RESUMEN
Fisetin is a natural flavonol present in edible vegetables, fruits and wine at 2-160 microg/g concentrations and an ingredient in nutritional supplements with much higher concentrations. The compound has been reported to exert anticarcinogenic effects as well as antioxidant and anti-inflammatory activity via its ability to act as an inhibitor of cell proliferation and free radical scavenger, respectively. Our cell-based high-throughput screen for small molecules that override chemically induced mitotic arrest identified fisetin as an antimitotic compound. Fisetin rapidly compromised microtubule drug-induced mitotic block in a proteasome-dependent manner in several human cell lines. Moreover, in unperturbed human cancer cells fisetin caused premature initiation of chromosome segregation and exit from mitosis without normal cytokinesis. To understand the molecular mechanism behind these mitotic errors, we analyzed the consequences of fisetin treatment on the localization and phoshorylation of several mitotic proteins. Aurora B, Bub1, BubR1 and Cenp-F rapidly lost their kinetochore/centromere localization and others became dephosphorylated upon addition of fisetin to the culture medium. Finally, we identified Aurora B kinase as a novel direct target of fisetin. The activity of Aurora B was significantly reduced by fisetin in vitro and in cells, an effect that can explain the observed forced mitotic exit, failure of cytokinesis and decreased cell viability. In conclusion, our data propose that fisetin perturbs spindle checkpoint signaling, which may contribute to the antiproliferative effects of the compound.
Asunto(s)
Flavonoides/farmacología , Mitosis/efectos de los fármacos , Huso Acromático/metabolismo , Aurora Quinasa B , Aurora Quinasas , Línea Celular Tumoral , Proteínas Cromosómicas no Histona/metabolismo , Activación Enzimática , Flavonoles , Humanos , Cinetocoros/efectos de los fármacos , Cinetocoros/fisiología , Proteínas de Microfilamentos/metabolismo , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Huso Acromático/efectos de los fármacosRESUMEN
Cenp-F (mitosin) is a large coiled-coil protein whose function has remained obscure since its identification a decade ago. It has been suggested that the protein plays a role in the kinetochore-mediated mitotic functions but until recently there was little evidence to support this postulation. Recent results from five laboratories have given insights on how Cenp-F may participate in the regulation of cell division. In this mini-review, we will summarize the current data regarding the mitotic tasks of Cenp-F as well as discuss how it is used as a proliferation marker of malignant cell growth in the clinic. Also, the protein's post-translational modification by farnesylation and potential contribution to cell cycle effects of farnesyl transferase inhibitors will be addressed.
Asunto(s)
Proteínas Cromosómicas no Histona/fisiología , Proteínas de Microfilamentos/fisiología , Mitosis , Transferasas Alquil y Aril/metabolismo , Animales , Biomarcadores/metabolismo , Proliferación Celular , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , Humanos , Cinetocoros/metabolismo , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Modelos Genéticos , Prenilación de ProteínaRESUMEN
DNA copy number amplification at the chromosomal region of 17q is frequent in gastric cancer. Recently 17q21 was identified as the critical region for the amplicon formation because this region harbors the ERBB2 oncogene and several other targets, such as TOP2A and DARPP32. In our study, we characterized the amplification (52 cases) and expression (29 cases) levels of ERBB2, TOP2A and DARPP32 in gastric cancer samples. These 3 genes were concomitantly amplified in 17% of the intestinal type of gastric adenocarcinoma. However, the expression levels were independent, showing overexpression of DARPP32 (48%), TOP2A (17%) and ERBB2 (3%) studied by quantitative real-time PCR. The most frequently overexpressed gene, DARPP32, exhibited strong protein overexpression in 45% (30/66) of the cases in immunohistochemical study of gastric cancer tumor tissue array. Additional studies are required to thoroughly understand the biological significance of these genes in gastric cancer.
Asunto(s)
ADN-Topoisomerasas de Tipo II/genética , Neoplasias Gastrointestinales/genética , Regulación Neoplásica de la Expresión Génica , Fosfoproteínas/genética , Receptor ErbB-2/genética , Adenocarcinoma/genética , Antígenos de Neoplasias/genética , Estudios de Casos y Controles , Cromosomas Humanos Par 17/genética , ADN de Neoplasias/genética , Proteínas de Unión al ADN , Fosfoproteína 32 Regulada por Dopamina y AMPc , Mucosa Gástrica/metabolismo , Amplificación de Genes , Dosificación de Gen , Humanos , Hibridación Fluorescente in Situ , Proteínas del Tejido Nervioso/genética , Proteínas de Unión a Poli-ADP-Ribosa , ARN Mensajero/genética , ARN Neoplásico/genética , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
DNA copy number gains and amplifications at 17q are frequent in gastriccancer, yet systematic analyses of the 17q amplicon have not been performed. In this study, we carried out a comprehensive analysis of copy number and expression levels of 636 chromosome 17-specific genes in gastric cancer by using a custom-made chromosome 17-specific cDNA microarray. Analysis of DNA copy number changes by comparative genomic hybridization on cDNA microarray revealed increased copy numbers of 11 known genes (ERBB2, TOP2A, GRB7, ACLY, PIP5K2B, MPRL45, MKP-L, LHX1, MLN51, MLN64, and RPL27) and seven expressed sequence tags (ESTs) that mapped to 17q12-q21 region. To investigate the genes transcribed at the 17q, we performed gene expression analyses on an identical cDNA microarray. Our expression analysis showed overexpression of 8 genes (ERBB2, TOP2A, GRB2, AOC3, AP2B1, KRT14, JUP, and ITGA3) and two ESTs. Of the commonly amplified transcripts, an uncharacterized EST AA552509 and the TOP2A gene were most frequently overexpressed in 82% of the samples. Additional studies will be initiated to understand the possible biological and clinical significance of these genes in gastric cancer development and progression.
