RESUMEN
Mechanisms regulating the transition of mammary epithelial cells (MECs) to mammary stem cells (MaSCs) and to tumor-initiating cells (TICs) have not been entirely elucidated. The p53 family member, p63, is critical for mammary gland development and contains transactivation domain isoforms, which have tumor-suppressive activities, and the ΔN isoforms, which act as oncogenes. In the clinic, p63 is often used as a diagnostic marker, and further analysis of the function of TAp63 in the mammary gland is critical for improved diagnosis and patient care. Loss of TAp63 in mice leads to the formation of aggressive metastatic mammary adenocarcinoma at 9-16 months of age. Here we show that TAp63 is crucial for the transition of mammary cancer cells to TICs. When TAp63 is lost, MECs express embryonic and MaSC signatures and activate the Hippo pathway. These data indicate a crucial role for TAp63 in mammary TICs and provide a mechanism for its role as a tumor- and metastasis-suppressor in breast cancer.
Asunto(s)
Neoplasias de la Mama/patología , Carcinogénesis , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal , Transactivadores/metabolismo , Proteínas Quinasas Activadas por AMP , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Recuento de Células , Línea Celular Tumoral , Polaridad Celular , Transformación Celular Neoplásica , Transición Epitelial-Mesenquimal , Femenino , Técnicas de Silenciamiento del Gen , Vía de Señalización Hippo , Humanos , Hiperplasia , Glándulas Mamarias Animales/patología , Glándulas Mamarias Animales/fisiopatología , Ratones , Células Madre Neoplásicas/patología , Fosfoproteínas/deficiencia , Fosfoproteínas/genética , Proteínas Serina-Treonina Quinasas/genética , Regeneración , Transactivadores/deficiencia , Transactivadores/genética , Transcripción GenéticaRESUMEN
We estimated the relative contributions of known ataxia genes (SCA1, 2, 3, 6, 7 and X25) in the patient population sent to our DNA diagnostic laboratory for diagnostic testing. Approximately 28% of these patients had an abnormal triplet repeat expansion in one of these ataxia genes (3% for SCA1, 8% for SCA2, 11% for SCA3/MJD, 2% for SCA6, 3% for SCA7, and 1.5% for X25). The lack of abnormal repeat expansions in the majority of ataxis patients tested suggests that the molecular defects associated with most ataxia cases are currently undetermined and that this population includes both familial and sporadic cases. In contrast, of the patients submitted for genetic testing for Friedrich's ataxia (FRDA), 44% (69/157) showed at least one expansion in the X25 gene, indicating that FRDA accounts for a significant proportion of the recessively inherited ataxias and appears to have a high rate of accurate clinical diagnosis. On the basis of our DNA studies, we propose a comprehensive and efficient approach for molecular analysis of ataxia patients.
Asunto(s)
Análisis Mutacional de ADN/métodos , Pruebas Genéticas/métodos , Repeticiones de Minisatélite , Degeneraciones Espinocerebelosas/genética , Genes Dominantes , Genes Recesivos , Humanos , Degeneraciones Espinocerebelosas/clasificación , Degeneraciones Espinocerebelosas/diagnósticoRESUMEN
To examine the relationship between replication timing and differential gene transcription in tissue-specific and imprinted settings we have studied the replication timing properties of the human Prader-Willi syndrome (PWS) region on human chromosome 15q11-13. Interphase fluorescence in situ hybridization with an overlapping series of cosmid clones was used to map a PWS replication timing domain to a 500- to 650-kb region that includes the SNRPN gene. This PWS domain replicates late in lymphocytes but predominantly early in neuroblasts, with replication asynchrony observed in both tissues, and appears to colocalize with a genetically imprinted transcription domain showing prominent expression in the brain. A 5- to 30-kb deletion in the 5' region of SNRPN results in the loss of late replication control of this domain in lymphocytes when the deleted chromosome is inherited paternally. This potential allele-specific replication timing control region also appears to colocalize with a putative imprinting control region that has been shown previously to abolish the expression of three imprinted transcripts in this same region.
Asunto(s)
Replicación del ADN , Impresión Genómica , Síndrome de Prader-Willi/genética , Ribonucleoproteínas Nucleares Pequeñas/genética , Células Cultivadas , Cromosomas Humanos Par 15 , Femenino , Humanos , Linfocitos/metabolismo , Masculino , Neuronas/metabolismo , Eliminación de Secuencia , Transcripción Genética , Células Tumorales CultivadasRESUMEN
The Smith-Magenis syndrome (SMS) appears to be a contiguous-gene-deletion syndrome associated with a proximal deletion of the short arm of chromosome 17 in band p11.2. The spectrum of clinical findings includes short stature, brachydactyly, developmental delay, dysmorphic features, sleep disturbances, and behavioral problems. The complex phenotypic features suggest deletion of several contiguous genes. However, to date, no protein-encoding gene has been mapped to the SMS critical region. Recently, the Drosophila melanogaster flightless-I gene, fliI, and the homologous human cDNA have been isolated. Mutations in fliI result in loss of flight ability and, when severe, cause lethality due to incomplete cellularization with subsequent abnormal gastrulation. Here, we demonstrate that the human homologue (FLI) maps within the SMS critical region. Genomic cosmids were used as probes for FISH, which localized this gene to the 17p11.2 region. Somatic-cell hybrid-panel mapping further localized this gene to the SMS critical region. Southern blot analysis of somatic-cell hybrids and/or FISH analysis of lymphoblastoid cell lines from 12 SMS patients demonstrates the deletion of one copy of FLI in all SMS patients analyzed.
Asunto(s)
Cromosomas Humanos Par 17 , Drosophila melanogaster/genética , Enanismo/genética , Gelsolina , Discapacidad Intelectual/genética , Proteínas/genética , Receptores Citoplasmáticos y Nucleares , Animales , Línea Celular Transformada , Mapeo Cromosómico , Cósmidos , Cricetinae , Eliminación de Gen , Genes , Genes de Insecto , Humanos , Células Híbridas , Hibridación Fluorescente in Situ , Linfocitos/ultraestructura , Ratones , Proteínas de Microfilamentos , Morfogénesis/genética , Síndrome , TransactivadoresRESUMEN
A 4-kb HindIII fragment that supported the efficient autonomous replication of plasmid vector pDY-, a replication-defective construct based on Epstein-Barr virus sequences, in human K562 cells was rescued from amplified double-minute chromosomes containing the murine adenosine deaminase locus. Polymerase chain reaction assays of size-fractionated nascent strands demonstrated that replication initiation occurred within the same 1- to 2-kb region of this fragment in autonomously replicating plasmids containing the sequence in either orientation, in double-minute chromosomes, and in the single-copy locus at its normal chromosomal location. The complete sequence of this fragment was determined; it contains a 248-bp polypurine tract and consensus binding site sequences for several putative transcription and replication factors.
Asunto(s)
Adenosina Desaminasa/genética , Replicación del ADN , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Sitios de Unión , Línea Celular , Clonación Molecular , Secuencia de Consenso , ADN , Genoma , Humanos , Ratones , Datos de Secuencia Molecular , Plásmidos , Reacción en Cadena de la Polimerasa , Homología de Secuencia de Ácido Nucleico , Transcripción GenéticaRESUMEN
The zeste locus plays a central role in transvection phenomena, where the synaptic pairing of chromosomes carrying genes with which zeste interacts influences the expression of these genes. To explore the possible functions of the zeste gene product in this process, we have determined the DNA sequences both of a fragment of Drosophila genomic DNA capable of rescuing mutant zeste phenotypes, and of a near full-length cDNA clone derived from the 2.4-kb zeste mRNA. These data show that the zeste gene is interrupted by two small introns, and suggest that the majority of zeste sequences are contained within an intron of another transcriptional unit of opposite polarity. A large region of the predicted zeste product is comprised almost exclusively of glutamine and alanine residues. A domain near the N terminus of this protein, which is sufficient for site-specific DNA binding, is highly charged, as is the C-terminal region of the protein. A breakpoint of the rearrangement In (1)e(bx), which is associated with a za-like phenotype, is found within sequences encoding the zeste product, and would produce a truncated protein. The neomorphic mutation zv77h is correlated with a 300-bp deletion of sequences determining the untranslated 5' leader of the zeste messenger, but may also remove the initiating ATG codon, resulting in a zeste protein with an altered N terminus.
Asunto(s)
Drosophila melanogaster/genética , Meiosis , Complejo Sinaptonémico , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Proteínas de Unión al ADN/genética , Regulación de la Expresión Génica , Genes , Prueba de Complementación Genética , Datos de Secuencia Molecular , MutaciónRESUMEN
Approximately 170 kilobase pairs (kb) of contiguous DNA sequences derived from bands 3A3,4 of the Drosophila melanogaster X chromosome have been isolated by molecular cloning. Sequences required for the wild-type expression of the zeste locus are located within a 6-kb fragment of this chromosomal region, as shown by phenotypic rescue of zeste mutants in P element-mediated germ-line transformation. Expression of zeste is correlated with a 2.2-kb poly(A)+ RNA species transcribed at all postzygotic stages of Drosophila development. Many zeste alleles, including several producing neomorphic phenotypes, are not associated with detectable rearrangements of DNA.