scholarly journals p62cdc23 of Saccharomyces cerevisiae: a nuclear tetratricopeptide repeat protein with two mutable domains.

1993 ◽  
Vol 13 (2) ◽  
pp. 1212-1221 ◽  
Author(s):  
R S Sikorski ◽  
W A Michaud ◽  
P Hieter
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle Progression ◽  
Single Amino Acid ◽  
Temperature Sensitive ◽  
Cycle Progression ◽  
Tetratricopeptide Repeats ◽  
Tetratricopeptide Repeat Protein ◽  
Cdc Genes

CDC23 is required in Saccharomyces cerevisiae for cell cycle progression through the G2/M transition. The CDC23 gene product contains tandem, imperfect repeats, termed tetratricopeptide repeats, (TPR) units common to a protein family that includes several other nuclear division CDC genes. In this report we have used mutagenesis to probe the functional significance of the TPR units within CDC23. Analysis of truncated derivatives indicates that the TPR block of CDC23 is necessary for the function or stability of the polypeptide. In-frame deletion of a single TPR unit within the repeat block proved sufficient to inactivate CDC23 in vivo, though this allele could rescue the temperature-sensitive defect of a cdc23 point mutant by intragenic complementation. By both in vitro and in vivo mutagenesis techniques, 17 thermolabile cdc23 alleles were produced and examined. Fourteen alleles contained single amino acid changes that were found to cluster within two distinct mutable domains, one of which encompasses the most canonical TPR unit found in CDC23. In addition, we have characterized CDC23 as a 62-kDa protein (p62cdc23) that is localized to the yeast nucleus. Our mutagenesis results suggest that TPR blocks form an essential domain within members of the TPR family.

scholarly journals p62cdc23 of Saccharomyces cerevisiae: a nuclear tetratricopeptide repeat protein with two mutable domains

1993 ◽  
Vol 13 (2) ◽  
pp. 1212-1221
Author(s):  
R S Sikorski ◽  
W A Michaud ◽  
P Hieter
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle Progression ◽  
Single Amino Acid ◽  
Temperature Sensitive ◽  
Cycle Progression ◽  
Tetratricopeptide Repeats ◽  
Tetratricopeptide Repeat Protein ◽  
Cdc Genes

CDC23 is required in Saccharomyces cerevisiae for cell cycle progression through the G2/M transition. The CDC23 gene product contains tandem, imperfect repeats, termed tetratricopeptide repeats, (TPR) units common to a protein family that includes several other nuclear division CDC genes. In this report we have used mutagenesis to probe the functional significance of the TPR units within CDC23. Analysis of truncated derivatives indicates that the TPR block of CDC23 is necessary for the function or stability of the polypeptide. In-frame deletion of a single TPR unit within the repeat block proved sufficient to inactivate CDC23 in vivo, though this allele could rescue the temperature-sensitive defect of a cdc23 point mutant by intragenic complementation. By both in vitro and in vivo mutagenesis techniques, 17 thermolabile cdc23 alleles were produced and examined. Fourteen alleles contained single amino acid changes that were found to cluster within two distinct mutable domains, one of which encompasses the most canonical TPR unit found in CDC23. In addition, we have characterized CDC23 as a 62-kDa protein (p62cdc23) that is localized to the yeast nucleus. Our mutagenesis results suggest that TPR blocks form an essential domain within members of the TPR family.

scholarly journals Activation of the Bur1-Bur2 Cyclin-Dependent Kinase Complex by Cak1

2002 ◽  
Vol 22 (19) ◽  
pp. 6750-6758 ◽  
Author(s):  
Sheng Yao ◽  
Gregory Prelich
Keyword(s):  
Cell Cycle ◽  
Rna Polymerase Ii ◽  
Cell Cycle Progression ◽  
Single Gene ◽  
Eukaryotic Cell ◽  
Temperature Sensitive ◽  
Cycle Progression ◽  
Carboxy Terminal

ABSTRACT Cyclin-dependent kinases (Cdks) were originally identified as regulators of eukaryotic cell cycle progression, but several Cdks were subsequently shown to perform important roles as transcriptional regulators. While the mechanisms regulating the Cdks involved in cell cycle progression are well documented, much less is known regarding how the Cdks that are involved in transcription are regulated. In Saccharomyces cerevisiae, Bur1 and Bur2 comprise a Cdk complex that is involved in transcriptional regulation, presumably mediated by its phosphorylation of the carboxy-terminal domain (CTD) of the largest subunit of RNA polymerase II. To investigate the regulation of Bur1 in vivo, we searched for high-copy-number suppressors of a bur1 temperature-sensitive mutation, identifying a single gene, CAK1. Cak1 is known to activate two other Cdks in yeast by phosphorylating a threonine within their conserved T-loop domains. Bur1 also has the conserved threonine within its T loop and is therefore a potential direct target of Cak1. Additional tests establish a direct functional interaction between Cak1 and the Bur1-Bur2 Cdk complex: Bur1 is phosphorylated in vivo, both the conserved Bur1 T-loop threonine and Cak1 are required for phosphorylation and Bur1 function in vivo, and recombinant Cak1 stimulates CTD kinase activity of the purified Bur1-Bur2 complex in vitro. Thus, both genetic and biochemical evidence demonstrate that Cak1 is a physiological regulator of the Bur1 kinase.

scholarly journals Esa1p Is an Essential Histone Acetyltransferase Required for Cell Cycle Progression

1999 ◽  
Vol 19 (4) ◽  
pp. 2515-2526 ◽  
Author(s):  
Astrid S. Clarke ◽  
Joanna E. Lowell ◽  
Sandra J. Jacobson ◽  
Lorraine Pillus
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Histone Acetyltransferase ◽  
Essential Gene ◽  
Cell Cycle Control ◽  
Biological Significance ◽  
Temperature Sensitive ◽  
Cycle Progression

ABSTRACT Histones are dynamically modified during chromatin assembly, as specific transcriptional patterns are established, and during mitosis and development. Modifications include acetylation, phosphorylation, ubiquitination, methylation, and ADP-ribosylation, but the biological significance of each of these is not well understood. For example, distinct acetylation patterns correlate with nucleosome formation and with transcriptionally activated or silenced chromatin, yet mutations in genes encoding several yeast histone acetyltransferase (HAT) activities result in either no cellular phenotype or only modest growth defects. Here we report characterization of ESA1, an essential gene that is a member of the MYST family that includes two yeast silencing genes, human genes associated with leukemia and with the human immunodeficiency virus type 1 Tat protein, and Drosophila mof, a gene essential for male dosage compensation. Esa1p acetylates histones in a pattern distinct from those of other yeast enzymes, and temperature-sensitive mutant alleles abolish enzymatic activity in vitro and result in partial loss of an acetylated isoform of histone H4 in vivo. Strains carrying these mutations are also blocked in the cell cycle such that at restrictive temperatures,esa1 mutants succeed in replicating their DNA but fail to proceed normally through mitosis and cytokinesis. Recent studies show that Esa1p enhances transcription in vitro and thus may modulate expression of genes important for cell cycle control. These observations therefore link an essential HAT activity to cell cycle progression, potentially through discrete transcriptional regulatory events.

Functional interaction between p21rap1A and components of the budding pathway in Saccharomyces cerevisiae

1992 ◽  
Vol 12 (9) ◽  
pp. 4084-4092
Author(s):  
P C McCabe ◽  
H Haubruck ◽  
P Polakis ◽  
F McCormick ◽  
M A Innis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mammalian Cells ◽  
Bound States ◽  
Yeast Cells ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Amino Terminal ◽  
Loop Region

The rap1A gene encodes a 21-kDa, ras-related GTP-binding protein (p21rap1A) of unknown function. A close structural homolog of p21rap1A (65% identity in the amino-terminal two-thirds) is the RSR1 gene product (Rsr1p) of Saccharomyces cerevisiae. Although Rsr1p is not essential for growth, its presence is required for nonrandom selection of bud sites. To assess the similarity of these proteins at the functional level, wild-type and mutant forms of p21rap1A were tested for complementation of activities known to be fulfilled by Rsr1p. Expression of p21rap1A, like multicopy expression of RSR1, suppressed the conditional lethality of a temperature-sensitive cdc24 mutation. Point mutations predicted to affect the localization of p21rap1A or its ability to cycle between GDP and GTP-bound states disrupted suppression of cdc24ts, while other mutations in the 61-65 loop region improved suppression. Expression of p21rap1A could not, however, suppress the random budding phenotype of rsr1 cells. p21rap1A also apparently interfered with the normal activity of Rsrlp, causing random budding in diploid wild-type cells, suggesting an inability of p21rap1A to interact appropriately with Rsr1p regulatory proteins. Consistent with this hypothesis, we found an Rsr1p-specific GTPase-activating protein (GAP) activity in yeast membranes which was not active toward p21rap1A, indicating that p21rap1A may be predominantly GTP bound in yeast cells. Coexpression of human Rap1-specific GAP suppressed the random budding due to expression of p21rap1A or its derivatives, including Rap1AVal-12. Although Rap1-specific GAP stimulated the GTPase of Rsr1p in vitro, it did not dominantly interfere with Rsr1p function in vivo. A chimera consisting of Rap1A1-165::Rsr1p166-272 did not exhibit normal Rsr1p function in the budding pathway. These results indicated that p21rap1A and Rsr1p share at least partial functional homology, which may have implications for p21rap1A function in mammalian cells.

scholarly journals [Corrigendum] LRIG2 promotes the proliferation and cell cycle progression of glioblastoma cells in vitro and in vivo through enhancing PDGFRβ signaling

2022 ◽  
Vol 60 (2) ◽  
Author(s):  
Qungen Xiao ◽  
Minhai Dong ◽  
Fangling Cheng ◽  
Feng Mao ◽  
Weifeng Zong ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Glioblastoma Cells ◽  
Cycle Progression

scholarly journals Translation initiation factor 4A from Saccharomyces cerevisiae: analysis of residues conserved in the D-E-A-D family of RNA helicases.

1991 ◽  
Vol 11 (7) ◽  
pp. 3463-3471 ◽  
Author(s):  
S R Schmid ◽  
P Linder
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Translation Initiation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Clear Correlation ◽  
Temperature Sensitive ◽  
Rna Helicases ◽  
Initiation Of Translation

The eukaryotic translation initiation factor 4A (eIF-4A) possesses an in vitro helicase activity that allows the unwinding of double-stranded RNA. This activity is dependent on ATP hydrolysis and the presence of another translation initiation factor, eIF-4B. These two initiation factors are thought to unwind mRNA secondary structures in preparation for ribosome binding and initiation of translation. To further characterize the function of eIF-4A in cellular translation and its interaction with other elements of the translation machinery, we have isolated mutations in the TIF1 and TIF2 genes encoding eIF-4A in Saccharomyces cerevisiae. We show that three highly conserved domains of the D-E-A-D protein family, encoding eIF-4A and other RNA helicases, are essential for protein function. Only in rare cases could we make a conservative substitution without affecting cell growth. The mutants show a clear correlation between their growth and in vivo translation rates. One mutation that results in a temperature-sensitive phenotype reveals an immediate decrease in translation activity following a shift to the nonpermissive temperature. These in vivo results confirm previous in vitro data demonstrating an absolute dependence of translation on the TIF1 and TIF2 gene products.

scholarly journals Regulation of Focal Adhesion Kinase by a Novel Protein Inhibitor FIP200

2002 ◽  
Vol 13 (9) ◽  
pp. 3178-3191 ◽  
Author(s):  
Smita Abbi ◽  
Hiroki Ueda ◽  
Chuanhai Zheng ◽  
Lee Ann Cooper ◽  
Jihe Zhao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Cell Cycle Progression ◽  
Protein Inhibitor ◽  
Cellular Functions ◽  
Cycle Progression ◽  
Novel Protein

Focal adhesion kinase (FAK) is a major mediator of integrin signaling pathways. The mechanisms of regulation of FAK activity and its associated cellular functions are not very well understood. Here, we present data suggesting that a novel protein FIP200 functions as an inhibitor for FAK. We show the association of endogenous FIP200 with FAK, which is decreased upon integrin-mediated cell adhesion concomitant with FAK activation. In vitro- and in vivo-binding studies indicate that FIP200 interacts with FAK through multiple domains directly. FIP200 bound to the kinase domain of FAK inhibited its kinase activity in vitro and its autophosphorylation in vivo. Overexpression of FIP200 or its segments inhibited cell spreading, cell migration, and cell cycle progression, which correlated with their inhibition of FAK activity in vivo. The inhibition of these cellular functions by FIP200 could be rescued by coexpression of FAK. Last, we show that disruption of the functional interaction between endogenous FIP200 with FAK leads to increased FAK phosphorylation and partial restoration of cell cycle progression in cells plated on poly-l-lysine, providing further support for FIP200 as a negative regulator of FAK. Together, these results identify FIP200 as a novel protein inhibitor for FAK.

scholarly journals Anticancer Activity of Cynomorium coccineum

Cancers ◽  
2018 ◽  
Vol 10 (10) ◽  
pp. 354 ◽  
Author(s):  
Mouna Sdiri ◽  
Xiangmin Li ◽  
William Du ◽  
Safia El-Bok ◽  
Yi-Zhen Xie ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Anticancer Activity ◽  
Cancer Cells ◽  
Cell Lines ◽  
Cancer Cell ◽  
Cell Cycle Progression ◽  
Cycle Progression

The extensive applications of Cynomorium species and their rich bioactive secondary metabolites have inspired many pharmacological investigations. Previous research has been conducted to examine the biological activities and numerous interesting pharmaceutical activities have been reported. However, the antitumor activities of these species are unclear. To understand the potential anticancer activity, we screened Cynomorium coccineum and Cynomorium songaricum using three different extracts of each species. In this study, the selected extracts were evaluated for their ability to decrease survival rates of five different cancer cell lines. We compared the cytotoxicity of the three different extracts to the anticancer drug vinblastine and one of the most well-known medicinal mushrooms Amaurederma rude. We found that the water and alcohol extracts of C. coccineum at the very low concentrations possessed very high capacity in decreasing the cancer cells viability with a potential inhibition of tumorigenesis. Based on these primitive data, we subsequently tested the ethanol and the water extracts of C. coccineum, respectively in in vitro and in vivo assays. Cell cycle progression and induction of programmed cell death were investigated at both biological and molecular levels to understand the mechanism of the antitumor inhibitory action of the C. coccineum. The in vitro experiments showed that the treated cancer cells formed fewer and smaller colonies than the untreated cells. Cell cycle progression was inhibited, and the ethanol extract of C. coccineum at a low concentration induced accumulation of cells in the G1 phase. We also found that the C. coccineum’s extracts suppressed viability of two murine cancer cell lines. In the in vivo experiments, we injected mice with murine cancer cell line B16, followed by peritoneal injection of the water extract. The treatment prolonged mouse survival significantly. The tumors grew at a slower rate than the control. Down-regulation of c-myc expression appeared to be associated with these effects. Further investigation showed that treatment with C. coccineum induced the overexpression of the tumor suppressor Foxo3 and other molecules involved in inducing autophagy. These results showed that the C. coccineum extract exerts its antiproliferative activity through the induction of cell death pathway. Thus, the Cynomorium plants appear to be a promising source of new antineoplastic compounds.

Bioflavonoids protect cells against halogenated boroxine-induced genotoxic damage by upregulation of hTERT expression

2019 ◽  
Vol 74 (5-6) ◽  
pp. 125-129 ◽  
Author(s):  
Maida Hadzic ◽  
Sanin Haveric ◽  
Anja Haveric ◽  
Naida Lojo-Kadric ◽  
Borivoj Galic ◽  
...  
Keyword(s):  
Cell Cycle Progression ◽  
Housekeeping Genes ◽  
Inhibitory Effects ◽  
Human Telomerase Reverse Transcriptase ◽  
Cycle Progression ◽  
Genotoxic Damage ◽  
Malignant Skin ◽  
Human Telomerase

Abstract Plant bioflavonoids are widely present in the human diet and have various protective properties. In this study, we have demonstrated the capacity of delphinidin and luteolin to increase human telomerase reverse transcriptase (hTERT) expression level and act as protective agents against halogenated boroxine-induced genotoxic damage. Halogenated boroxine K2(B3O3F4OH) (HB), is a novel compound with potential for the treatment of both benign and malignant skin changes. In vivo and in vitro studies have confirmed the inhibitory effects of HB on carcinoma cell proliferation and cell cycle progression as well as enzyme inhibition. However, minor genotoxic effects of HB are registered in higher applied concentrations, but those can be suppressed by in vitro addition of delphinidin and luteolin in appropriate concentrations. Fresh peripheral blood samples were cultivated for 72 h followed by independent and concomitant treatments of HB with luteolin or delphinidin. We analyzed the differences in relative hTERT expression between series of treatments compared with controls, which were based on normalized ratios with housekeeping genes. The obtained results have shown that selected bioflavonoids induce upregulation of hTERT that may contribute to the repair of genotoxic damage in vitro.

scholarly journals (S)-10-Hydroxycamptothecin Inhibits Esophageal Squamous Cell Carcinoma Growth In Vitro and In Vivo Via Decreasing Topoisomerase I Enzyme Activity

Cancers ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 1964 ◽  
Author(s):  
Mengqiu Song ◽  
Shuying Yin ◽  
Ran Zhao ◽  
Kangdong Liu ◽  
Joydeb Kumar Kundu ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Enzymatic Activity ◽  
Topoisomerase I ◽  
Cell Cycle Progression ◽  
Caspase 3 ◽  
Cyclin B1 ◽  
Phase Cell ◽  
Cycle Progression

Topoisomerase (TOP) I plays a major role in the process of supercoiled DNA relaxation, thereby facilitating DNA replication and cell cycle progression. The expression and enzymatic activity of TOP I is positively correlated with tumor progression. Although the anticancer activity of (S)-10-Hydroxycamptothecin (HCPT), a TOP I specific inhibitor, has been reported in various cancers, the effect of HCPT on esophageal cancer is yet to be examined. In this study, we investigate the potential of HCPT to inhibit the growth of ESCC cells in vitro and verify its anti-tumor activity in vivo by using a patient-derived xenograft (PDX) tumor model in mice. Our study revealed the overexpression of TOP I in ESCC cells and treatment with HCPT inhibited TOP I enzymatic activity at 24 h and decreased expression at 48 h and 72 h. HCPT also induced DNA damage by increasing the expression of H2A.XS139. HCPT significantly decreased the proliferation and anchorage-independent growth of ESCC cells (KYSE410, KYSE510, KYSE30, and KYSE450). Mechanistically, HCPT inhibited the G2/M phase cell cycle transition, decreased the expression of cyclin B1, and elevated p21 expression. In addition, HCPT stimulated ESCC cells apoptosis, which was associated with elevated expression of cleaved PARP, cleaved caspase-3, cleaved caspase-7, Bax, Bim, and inhibition of Bcl-2 expression. HCPT dramatically suppressed PDX tumor growth and decreased the expression of Ki-67 and TOP I and increased the level of cleaved caspase-3 and H2A.XS139 expression. Taken together, our data suggested that HCPT inhibited ESCC growth, arrested cell cycle progression, and induced apoptosis both in vitro and in vivo via decreasing the expression and activity of TOP I enzyme.

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