RESUMO
Neuroblastoma (NB) is a common childhood malignant tumor of the neural crest-derived sympathetic nervous system. In NB the frequent loss of heterozygosity (LOH) on chromosome 1p raises the possibility that this region contains tumor-suppressor genes whose inactivation contributes to tumorigenesis. The human homolog of the Drosophila neural fate determination gene CASZ1, a zinc-finger transcription factor, maps to chromosome 1p36.22, a region implicated in NB tumorigenesis. Quantitative real-time PCR analysis showed that low-CASZ1 expression is significantly correlated with increased age (≥18 months), Children's Oncology Group high-risk classification, 1p LOH and MYCN amplification (all P<0.0002) and decreased survival probability (P=0.0009). CASZ1 was more highly expressed in NB with a differentiated histopathology (P<0.0001). Retinoids and epigenetic modification agents associated with regulation of differentiation induced CASZ1 expression. Expression profiling analysis revealed that CASZ1 regulates the expression of genes involved in regulation of cell growth and developmental processes. Specific restoration of CASZ1 in NB cells induced cell differentiation, enhanced cell adhesion, inhibited migration and suppressed tumorigenicity. These data are consistent with CASZ1 being a critical modulator of neural cell development, and that somatically acquired disruption of normal CASZ1 expression contributes to the malignant phenotype of human NB.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Genes Supressores de Tumor , Neuroblastoma/genética , Fatores de Transcrição/metabolismo , Adesão Celular , Diferenciação Celular , Linhagem Celular Tumoral , Movimento Celular , Reprogramação Celular , Cromossomos Humanos Par 1 , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Lactente , Estimativa de Kaplan-Meier , Proteína Proto-Oncogênica N-Myc , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Retinoides/farmacologia , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genéticaRESUMO
The polymerization of alpha- and beta-tubulin into microtubules results in a complex network of microfibrils that have important structural and functional roles in all eukaryotic cells. In addition, microtubules can interact with a diverse family of polypeptides which are believed to directly promote the assembly of microtubules and to modulate their functional activity. We have demonstrated that the c-Myc oncoprotein interacts in vivo and in vitro with alpha-tubulin and with polymerized microtubules and have defined the binding site to the N-terminal region within the transactivation domain of c-Myc. In addition, we have shown that c-Myc colocalizes with microtubules and remains tightly bound to the microtubule network after detergent extraction of intact cells. These findings suggest a potential role for Myc-tubulin interaction in vivo.
Assuntos
Microtúbulos/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Tubulina (Proteína)/metabolismo , Sequência de Bases , Sítios de Ligação , Compartimento Celular , Análise Mutacional de DNA , Imunofluorescência , Humanos , Dados de Sequência Molecular , Ligação Proteica , Deleção de Sequência , Células Tumorais CultivadasRESUMO
We have analyzed the CTF4 (CHL15) gene, earlier identified in two screens for yeast mutants with increased rates of mitotic loss of chromosome III and artificial circular and linear chromosomes. Analysis of the segregation properties of circular minichromosomes and chromosome fragments indicated that sister chromatid loss (1:0 segregation) is the predominant mode of chromosome destabilization in ctf4 mutants, though nondisjunction events (2:0 segregation) also occur at an increased rate. Both inter- and intrachromosomal mitotic recombination levels are elevated in ctf4 mutants, whereas spontaneous mutation to canavanine resistance was not elevated. A genomic clone of CTF4 was isolated and used to map its physical and genetic positions on chromosome XVI. Nucleotide sequence analysis of CTF4 revealed a 2.8-kb open reading frame with a 105-kDa predicted protein sequence. The CTF4 DNA sequence is identical to that of POB1, characterized as a gene encoding a protein that associates in vitro with DNA polymerase alpha. At the N-terminal region of the protein sequence, zinc finger motifs which define potential DNA-binding domains were found. The C-terminal region of the predicted protein displayed similarity to sequences of regulatory proteins known as the helix-loop-helix proteins. Data on the effects of a frameshift mutation suggest that the helix-loop-helix domain is essential for CTF4 function. Analysis of sequences upstream of the CTF4 open reading frame revealed the presence of a hexamer element, ACGCGT, a sequence associated with many DNA metabolism genes in budding yeasts. Disruption of the coding sequence of CTF4 did not result in inviability, indicating that the CTF4 gene is nonessential for mitotic cell division. However, ctf4 mutants exhibit an accumulation of large budded cells with the nucleus in the neck. ctf4 rad52 double mutants grew very slowly and produced extremely high levels (50%) of inviable cell division products compared with either single mutant alone, which is consistent with a role for CTF4 in DNA metabolism.
Assuntos
DNA Fúngico/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas Fúngicas/genética , Genes Fúngicos , Proteínas , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Sequência de Bases , Mapeamento Cromossômico , Clonagem Molecular , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Teste de Complementação Genética , Cinética , Dados de Sequência Molecular , Mutação , Mapeamento por Restrição , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiaeRESUMO
A genomic fragment containing the K51 gene previously isolated from a rat genomic library by hybridization with the v-mos probe in nonstringent conditions [Chumakov et al., Dokl. Akad. Nauk SSSR 290 (1986) 1252-1254], resembles a human keratin type-I-encoding gene [Shvets et al., Mol. Biol. 24 (1990) 663-677]. This genomic clone, K51, has been used as a probe to search for related human genes. A recombinant clone, HK51, with a 1.5-kb insert, was isolated from a human embryonic skin cDNA library, and its nucleotide (nt) sequence was determined. Analysis has shown that the cloned cDNA encodes human keratin 10 (Ker10). All presently known nt sequences of the human Ker10-encoding gene (Ker10) are not identical. Differences are concentrated in the 5'-end of the first exon and in the middle of the seventh exon within repeats. In spite of structural rearrangements in two of eight exons, the reading frame and position of the stop codon are preserved. The genetic rearrangements cause changes in hydrophobicity profiles of the N and C termini of Ker10. It was also noticed that insertion of one nt leads to the formation of an unusual 3'-end of the transcript.