Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 8 de 8
Filtrar
Mais filtros

Base de dados
Tipo de documento
Intervalo de ano de publicação
1.
Chromosoma ; 2023 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-37728741

RESUMO

Mitosis is an essential process in which the duplicated genome is segregated equally into two daughter cells. CTCF has been reported to be present in mitosis and has a role in localizing CENP-E, but its importance for mitotic fidelity remains to be determined. To evaluate the importance of CTCF in mitosis, we tracked mitotic behaviors in wild-type and two different CTCF CRISPR-based genetic knockdowns. We find that knockdown of CTCF results in prolonged mitoses and failed anaphase segregation via time-lapse imaging of SiR-DNA. CTCF knockdown did not alter cell cycling or the mitotic checkpoint, which was activated upon nocodazole treatment. Immunofluorescence imaging of the mitotic spindle in CTCF knockdowns revealed disorganization via tri/tetrapolar spindles and chromosomes behind the spindle pole. Imaging of interphase nuclei showed that nuclear size increased drastically, consistent with failure to divide the duplicated genome in anaphase. Long-term inhibition of CNEP-E via GSK923295 recapitulates CTCF knockdown abnormal mitotic spindles with polar chromosomes and increased nuclear sizes. Population measurements of nuclear shape in CTCF knockdowns do not display decreased circularity or increased nuclear blebbing relative to wild-type. However, failed mitoses do display abnormal nuclear morphologies relative to successful mitoses, suggesting that population images do not capture individual behaviors. Thus, CTCF is important for both proper metaphase organization and anaphase segregation which impacts the size and shape of the interphase nucleus likely through its known role in recruiting CENP-E.

2.
Chromosome Res ; 30(1): 123-136, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35239049

RESUMO

More than one third of the mammalian genome is in a close association with the nuclear lamina, thus these genomic regions were termed lamina-associated domains (LADs). This association is fundamental for many aspects of chromatin biology including transcription, replication, and DNA damage repair. LADs association with the nuclear envelope is thought to be dependent on two major mechanisms: The first mechanism is the interaction between nuclear membrane proteins such as LBR with heterochromatin modifications that are enriched in LADs chromatin. The second mechanism is based on proteins that bind the borders of the LADs and support the association of the LADs with the nuclear envelope. Two factors were suggested to support the second mechanism: CCCTC-binding factor (CTCF) and YY1 based on their enriched binding to LADs borders. However, this mechanism has not been proven yet at a whole genome level. Here, to test if CTCF supports the LADs landscape, we generated melanoma cells with a partial loss of function (pLoF) of CTCF by the CRISPR-Cas9 system and determined the LADs landscape by lamin B ChIP-seq analysis. We found that under regular growth conditions, CTCF pLoF led to modest changes in the LADs landscape that included an increase in the signal of 2% of the LADs and a decrease in the signal of 8% of the LADs. However, CTCF importance for the LADs landscape was much higher upon induction of a chromatin stress. We induced chromatin stress by inhibiting RNA polymerase II, an intervention that is known to alter chromatin compaction and supercoiling. Notably, only in CTCF pLoF cells, the chromatin stress led to the dissociation of 7% of the LADs from the lamina. The CTCF-dependent LADs had almost three times shorter median length than the non-affected LADs, were enriched in CTCF binding at their borders, and were higher in their facultative-status (cell-type specific). Thus, it appears that CTCF is a key factor in facilitating the association of short facultative LADs with the nuclear lamina upon chromatin stress.


Assuntos
Cromatina , Lâmina Nuclear , Animais , Cromatina/genética , Cromatina/metabolismo , Genoma , Genômica , Heterocromatina/metabolismo , Mamíferos/genética , Lâmina Nuclear/química , Lâmina Nuclear/genética , Lâmina Nuclear/metabolismo
3.
Environ Chem Lett ; 19(6): 3967-3972, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34456659

RESUMO

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has raised the need of versatile means for virus decontamination. Millimeter waves are used in biochemical research in dynamic nuclear polarization enhanced nuclear magnetic resonance (DNP/NMR) spectroscopy. However, their efficiency in object decontamination for viruses has not been tested yet. Here we report the high efficiency of 95 GHz waves in killing both coronavirus 229E and poliovirus. An exposure of 2 s to 95 GHz waves reduced the titer of these viruses by 99.98% and 99.375%, respectively, and formed holes in the envelope of 229E virions as detected by scanning electron microscopy (SEM) analysis. The ability of 95 GHz waves to reduce the coronavirus titer to a range of limited infective dose of SARS-CoV-2 for humans and animal models along with precise focusing capabilities for these waves suggest 95 GHz waves as an effective way to decontaminate objects.

4.
Eur J Cell Biol ; 103(4): 151454, 2024 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-39232451

RESUMO

CTCF is a key factor in three-dimensional chromatin folding and transcriptional control that was found to affect cancer cell migration by a mechanism that is still poorly understood. To identify this mechanism, we used mouse melanoma cells with a partial loss of function (pLoF) of CTCF. We found that CTCF pLoF inhibits cell migration rate while leading to an increase in the expression of multiple enzymes in the cholesterol biosynthesis pathway along with an elevation in the cellular cholesterol level. In agreement with the cholesterol change we detected altered membrane dynamics in CTCF pLoF cells as measured by reduced formation of migrasomes, extracellular vesicles formed at the rear side of migrating cells. Inhibition of cholesterol synthesis in CTCF pLoF cells restored the cellular migration rate and migrasome formation, suggesting that CTCF supports cell migration by suppressing cholesterol synthesis. Detailed analysis of the promoter of Hmgcs1, an early enzyme in the cholesterol synthesis pathway, revealed that CTCF prevents formation of a loop between that promoter and another promoter 200 kb away. CTCF also supports PRC2 recruitment to the promoter and deposition of H3K27me3. H3K27me3 at the promoter of Hmgcs1 prevents SREBP2 binding and activation of transcription. By this mechanism, CTCF fine-tunes cholesterol levels to support cell migration. Notably, genome wide association studies suggest a link between CTCF and cholesterol-associated diseases, thus CTCF emerges as a new regulator of cholesterol biosynthesis.

5.
Cell Rep ; 42(8): 112864, 2023 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-37494182

RESUMO

Lymphocyte priming in lymph nodes (LNs) was postulated to depend on the formation of stable T cell receptor (TCR)-specific immune synapses (ISs) with antigen (Ag)-presenting dendritic cells (DCs). The high-affinity LFA-1 ligand ICAM-1 was implicated in different ISs studied in vitro. We dissect the in vivo roles of endogenous DC ICAM-1 in Ag-stimulated T cell proliferation and differentiation and find that under type 1 polarizing conditions in vaccinated or vaccinia virus-infected skin-draining LNs, Ag-presenting DCs engage in ICAM-1-dependent stable conjugates with a subset of Ag-specific CD8 blasts. Nevertheless, in the absence of these conjugates, CD8 lymphocyte proliferation and differentiation into functional cytotoxic T cells (CTLs) and skin homing effector lymphocytes takes place normally. Our results suggest that although CD8 T cell blasts engage in tight ICAM-1-dependent DC-T ISs, firm ISs are dispensable for TCR-triggered proliferation and differentiation into productive effector lymphocytes.


Assuntos
Células Dendríticas , Molécula 1 de Adesão Intercelular , Molécula 1 de Adesão Intercelular/metabolismo , Células Dendríticas/metabolismo , Linfócitos T CD8-Positivos , Ativação Linfocitária , Antígenos/metabolismo , Diferenciação Celular , Receptores de Antígenos de Linfócitos T/metabolismo
6.
bioRxiv ; 2023 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-36712070

RESUMO

Mitosis is an essential process in which the duplicated genome is segregated equally into two daughter cells. CTCF has been reported to be present in mitosis but its importance for mitotic fidelity remains to be determined. To evaluate the importance of CTCF in mitosis, we tracked mitotic behaviors in wild type and two different CTCF CRISPR-based genetic knockdowns. We find that knockdown of CTCF results in prolonged mitoses and failed anaphase segregation via time lapse imaging of SiR-DNA. CTCF knockdown did not alter cell cycling or the mitotic checkpoint, which was activated upon nocodazole treatment. Immunofluorescence imaging of the mitotic spindle in CTCF knockdowns revealed disorganization via tri/tetrapolar spindles and chromosomes behind the spindle pole. Imaging of interphase nuclei showed that nuclear size increased drastically, consistent with failure to divide the duplicated genome in anaphase. Population measurements of nuclear shape in CTCF knockdowns do not display decreased circularity or increased nuclear blebbing relative to wild type. However, failed mitoses do display abnormal nuclear morphologies relative to successful mitoses, suggesting population images do not capture individual behaviors. Thus, CTCF is important for both proper metaphase organization and anaphase segregation which impacts the size and shape of the interphase nucleus.

7.
Front Immunol ; 13: 849701, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35911772

RESUMO

Breast tumors and their derived circulating cancer cells express the leukocyte ß2 integrin ligand Intercellular adhesion molecule 1 (ICAM-1). We found that elevated ICAM-1 expression in breast cancer cells results in a favorable outcome and prolonged survival of breast cancer patients. We therefore assessed the direct in vivo contribution of ICAM-1 expressed by breast cancer cells to breast tumorigenesis and lung metastasis in syngeneic immunocompetent mice hosts using spontaneous and experimental models of the lung metastasis of the C57BL/6-derived E0771 cell line, a luminal B breast cancer subtype. Notably, the presence of ICAM-1 on E0771 did not alter tumor growth or the leukocyte composition in the tumor microenvironment. Interestingly, the elimination of Tregs led to the rapid killing of primary tumor cells independently of tumor ICAM-1 expression. The in vivo elimination of a primary E0771 tumor expressing the ovalbumin (OVA) model neoantigen by the OVA-specific OVA-tcr-I mice (OT-I) transgenic cytotoxic T lymphocytes (CTLs) also took place normally in the absence of ICAM-1 expression by E0771 breast cancer target cells. The whole lung imaging of these cells by light sheet microscopy (LSM) revealed that both Wild type (WT)- and ICAM-1-deficient E0771 cells were equally disseminated from resected tumors and accumulated inside the lung vasculature at similar magnitudes. ICAM-1-deficient breast cancer cells developed, however, much larger metastatic lesions than their control counterparts. Strikingly, the vast majority of these cells gave rise to intravascular tumor colonies both in spontaneous and experimental metastasis models. In the latter model, ICAM-1 expressing E0771- but not their ICAM-1-deficient counterparts were highly susceptible to elimination by neutrophils adoptively transferred from E0771 tumor-bearing donor mice. Ex vivo, neutrophils derived from tumor-bearing mice also killed cultured E0771 cells via ICAM-1-dependent interactions. Collectively, our results are a first indication that ICAM-1 expressed by metastatic breast cancer cells that expand inside the lung vasculature is involved in innate rather than in adaptive cancer cell killing. This is also a first indication that the breast tumor expression of ICAM-1 is not required for CTL-mediated killing but can function as a suppressor of intravascular breast cancer metastasis to lungs.


Assuntos
Neoplasias Pulmonares , Linfócitos T Citotóxicos , Animais , Linhagem Celular Tumoral , Molécula 1 de Adesão Intercelular/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina , Microambiente Tumoral
8.
Cancers (Basel) ; 13(10)2021 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-34069191

RESUMO

The mechanisms by which the nuclear lamina of tumor cells influences tumor growth and migration are highly disputed. Lamin A and its variant lamin C are key lamina proteins that control nucleus stiffness and chromatin conformation. Downregulation of lamin A/C in two prototypic metastatic lines, B16F10 melanoma and E0771 breast carcinoma, facilitated cell squeezing through rigid pores, and reduced heterochromatin content. Surprisingly, both lamin A/C knockdown cells grew poorly in 3D spheroids within soft agar, and lamin A/C deficient cells derived from spheroids transcribed lower levels of the growth regulator Yap1. Unexpectedly, the transendothelial migration of both cancer cells in vitro and in vivo, through lung capillaries, was not elevated by lamin A/C knockdown and their metastasis in lungs was even dramatically reduced. Our results are the first indication that reduced lamin A/C content in distinct types of highly metastatic cancer cells does not elevate their transendothelial migration (TEM) capacity and diapedesis through lung vessels but can compromise lung metastasis at a post extravasation level.

SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA