Targeted point mutations of p53 lead to dominant-negative inhibition of wild-type p53 function.
ABSTRACT The p53 tumor suppressor gene is the most frequently mutated gene in human cancers, and germ-line p53 mutations cause a familial predisposition for cancer. Germ-line or sporadic p53 mutations are usually missense and typically affect the central DNA-binding domain of the protein. Because p53 functions as a tetrameric transcription factor, mutant p53 is thought to inhibit the function of wild-type p53 protein. Here, we studied the possible dominant-negative inhibition of wild-type p53 protein by two different, frequently occurring point mutations. The R270H and P275S mutations were targeted into the genome of mouse embryonic stem cells to allow the analysis of the effects of the mutant proteins expressed in normal cells at single-copy levels. In embryonic stem cells, the presence of a heterozygous point-mutated allele resulted in delayed transcriptional activation of several p53 downstream target genes on exposure to gamma irradiation. Doxorubicin-induced apoptosis was severely affected in the mutant embryonic stem cells compared with wild-type cells. Heterozygous mutant thymocytes had a severe defect in p53-dependent apoptotic pathways after treatment with gamma irradiation or doxorubicin, whereas p53-independent apoptotic pathways were intact. Together these data demonstrate that physiological expression of point-mutated p53 can strongly limit overall cellular p53 function, supporting the dominant-negative action of such mutants. Also, cells heterozygous for such mutations may be compromised in terms of tumor suppression and response to chemotherapeutic agents.
SourceAvailable from: Lorenzo Galluzzi[Show abstract] [Hide abstract]
ABSTRACT: Autophagy plays a key role in the maintenance of cellular homeostasis. In healthy cells, such a homeostatic activity constitutes a robust barrier against malignant transformation. Accordingly, many oncoproteins inhibit, and several oncosuppressor proteins promote, autophagy. Moreover, autophagy is required for optimal anticancer immunosurveillance. In neoplastic cells, however, autophagic responses constitute a means to cope with intracellular and environmental stress, thus favoring tumor progression. This implies that at least in some cases, oncogenesis proceeds along with a temporary inhibition of autophagy or a gain of molecular functions that antagonize its oncosuppressive activity. Here, we discuss the differential impact of autophagy on distinct phases of tumorigenesis and the implications of this concept for the use of autophagy modulators in cancer therapy. © 2015 The Authors.The EMBO Journal 02/2015; 34(7). DOI:10.15252/embj.201490784 · 10.75 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: Inherited mutations in DNA repair genes are major contributors to familial cancer syndromes. Most pathogenic mutations introduce premature stop codons resulting in the loss of protein function. Missense variants of uncertain pathogenic significance are also often observed. In an attempt to understand the physiological attributes of this category of variants, this thesis analysed two recurrent coding variants using a multi-disciplinary approach. The first variant, PMS2 c.2002A>G, was identified in the Inuit population of Quebec and early onset cancers co-segregate with homozygous status. The second, BRCA2 c.6853A>G, appears to be restricted to the Ashkenazi Jewish population and is frequently recorded in the database of Breast Cancer Information Core (BIC, http://research.nhgri.nih.gov/bic/). Both variants not only cause the substitution of an isoleucine to a valine in the proteins, but also interfere with RNA splicing. The variant PMS2 c.2002 A>G creates a novel 5' splicing site which competes with the authentic one, resulting in biased gene expression. The majority of the transcripts are aberrant ones with a five-base pair deletion and a minority of intact transcripts are still produced. The intact transcripts are translated into a full-length proteins which contribute to the milder phenotype associated with this variant, when compared to phenotypes caused by homozygous truncating PMS2 mutations. The variant BRCA2 c.6853A>G disrupts a splicing enhancer promoting the skipping of exon 12, an in-frame exon that does not code for any known functional domains. No detectable phenotype is caused by this variant. The work described in this thesis generated significant medical insights into hereditary syndrome. First, we demonstrated that the inclusion of expression analysis in gene test can improve the accuracy of predicting the functional significance of a candidate variant. Second, manipulating gene expression may be a potential avenue of preventing recessive diseases. Finally, phenotypic variation may be caused by multiple factors, so predicting disease risk solely based on DNA sequence should be practised cautiously with full awareness of the limitation.04/2014, Degree: PhD, Supervisor: William D Foulkes
Carcinogenesis 01/2003; 24(1):113-119. DOI:10.1093/carcin/24.1.113 · 5.27 Impact Factor