Signs of positive selection of somatic mutations in human cancers detected by EST sequence analysis

BACKGROUND: Carcinogenesis typically involves multiple somatic mutations in caretaker (DNA repair) and gatekeeper (tumor suppressors and oncogenes) genes. Analysis of mutation spectra of the tumor suppressor that is most commonly mutated in human cancers, p53, unexpectedly suggested that somatic evolution of the p53 gene during tumorigenesis is dominated by positive selection for gain of function. This conclusion is supported by accumulating experimental evidence of evolution of new functions of p53 in tumors. These findings prompted a genome-wide analysis of possible positive selection during tumor evolution. METHODS: A comprehensive analysis of probable somatic mutations in the sequences of Expressed Sequence Tags (ESTs) from malignant tumors and normal tissues was performed in order to access the prevalence of positive selection in cancer evolution. For each EST, the numbers of synonymous and non-synonymous substitutions were calculated. In order to identify genes with a signature of positive selection in cancers, these numbers were compared to: i) expected numbers and ii) the numbers for the respective genes in the ESTs from normal tissues. RESULTS: We identified 112 genes with a signature of positive selection in cancers, i.e., a significantly elevated ratio of non-synonymous to synonymous substitutions, in tumors as compared to 37 such genes in an approximately equal-sized EST collection from normal tissues. A substantial fraction of the tumor-specific positive-selection candidates have experimentally demonstrated or strongly predicted links to cancer. CONCLUSION: The results of EST analysis should be interpreted with extreme caution given the noise introduced by sequencing errors and undetected polymorphisms. Furthermore, an inherent limitation of EST analysis is that multiple mutations amenable to statistical analysis can be detected only in relatively highly expressed genes. Nevertheless, the present results suggest that positive selection might affect a substantial number of genes during tumorigenic somatic evolution

Structural basis for DNA damage-induced phosphoregulation of MDM2 RING domain

Phosphorylation of MDM2 by ATM upon DNA damage is an important mechanism for deregulating MDM2, thereby leading to p53 activation. ATM phosphorylates multiple residues near the RING domain of MDM2, but the underlying molecular basis for deregulation remains elusive. Here we show that Ser429 phosphorylation selectively enhances the ubiquitin ligase activity of MDM2 homodimer but not MDM2-MDMX heterodimer. A crystal structure of phospho-Ser429 (pS429)-MDM2 bound to E2–ubiquitin reveals a unique 310-helical feature present in MDM2 homodimer that allows pS429 to stabilize the closed E2–ubiquitin conformation and thereby enhancing ubiquitin transfer. In cells Ser429 phosphorylation increases MDM2 autoubiquitination and degradation upon DNA damage, whereas S429A substitution protects MDM2 from auto-degradation. Our results demonstrate that Ser429 phosphorylation serves as a switch to boost the activity of MDM2 homodimer and promote its self-destruction to enable rapid p53 stabilization and resolve a long-standing controversy surrounding MDM2 auto-degradation in response to DNA damage

14-3-3ζ Interacts with Stat3 and Regulates Its Constitutive Activation in Multiple Myeloma Cells

The 14-3-3 proteins are a family of regulatory signaling molecules that interact with other proteins in a phosphorylation-dependent manner and function as adapter or scaffold proteins in signal transduction pathways. One family member, 14-3-3ζ, is believed to function in cell signaling, cycle control, and apoptotic death. A systematic proteomic analysis done in our laboratory has identified signal transducers and activators of transcription 3 (Stat3) as a novel 14-3-3ζ interacting protein. Following our initial finding, in this study, we provide evidence that 14-3-3ζ interacts physically with Stat3. We further demonstrate that phosphorylation of Stat3 at Ser727 is vital for 14-3-3ζ interaction and mutation of Ser727 to Alanine abolished 14-3-3ζ/Stat3 association. Inhibition of 14-3-3ζ protein expression in U266 cells inhibited Stat3 Ser727 phosphorylation and nuclear translocation, and decreased both Stat3 DNA binding and transcriptional activity. Moreover, 14-3-3ζ is involved in the regulation of protein kinase C (PKC) activity and 14-3-3ζ binding to Stat3 protects Ser727 dephosphorylation from protein phosphatase 2A (PP2A). Taken together, our findings support the model that multiple signaling events impinge on Stat3 and that 14-3-3ζ serves as an essential coordinator for different pathways to regulate Stat3 activation and function in MM cells

Should all adjunctive corticosteroid therapy be avoided in the management of hemodynamically stabile Staphylococcus aureus bacteremia?

The purpose of this study was to examine the prognostic impact of corticosteroids in hemodynamically stabile Staphylococcus aureus bacteremia (SAB). There were 361 hemodynamically stabile methicillin-sensitive SAB patients with prospective follow-up and grouping according to time-point, dose and indication for corticosteroid therapy. To enable analyses without external interfering corticosteroid therapy all patients with corticosteroid therapy equivalent to prednisone > 10 mg/day for >= 1 month prior to positive blood culture results were excluded. Twenty-five percent (92) of patients received corticosteroid therapy of which 11 % (40) had therapy initiated within 1 week (early initiation) and 9 % (31) had therapy initiated 2-4 weeks after (delayed initiation) positive blood culture. Twenty-one patients (6 %) had corticosteroid initiated after 4 weeks and were not included in the analyses. A total of 55 % (51/92) received a weekly prednisone dose > 100 mg. Patients with early initiated corticosteroid therapy had higher mortality compared to patients treated without corticosteroid therapy at 28 days (20 % vs. 7 %) (OR, 3.11; 95% CI, 1.27-7.65; p = 100 mg/week the negative prognostic impact on 28-day mortality was accentuated (HR 4.8, p = 0.001). Corticosteroid therapy initiation after 1 week of positive blood cultures had no independent prognostic impact. Early initiation of corticosteroid therapy may be associate to increased mortality in hemodynamically stabile SAB.Peer reviewe

Rh D foeto-maternal alloimmunization prophylaxis with anti-D immunoglobulins reviewed in the era of foetal RHD genotyping.

In Belgium, prevention of anti-D immunization is currently based on systematic postnatal prophylaxis associated with targeted antenatal injection in high-risk situations of foeto-maternal haemorrhage.The failures of prevention are mainly due to the non-respect of established guidelines for RhlG prophylaxis, and to spontaneous undetected foetal-maternal haemorrhages without any obvious cause during the third trimester of pregnancy. In order to reduce the rate of residual post-pregnancy anti-D immunization, several countries decided to associate the classical prophylaxis to a routine antenatal anti-D prophylaxis (RAADP) during the 28th or 29th week of gestation. Since a few years, the foetal RHD genotyping in maternal plasma enables us to limit the antenatal prophylaxis only to those D- women carrying a D+ foetus. This paper deals with: the advantages of an antenatal prevention in the light of non-invasive foetal RHD genotyping, the rules rendering prevention protocols efficient whatever the algorithm applied, and the recommended immuno-haematology follow-up of women who received RhlG

Roles of brca2 (fancd1) in Oocyte Nuclear Architecture, Gametogenesis, Gonad Tumors, and Genome Stability in Zebrafish

Mild mutations in BRCA2 (FANCD1) cause Fanconi anemia (FA) when homozygous, while severe mutations cause common cancers including breast, ovarian, and prostate cancers when heterozygous. Here we report a zebrafish brca2 insertional mutant that shares phenotypes with human patients and identifies a novel brca2 function in oogenesis. Experiments showed that mutant embryos and mutant cells in culture experienced genome instability, as do cells in FA patients. In wild-type zebrafish, meiotic cells expressed brca2; and, unexpectedly, transcripts in oocytes localized asymmetrically to the animal pole. In juvenile brca2 mutants, oocytes failed to progress through meiosis, leading to female-to-male sex reversal. Adult mutants became sterile males due to the meiotic arrest of spermatocytes, which then died by apoptosis, followed by neoplastic proliferation of gonad somatic cells that was similar to neoplasia observed in ageing dead end (dnd)-knockdown males, which lack germ cells. The construction of animals doubly mutant for brca2 and the apoptotic gene tp53 (p53) rescued brca2-dependent sex reversal. Double mutants developed oocytes and became sterile females that produced only aberrant embryos and showed elevated risk for invasive ovarian tumors. Oocytes in double-mutant females showed normal localization of brca2 and pou5f1 transcripts to the animal pole and vasa transcripts to the vegetal pole, but had a polarized rather than symmetrical nucleus with the distribution of nucleoli and chromosomes to opposite nuclear poles; this result revealed a novel role for Brca2 in establishing or maintaining oocyte nuclear architecture. Mutating tp53 did not rescue the infertility phenotype in brca2 mutant males, suggesting that brca2 plays an essential role in zebrafish spermatogenesis. Overall, this work verified zebrafish as a model for the role of Brca2 in human disease and uncovered a novel function of Brca2 in vertebrate oocyte nuclear architecture