expression as iDKO, and prolonged treatment of mice with a CDK8/19 inhibitor did not affect the size of testes. iDKO, in contrast to single knockouts or treatment with a CDK8/19 kinase inhibitor, led to depletion of cyclin C (CcnC), the binding partner of CDK8/19 that has been implicated in CDK8/19-independent functions. This suggests that the observed phenotype was likely mediated through kinase-independent activities of CDK8/19, such as CcnC stabilization.

developmental biology↗

bioRxiv2023

CDK8/19 Inhibition Attenuates G1 Arrest Induced by BCR-ABL Antagonists and Accelerates Death of Chronic Myelogenous Leukemia Cells

Imatinib mesylate (IM) and other BCR-ABL tyrosine kinase inhibitors (BCR-ABLi) are the mainstay of chronic myelogenous leukemia (CML) treatment. However, activation of circumventing signaling pathways and quiescence may limit BCR-ABLi efficacy. CDK8/19 Mediator kinases have been implicated in the emergence of non-genetic drug resistance. Dissecting the effects of pharmacological CDK8/19 inhibition on CML survival in response to BCR-ABLi, we found that a selective, non-toxic CDK8/19 inhibitor (CDK8/19i) Senexin B (SenB) and other CDK8/19i sensitized K562 cells to different BCR-ABLi via attenuation of cell cycle arrest. In particular, SenB prevented IM-induced upregulation of genes that negatively regulate cell cycle progression. SenB also antagonized IM-activated p27Kip1 elevation thereby diminishing the population of G1-arrested cells. After transient G1 arrest, cells treated with IM+SenB re-entered the S phase, where they were halted and underwent replicative stress. Consequently, the combination of IM and SenB intensified apoptotic cell death, measured by activation of caspase 9 and 3, subsequent cleavage of poly(ADPriboso)polymerase 1, positive Annexin V staining and increase of subG1 fraction. In contrast, IM-treated BCR-ABL-positive KU812 CML cells, which did not induce p27Kip1, readily died regardless of SenB treatment. Thus, CDK8/19i prevent the quiescence-mediated escape from BCR-ABLi-induced apoptosis, suggesting a strategy for avoiding the CML relapse.

cancer biology↗