Browsing by Subject "Mouse embryonic fibroblasts"

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    Histone H3.3 regulates mitotic progression in mouse embryonic fibroblasts
    (Canadian Science Publishing, 2017) Ors, A.; Papin, C.; Favier, B.; Roulland, Y.; Dalkara, D.; Ozturk, M.; Hamiche, A.; Dimitrov, S.; Padmanabhan, K.
    H3.3 is a histone variant that marks transcription start sites as well as telomeres and heterochromatic sites on the genome. The presence of H3.3 is thought to positively correlate with the transcriptional status of its target genes. Using a conditional genetic strategy against H3.3B, combined with short hairpin RNAs against H3.3A, we essentially depleted all H3.3 gene expression in mouse embryonic fibroblasts. Following nearly complete loss of H3.3 in the cells, our transcriptomic analyses show very little impact on global gene expression or on the localization of histone variant H2A.Z. Instead, fibroblasts displayed slower cell growth and an increase in cell death, coincident with large-scale chromosome misalignment in mitosis and large polylobed or micronuclei in interphase cells. Thus, we conclude that H3.3 may have an important under-explored additional role in chromosome segregation, nuclear structure, and the maintenance of genome integrity. © 2017 Published by NRC Research Press.
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    Molecular mechanisms of PI3K isoform dependence in embryonic growth
    (Galenos Yayınevi, 2024-08-29) Atıcı, Sena; Çizmecioğlu, Onur
    Objective The phosphoinositide 3-kinase (PI3K) pathway is an important signaling mechanism for cell proliferation and metabolism. Mutations that activate PIK3CA may make cells p110a dependent, but when phosphatase tensin homolog (PTEN) is lost, the p110b isoform of PI3Ks becomes more important. However, the exact mechanism underlying the prevalence of p110s remains unclear. In this study, our aim was to elucidate the processes behind PI3K isoform dependency in a cellular model of embryonic development. Material and Methods In order to understand PI3K isoform prevalence, mouse embryonic fibroblasts (MEFs) were used and p110b, PTEN and Rac1 activity was modulated using retroviral plasmids. Expression levels and cellular growth were assessed by performing immunoblots and crystal violet assays. Results The levels of PTEN had only a partial effect on the prevalence of PI3K isoforms in MEFs. The dependency on p110a diminished when PTEN was depleted. Of note, when PTEN expression was repressed, there was no full transition in dependency from one PI3K isoform to the other. Interestingly, the viability of PTEN-depleted MEFs became less dependent on p110a and more dependent on p110b when p110b was overexpressed. Nevertheless, the overexpression of p110b in conjunction with PTEN knock-downs did not result in a complete shift of isoforms in PI3Ks. Finally, we investigated Rac1 activation with a mutant allele and determined a more potent increase in p110b prominence in MEFs. Conclusion These findings suggest that multiple cellular parameters, including PTEN status, PI3K isoform levels, and Rac1 activity, combine to influence PI3K isoform prevalence, rather than a single determinant.