RESUMEN
Dynamic interaction between BRCA2 and telomeric G-quadruplexes (G4) is crucial for maintaining telomere replication homeostasis. Cells lacking BRCA2 display telomeric damage with a subset of these cells bypassing senescence to initiate break-induced replication (BIR) for telomere synthesis. Here we show that the abnormal stabilization of telomeric G4 following BRCA2 depletion leads to telomeric repeat-containing RNA (TERRA)-R-loop accumulation, triggering liquid-liquid phase separation (LLPS) and the assembly of Alternative Lengthening of Telomeres (ALT)-associated promyelocytic leukemia (PML) bodies (APBs). Disruption of R-loops abolishes LLPS and impairs telomere synthesis. Artificial engineering of telomeric LLPS restores telomere synthesis, underscoring the critical role of LLPS in ALT. TERRA-R-loops also recruit Polycomb Repressive Complex 2 (PRC2), leading to tri-methylation of Lys27 on histone H3 (H3K27me3) at telomeres. Half of paraffin-embedded tissue sections from human breast cancers exhibit APBs and telomere length heterogeneity, suggesting that BRCA2 mutations can predispose individuals to ALT-type tumorigenesis. Overall, BRCA2 abrogation disrupts the dynamicity of telomeric G4, producing TERRA-R-loops, finally leading to the assembly of telomeric liquid condensates crucial for ALT. We propose that modulating the dynamicity of telomeric G4 and targeting TERRA-R-loops in telomeric LLPS maintenance may represent effective therapeutic strategies for treating ALT-like cancers with APBs, including those with BRCA2 disruptions.
Asunto(s)
Proteína BRCA2 , Replicación del ADN , G-Cuádruplex , Homeostasis del Telómero , Telómero , Humanos , Telómero/metabolismo , Telómero/genética , Proteína BRCA2/genética , Proteína BRCA2/metabolismo , Homeostasis del Telómero/genética , Replicación del ADN/genética , Histonas/metabolismo , Histonas/genética , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Estructuras R-Loop , Complejo Represivo Polycomb 2/metabolismo , Complejo Represivo Polycomb 2/genética , Línea Celular Tumoral , Femenino , Separación de FasesRESUMEN
Liquid-liquid phase separation (LLPS) facilitates the formation of membraneless organelles within cells, with implications in various biological processes and disease states. AT-rich interactive domain-containing protein 1A (ARID1A) is a chromatin remodeling factor frequently associated with cancer mutations, yet its functional mechanism remains largely unknown. Here, we find that ARID1A harbors a prion-like domain (PrLD), which facilitates the formation of liquid condensates through PrLD-mediated LLPS. The nuclear condensates formed by ARID1A LLPS are significantly elevated in Ewing's sarcoma patient specimen. Disruption of ARID1A LLPS results in diminished proliferative and invasive abilities in Ewing's sarcoma cells. Through genome-wide chromatin structure and transcription profiling, we identify that the ARID1A condensate localizes to EWS/FLI1 target enhancers and induces long-range chromatin architectural changes by forming functional chromatin remodeling hubs at oncogenic target genes. Collectively, our findings demonstrate that ARID1A promotes oncogenic potential through PrLD-mediated LLPS, offering a potential therapeutic approach for treating Ewing's sarcoma.
Asunto(s)
Ensamble y Desensamble de Cromatina , Proteínas de Unión al ADN , Proteína EWS de Unión a ARN , Sarcoma de Ewing , Factores de Transcripción , Humanos , Sarcoma de Ewing/genética , Sarcoma de Ewing/metabolismo , Sarcoma de Ewing/patología , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ADN/genética , Línea Celular Tumoral , Proteína EWS de Unión a ARN/metabolismo , Proteína EWS de Unión a ARN/genética , Regulación Neoplásica de la Expresión Génica , Proliferación Celular , Proteínas de Fusión Oncogénica/metabolismo , Proteínas de Fusión Oncogénica/genética , Proteína Proto-Oncogénica c-fli-1/metabolismo , Proteína Proto-Oncogénica c-fli-1/genética , Cromatina/metabolismo , Carcinogénesis/genética , Animales , Ratones , Dominios Proteicos , Neoplasias Óseas/genética , Neoplasias Óseas/metabolismo , Neoplasias Óseas/patología , Separación de FasesRESUMEN
Phase separation is a key mechanism for intracellular organization, driving the segregation of biomolecules into distinct condensates. Intracellular condensates play diverse functional roles including gene expression, stress response, and cell signaling. Technologies that enable the control of intracellular phase separation can be highly useful not only for a better understanding of the biophysical principles of phase separation processes but also for engineering novel condensates. Here, we describe an optogenetic approach for spatiotemporal control of phase separation in living cells.
Asunto(s)
Optogenética , Biofisica , Citoplasma/metabolismoRESUMEN
Newly synthesized proteins in the endoplasmic reticulum (ER) are sorted by coat protein complex II (COPII) at the ER exit site en route to the Golgi. Under cellular stresses, COPII proteins become targets of regulation to control the transport. Here, we show that the COPII outer coat proteins Sec31 and Sec13 are selectively sequestered into the biomolecular condensate of SCOTIN/SHISA-5, which interferes with COPII vesicle formation and inhibits ER-to-Golgi transport. SCOTIN is an ER transmembrane protein with a cytosolic intrinsically disordered region (IDR), which is required and essential for the formation of condensates. Upon IFN-γ stimulation, which is a cellular condition that induces SCOTIN expression and condensation, ER-to-Golgi transport was inhibited in a SCOTIN-dependent manner. Furthermore, cancer-associated mutations of SCOTIN perturb its ability to form condensates and control transport. Together, we propose that SCOTIN impedes the ER-to-Golgi transport through its ability to form biomolecular condensates at the ER membrane.
Asunto(s)
Retículo Endoplásmico , Proteínas de Transporte Vesicular , Proteínas de Transporte Vesicular/metabolismo , Transporte Biológico , Transporte de Proteínas/fisiología , Retículo Endoplásmico/metabolismo , Aparato de Golgi/metabolismoRESUMEN
4-Nonylphenol (NP) is a surfactant that is a well-known and widespread estrogenic endocrine disrupting chemical (EDC). Although it has been known that the affinity of NP to ERs is low, it has been suggested that low-dose NP has toxicity. In the present study, the endocrine disrupting effects on reproduction, and the weight of gonads, epididymis, and uterus were evaluated with the chronic lower-dose NP exposing. This study was designed by following the OECD test guideline 443 and subjected to a complete necropsy. In male, NP had an effect on the weight of the testis and epididymis in both F0 and F1. In females, NP decreased the weight of ovary and uterus in F0 but not in pre-pubertal F1 pubs. Fertility of male and female in F0 or F1 was no related with NP administration. The number of caudal-epididymal sperm by body weight (BW) was not different between groups in both F0 and F1. Besides, the difference of the sperm number between generations was not detected. The number of ovulated oocytes was similar between groups in F0, but significantly decreased in NP 50 group of F1. The litter size and sex ratios of offspring in F1 and F2 were not different. The accumulated mating rate and gestation period were not affected by the NP administration. Those results shows that chronic lower-dose NP administration has an effect of endocrine disruptor on the weight of gonads and epididymis of F0 and F1 but not in reproduction. Based on the results, it is suggested that chronic lower-dose NP exposing causes endocrine disruption in the weight of gonad and epididymis but not in the reproductive ability of next generations.
RESUMEN
Diphlorethohydroxycarmalol (DPHC) is a known to modulate the expression of extracellular matrix (ECM) components in 3T3-L1. However, the possible role of DPHC in integument stability during obesity induction is not clear yet. We evaluated the effects of DPHC on collagen or elastic fiber quantity in integument during obesity induction with high-fat diet. The dorsal back integument sections were stained with hematoxylin-eosin, Masson trichrome, and Verhoff-Van Gieson. The intensities of collagen fibers and elastin fibers were analyzed with ImageJ. The number of fibroblasts was counted at ×1,000 fields. The number of fibroblast was increased by obesity induction, but DPHC suppressed it in a concentrationdependent manner both in lean and obese mice. On the other hand, the intensities of collagen fibers were increased by DPHC treatment in obese mice groups but not in lean mice groups. The intensities of collagen fibers of obese mice were lower than that of the lean mice in 0% group. However, the number became similar between lean and obese mice by the treatment of DPHC. The intensity of elastic fibers was increased in the lean mice with the concentration of DPHC. In the obese mice group, there were increasing patterns but only significant at 10% DPHC group. The intensity of elastic fibers of obese mice was higher than lean mice in 0%, 1%, and 10% groups. Histologically epithelial cells and follicle cells which were diffused nuclear staining forms were increased by DPHC treatment. The results suggest that the activity of integument cells during obesity induction can be modulated by DPHC.