Cell cycle synchronization and BrdU incorporation as a tool to study the possible selective elimination of ErbB1 gene in the micronuclei in A549 cells

Lauand, Camila; Niero, Evandro Luís de Oliveira; Dias, Viviane Miranda; Machado-Santelli, Gláucia Maria

doi:10.1590/1414-431x20144262

Cited by 6 publications

(5 citation statements)

References 38 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…To dissociate the effect of p53 on HR from its cell cycle effect, we used nocodazole-synchronized cells pulse-treated with CPT at the entrance to S phase. The doubling time of A549 cells in our hands is 38 hours with S phase lasting approximately 16 hours (unpublished data), a time slightly shorter than that reported previously [ 35 ]. A significant delay of S phase duration (up to 27 hours) was observed following pulse-treatment with CPT (Figure 3A ), with all derivatives, wild-type, p53 - or p21 Waf1 -depleted cells proceeding through S phase with a similar rate and entering G2 (Figure 3A ).…”

Section: Resultscontrasting

confidence: 51%

Transcriptional activation of p21Waf1 contributes to suppression of HR by p53 in response to replication arrest induced by camptothecin

2018

View full text Add to dashboard Cite

The inhibitory effect of p53 on homologous recombination (HR) is exerted through sequestration of replication protein A (RPA). Release of the p53/RPA complex in response to replication stress is crucially dependent on the phosphorylation status of both proteins and is required for efficient DNA repair by HR. Phosphorylation of RPA within its RPA2 subunit by cyclin-dependent kinases (CDK) is an early event in the replication stress response. Here we investigated the role of transcriptional activation of the p53 downstream target, p21Waf1, on RPA2 phosphorylation, the stability of the p53/RPA complex and HR in cells undergoing replication arrest induced by camptothecin (CPT). We show that in CPT-treated cells, activation of p53 and p21Waf1 impedes RPA2 phosphorylation, while their depletion by siRNA stimulates it. The p53/RPA complex is more stable in wild-type cells than in cells depleted of p21Waf1. We used nocodazole-synchronized cells treated with CPT at the entrance to S phase to assess rates of HR. Regardless of their p53 or p21Waf1 status, the cells proceed through S phase at a similar rate and enter G2. While HR is low in wild-type cells and high in p53-depleted cells, only partial inhibition of HR is observed in the p21Waf1-depleted cells. This correlates with the extent of RPA sequestration by p53. Thus, in CPT-treated cells, p53-induced transcriptional activation of p21Waf1 regulates RPA2 phosphorylation, the stability of the p53/RPA complex and HR.

show abstract

Section: Resultscontrasting

confidence: 51%

Transcriptional activation of p21Waf1 contributes to suppression of HR by p53 in response to replication arrest induced by camptothecin

2018

View full text Add to dashboard Cite

show abstract

“…As an analog of thymidine, BrdU is incorporated into the newly synthesized DNA of replicating cells by substituting for thymidine during DNA replication. The expression of BrdU thus identifies cells that are actively replicating their DNA 15 . We administered BrdU 4 hours before harvesting the skin of mice; thus, the expression of BrdU in these samples revealed cells that had replicated in those 4 hours.…”

Section: Discussionmentioning

confidence: 99%

Immunohistochemical study of hair follicle stem cells in regenerated hair follicles induced by Wnt10b

Zhang¹,

Xing²,

Guo³

et al. 2016

Int. J. Med. Sci.

View full text Add to dashboard Cite

The regulation of the periodic regeneration of hair follicles is complicated. Although Wnt10b has been reported to induce hair follicle regeneration, the characteristics of induced hair follicles, especially the target cells of Wnt10b, have not yet been clearly elucidated. Thus, we systematically evaluated the expression and proliferation patterns of Wnt10b-induced hair follicles. We found that Wnt10b promoted the proliferation of hair follicle stem cells from 24 hours after AdWnt10b injection. Seventy-two hours after AdWnt10b injection, cells outside of bulge area began to proliferate. When the induced hair follicle entered full anagen, although the hair follicle stem cells were normal, canonical Wnt signaling was maintained in the hair precortex cells. Our results reveal that the target cells that overexpressed Wnt10b included hair follicle stem cells, hair precortex cells, and matrix cells.

show abstract

“…Extrapolated growth curves were constructed over a period of fourteen days by serial passaging and cell counting with a hemocytometer or by PrestoBlue® assay (Thermo Fisher Scientific, UK). Cell proliferation was also analyzed using the CellTrace™ CFSE Cell Proliferation Kit (Invitrogen, USA) in cells synchronized by gradual serum deprivation following published protocols 25 . Cells were harvested at 48 hours and subjected to division peak resolution by flow cytometry.…”

Section: Methodsmentioning

confidence: 99%

ARID2 deficiency promotes tumor progression and is associated with higher sensitivity to chemotherapy in lung cancer

et al. 2021

View full text Add to dashboard Cite

The survival rate in lung cancer remains stubbornly low and there is an urgent need for the identification of new therapeutic targets. In the last decade, several members of the SWI/SNF chromatin remodeling complexes have been described altered in different tumor types. Nevertheless, the precise mechanisms of their impact on cancer progression, as well as the application of this knowledge to cancer patient management are largely unknown. In this study, we performed targeted sequencing of a cohort of lung cancer patients on genes involved in chromatin structure. Additionally, we studied at the protein level the expression of these genes in cancer samples and performed functional experiments to identify the molecular mechanisms linking alterations of chromatin remodeling genes and tumor development. Remarkably, we found that 20% of lung cancer patients show ARID2 protein loss, partially explained by the presence of ARID2 mutations. Additionally, we showed that ARID2-deficiency provokes profound chromatin structural changes altering cell transcriptional programmes which bolsters the proliferative and metastatic potential of the cells both in vitro and in vivo . Moreover, we demonstrated that ARID2 deficiency impairs DNA repair, enhancing the sensitivity of the cells to DNA damaging agents. Our findings support that ARID2 is a bona-fide tumor suppressor gene in lung cancer that may be exploited therapeutically.

show abstract

Cell cycle synchronization and BrdU incorporation as a tool to study the possible selective elimination of ErbB1 gene in the micronuclei in A549 cells

Cited by 6 publications

References 38 publications

Transcriptional activation of p21Waf1 contributes to suppression of HR by p53 in response to replication arrest induced by camptothecin

Transcriptional activation of p21Waf1 contributes to suppression of HR by p53 in response to replication arrest induced by camptothecin

Immunohistochemical study of hair follicle stem cells in regenerated hair follicles induced by Wnt10b

ARID2 deficiency promotes tumor progression and is associated with higher sensitivity to chemotherapy in lung cancer

Contact Info

Product

Resources

About