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1.
J Cell Sci ; 136(22)2023 11 15.
Article in English | MEDLINE | ID: mdl-37921359

ABSTRACT

The nucleolus is sensitive to stress and can orchestrate a chain of cellular events in response to stress signals. Despite being a growth factor, FGF2 has antiproliferative and tumor-suppressive functions in some cellular contexts. In this work, we investigated how the antiproliferative effect of FGF2 modulates chromatin-, nucleolus- and rDNA-associated proteins. The chromatin and nucleolar proteome indicated that FGF2 stimulation modulates proteins related to transcription, rRNA expression and chromatin-remodeling proteins. The global transcriptional rate and nucleolus area increased along with nucleolar disorganization upon 24 h of FGF2 stimulation. FGF2 stimulation induced immature rRNA accumulation by increasing rRNA transcription. The rDNA-associated protein analysis reinforced that FGF2 stimulus interferes with transcription and rRNA processing. RNA Pol I inhibition partially reversed the growth arrest induced by FGF2, indicating that changes in rRNA expression might be crucial for triggering the antiproliferative effect. Taken together, we demonstrate that the antiproliferative FGF2 stimulus triggers significant transcriptional changes and modulates the main cell transcription site, the nucleolus.


Subject(s)
Cell Nucleolus , Fibroblast Growth Factor 2 , Fibroblast Growth Factor 2/genetics , Fibroblast Growth Factor 2/pharmacology , Fibroblast Growth Factor 2/metabolism , Cell Nucleolus/metabolism , RNA, Ribosomal/genetics , RNA, Ribosomal/metabolism , Transcription, Genetic , DNA, Ribosomal/genetics , Chromatin/genetics , Chromatin/metabolism
2.
Nucleic Acids Res ; 46(19): 10095-10105, 2018 11 02.
Article in English | MEDLINE | ID: mdl-30102332

ABSTRACT

ELL family transcription factors activate the overall rate of RNA polymerase II (Pol II) transcription elongation by binding directly to Pol II and suppressing its tendency to pause. In metazoa, ELL regulates Pol II transcription elongation as part of a large multisubunit complex referred to as the Super Elongation Complex (SEC), which includes P-TEFb and EAF, AF9 or ENL, and an AFF family protein. Although orthologs of ELL and EAF have been identified in lower eukaryotes including Schizosaccharomyces pombe, it has been unclear whether SEC-like complexes function in lower eukaryotes. In this report, we describe isolation from S. pombe of an ELL-containing complex with features of a rudimentary SEC. This complex includes S. pombe Ell1, Eaf1, and a previously uncharacterized protein we designate Ell1 binding protein 1 (Ebp1), which is distantly related to metazoan AFF family members. Like the metazoan SEC, this S. pombe ELL complex appears to function broadly in Pol II transcription. Interestingly, it appears to have a particularly important role in regulating genes involved in cell separation.


Subject(s)
RNA Polymerase II/genetics , Schizosaccharomyces pombe Proteins/genetics , Transcription Factors/genetics , Transcriptional Elongation Factors/genetics , Positive Transcriptional Elongation Factor B/chemistry , Positive Transcriptional Elongation Factor B/genetics , RNA Polymerase II/chemistry , Schizosaccharomyces/genetics , Schizosaccharomyces pombe Proteins/chemistry , Transcription Factors/chemistry , Transcription, Genetic , Transcriptional Elongation Factors/chemistry
3.
Proteomics ; 18(17): e1800203, 2018 09.
Article in English | MEDLINE | ID: mdl-30035358

ABSTRACT

Fibroblast growth factor 2 (FGF2) is a well-known cell proliferation promoter; however, it can also induce cell cycle arrest. To gain insight into the molecular mechanisms of this antiproliferative effect, for the first time, the early systemic proteomic differences induced by this growth factor in a K-Ras-driven mouse tumor cell line using a quantitative proteomics approach are investigated. More than 2900 proteins are quantified, indicating that terms associated with metabolism, RNA processing, replication, and transcription are enriched among proteins differentially expressed upon FGF2 stimulation. Proteomic trend dynamics indicate that, for proteins mainly associated with DNA replication and carbohydrate metabolism, an FGF2 stimulus delays their abundance changes, whereas FGF2 stimulation accelerates other metabolic programs. Transcription regulatory network analysis indicates master regulators of FGF2 stimulation, including two critical transcription factors, FOSB and JUNB. Their expression dynamics, both in the Y1 cell line (a murine model of adenocarcinoma cells) and in two other human cell lines (SK-N-MC and UM-UC-3) also susceptible to FGF2 antiproliferative effects, are investigated. Both protein expression levels depend on fibroblast growth factor receptor (FGFR) and src signaling. JUNB and FOSB knockdown do not rescue cells from the growth arrest induced by FGF2; however, FOSB knockdown rescue cells from DNA replication delay, indicating that FOSB expression underlies one of the FGF2 antiproliferative effects, namely, S-phase progression delay.


Subject(s)
Adrenal Cortex Neoplasms/metabolism , Adrenocortical Carcinoma/metabolism , Cell Proliferation , Fibroblast Growth Factor 2/pharmacology , Proteome/metabolism , Proto-Oncogene Proteins c-fos/metabolism , Transcription Factors/metabolism , Adrenal Cortex Neoplasms/drug therapy , Adrenal Cortex Neoplasms/pathology , Adrenocortical Carcinoma/drug therapy , Adrenocortical Carcinoma/pathology , Animals , Humans , Mice , Protein Interaction Maps , Proteome/analysis , Proto-Oncogene Proteins p21(ras)/genetics , Signal Transduction , Tumor Cells, Cultured , Urinary Bladder Neoplasms/drug therapy , Urinary Bladder Neoplasms/metabolism , Urinary Bladder Neoplasms/pathology
4.
Proteomics, v. 18, n. 17, 18002013, jul. 2018
Article in English | Sec. Est. Saúde SP, SESSP-IBPROD, Sec. Est. Saúde SP | ID: bud-2657

ABSTRACT

Fibroblast growth factor 2 (FGF2) is a well-known cell proliferation promoter; however, it can also induce cell cycle arrest. To gain insight into the molecular mechanisms of this antiproliferative effect, for the first time, the early systemic proteomic differences induced by this growth factor in a K-Ras-driven mouse tumor cell line using a quantitative proteomics approach are investigated. More than 2900 proteins are quantified, indicating that terms associated with metabolism, RNA processing, replication, and transcription are enriched among proteins differentially expressed upon FGF2 stimulation. Proteomic trend dynamics indicate that, for proteins mainly associated with DNA replication and carbohydrate metabolism, an FGF2 stimulus delays their abundance changes, whereas FGF2 stimulation accelerates other metabolic programs. Transcription regulatory network analysis indicates master regulators of FGF2 stimulation, including two critical transcription factors, FOSB and JUNB. Their expression dynamics, both in the Y1 cell line (a murine model of adenocarcinoma cells) and in two other human cell lines (SK-N-MC and UM-UC-3) also susceptible to FGF2 antiproliferative effects, are investigated. Both protein expression levels depend on fibroblast growth factor receptor (FGFR) and src signaling. JUNB and FOSB knockdown do not rescue cells from the growth arrest induced by FGF2; however, FOSB knockdown rescue cells from DNA replication delay, indicating that FOSB expression underlies one of the FGF2 antiproliferative effects, namely, S-phase progression delay.

5.
Proteomics ; 18(17): 1800203, 2018.
Article in English | Sec. Est. Saúde SP, SESSP-IBPROD, Sec. Est. Saúde SP | ID: but-ib15799

ABSTRACT

Fibroblast growth factor 2 (FGF2) is a well-known cell proliferation promoter; however, it can also induce cell cycle arrest. To gain insight into the molecular mechanisms of this antiproliferative effect, for the first time, the early systemic proteomic differences induced by this growth factor in a K-Ras-driven mouse tumor cell line using a quantitative proteomics approach are investigated. More than 2900 proteins are quantified, indicating that terms associated with metabolism, RNA processing, replication, and transcription are enriched among proteins differentially expressed upon FGF2 stimulation. Proteomic trend dynamics indicate that, for proteins mainly associated with DNA replication and carbohydrate metabolism, an FGF2 stimulus delays their abundance changes, whereas FGF2 stimulation accelerates other metabolic programs. Transcription regulatory network analysis indicates master regulators of FGF2 stimulation, including two critical transcription factors, FOSB and JUNB. Their expression dynamics, both in the Y1 cell line (a murine model of adenocarcinoma cells) and in two other human cell lines (SK-N-MC and UM-UC-3) also susceptible to FGF2 antiproliferative effects, are investigated. Both protein expression levels depend on fibroblast growth factor receptor (FGFR) and src signaling. JUNB and FOSB knockdown do not rescue cells from the growth arrest induced by FGF2; however, FOSB knockdown rescue cells from DNA replication delay, indicating that FOSB expression underlies one of the FGF2 antiproliferative effects, namely, S-phase progression delay.

6.
Genes Dev ; 25(8): 801-13, 2011 Apr 15.
Article in English | MEDLINE | ID: mdl-21498570

ABSTRACT

In the absence of growth signals, cells exit the cell cycle and enter into G0 or quiescence. Alternatively, cells enter senescence in response to inappropriate growth signals such as oncogene expression. The molecular mechanisms required for cell cycle exit into quiescence or senescence are poorly understood. The DREAM (DP, RB [retinoblastoma], E2F, and MuvB) complex represses cell cycle-dependent genes during quiescence. DREAM contains p130, E2F4, DP1, and a stable core complex of five MuvB-like proteins: LIN9, LIN37, LIN52, LIN54, and RBBP4. In mammalian cells, the MuvB core dissociates from p130 upon entry into the cell cycle and binds to BMYB during S phase to activate the transcription of genes expressed late in the cell cycle. We used mass spectroscopic analysis to identify phosphorylation sites that regulate the switch of the MuvB core from BMYB to DREAM. Here we report that DYRK1A can specifically phosphorylate LIN52 on serine residue 28, and that this phosphorylation is required for DREAM assembly. Inhibiting DYRK1A activity or point mutation of LIN52 disrupts DREAM assembly and reduces the ability of cells to enter quiescence or undergo Ras-induced senescence. These data reveal an important role for DYRK1A in the regulation of DREAM activity and entry into quiescence.


Subject(s)
Cellular Senescence/physiology , Protein Serine-Threonine Kinases/metabolism , Protein-Tyrosine Kinases/metabolism , Animals , Cell Cycle/genetics , Cell Cycle/physiology , Cell Line , Cell Line, Tumor , Cell Proliferation , Cells, Cultured , Cellular Senescence/genetics , E2F4 Transcription Factor/genetics , E2F4 Transcription Factor/metabolism , Humans , Mice , NIH 3T3 Cells , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Phosphorylation , Protein Serine-Threonine Kinases/genetics , Protein-Tyrosine Kinases/genetics , Retinoblastoma-Binding Protein 4/genetics , Retinoblastoma-Binding Protein 4/metabolism , Retinoblastoma-Like Protein p130/genetics , Retinoblastoma-Like Protein p130/metabolism , Trans-Activators/genetics , Trans-Activators/metabolism , Transcription Factor DP1/genetics , Transcription Factor DP1/metabolism , Tumor Suppressor Proteins/genetics , Tumor Suppressor Proteins/metabolism , Dyrk Kinases
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