Biomedical subjects

Arnold J Levine

Publications and source records attributed to Arnold J Levine.

At least 19 recordsLinked to original sources

PubMed2006

Comparison of avian and human influenza A viruses reveals a mutational bias on the viral genomes.

In the last few years, the genomic sequence data for thousands of influenza A virus strains, including the 1918 pandemic strain, and hundreds of isolates of the avian influenza virus H5N1, which is causing an increasing number of human fatalities, have become publicly available. This large quantity of sequence data allows us to do comparative genomics with the human and avian versions of the virus. We find that the nucleotide compositions of influenza A viruses infecting the two hosts are sufficiently different that we can determine the host at almost 100% accuracy. This assignment works at the segment level, which allows us to construct the reassortment history of individual segments within each strain. We suggest that the different nucleotide compositions can be explained by a host-dependent mutation bias. To support this idea, we estimate the fixation rates for the different polymerase segments and the ratios of synonymous to nonsynonymous changes. Additionally, we provide evidence supporting the hypothesis that the H1N1 influenza virus entered the human population just prior to the 1918 outbreak, with an earliest bound of 1910.

Amino Acid Sequence↗

PubMed2006

MDM2 SNP309 accelerates colorectal tumour formation in women.

Recent studies have shown that the G-allele of MDM2 SNP309 (T/G) in the p53 tumour suppressor pathway can accelerate tumorigenesis and alter the risk of various cancers in women and not in men. In this report, data are presented from two independent groups of patients that suggest that the G-allele of SNP309 accelerates colorectal tumour formation only in women, and that lend further support to the model that primarily female-specific hormones, such as oestrogen, could either directly or indirectly allow for the G-allele of SNP309 to accelerate tumour formation in women.

Adult↗

PubMed2006

MDM2 SNP309 accelerates tumor formation in a gender-specific and hormone-dependent manner.

The importance of the p53 stress response pathway in the suppression of tumor formation is well documented. In a previous report, a single nucleotide polymorphism (SNP309 T/G) was found in the promoter of the MDM2 gene resulting in higher levels of MDM2 RNA and protein and, consequently, in the attenuation of the p53 pathway both in vitro and in vivo. As the SNP309 locus is found in a region of the MDM2 promoter, which is regulated by hormonal signaling pathways, and the G-allele of SNP309 increases the affinity of a well-described cotranscriptional activator of nuclear hormone receptors (i.e., Sp1), the hypothesis that the SNP309 locus could alter the effects of hormones on tumorigenesis was tested in vivo in humans. Data obtained from patients with three different sporadic cancers, from four independent case studies, support this hypothesis, providing an example for the genetic basis of gender differences in cancer and showing that the genotype at a specific locus can affect how hormones, like estrogen, affect tumorigenesis in humans.

Adolescent↗

PubMed2006

The regulation of exosome secretion: a novel function of the p53 protein.

The p53 protein responds to stress signals by regulating the transcription of a variety of genes. Some of these genes encode secreted proteins that may be involved in the communication between adjacent cells. In this study, a proteomics approach was employed to identify proteins secreted by cells in a p53-dependent manner after DNA damage. In addition to the known transcriptional targets of p53, a set of proteins encoded by genes that are not transcriptional targets of p53 were found to increase in the culture medium after p53 activation. These proteins exit the cell via small, secreted vesicles called exosomes and exosome production by cells was found to be regulated by the p53 response. A p53-regulated gene product, TSAP6, was shown to enhance exosome production in cells undergoing a p53 response to stress. Thus, the p53 pathway regulates the production of exosomes into the medium and these vesicles can communicate with adjacent cells and even cells of the immune system.

Alleles↗

PubMed2006

Comparison of the human and worm p53 structures suggests a way for enhancing stability.

Maintaining the native conformation is essential for the proper function of tumor suppressor protein p53. However, p53 is a low-stability protein that can easily lose its function upon structural perturbations such as those resulting from missense mutations, leading to the development of cancer. Therefore, it is important to develop strategies to design stable p53 which still maintains its normal function. Here, we compare the stabilities of the human and worm p53 core domains using molecular dynamics simulations. We find that the worm p53 is significantly more stable than the human form. Detailed analysis of the structural fluctuations shows that the stability difference lies in the peripheral structural motifs that contrast in their structural features and flexibility. The most dramatic difference in stability originates from loop L1, from the turn between helix H1 and beta-strand S5, and from the turn that connects beta-strands S7 and S8. Structural analysis shows significant differences for these motifs between the two proteins. Loop L1 lacks secondary structure, and the turns between helix H1 and strand S5 and between strands S7 and S8 are much longer in the human form p53. On the basis of these differences, we designed a mutant by shortening the turn between strands S7 and S8 to enhance the stability. Surprisingly, this mutant was very stable when probed by molecular dynamics simulations. In addition, the stabilization was not localized in the turn region. Loop L1 was also significantly stabilized. Our results show that stabilizing peripheral structural motifs can greatly enhance the stability of the p53 core domain and therefore is likely to be a viable alternative in the development of stable p53. In addition, loop- or turn-related mutants with different stabilities may also be used to probe the relationship between function, a particular structural motif, and its flexibility.

Animals↗

PubMed2006

Coordination and communication between the p53 and IGF-1-AKT-TOR signal transduction pathways.

Over the past 10 years the signal transduction networks for p53, IGF-1-AKT, and TOR pathways have been assembled in worms, flies, and mammals, and their functions elucidated. In the past 1-2 years a number of genes and their proteins have been identified that permit extensive communication and coordination between these pathways. These three pathways are involved in sensing and integrating signals arising from nutrient and growth factor availability, signals from sensory and sexual organs, and intrinsic and extrinsic stress signals. In turn these pathways regulate cell growth, proliferation, and death. These networks are central to our understanding of a variety of physiological and pathological conditions, including cancer, diabetes, and longevity.

Animals↗

PubMed2006

A robust meta-classification strategy for cancer detection from MS data.

We propose a novel method for phenotype identification involving a stringent noise analysis and filtering procedure followed by combining the results of several machine learning tools to produce a robust predictor. We illustrate our method on SELDI-TOF MS prostate cancer data (http://home.ccr.cancer.gov/ncifdaproteomics/ppatterns.asp). Our method identified 11 proteomic biomarkers and gave significantly improved predictions over previous analyses with these data. We were able to distinguish cancer from non-cancer cases with a sensitivity of 90.31% and a specificity of 98.81%. The proposed method can be generalized to multi-phenotype prediction and other types of data (e.g., microarray data).

Biomarkers, Tumor↗

PubMed2006

Beyond PTEN mutations: the PI3K pathway as an integrator of multiple inputs during tumorigenesis.

The tumour-suppressor phosphatase with tensin homology (PTEN) is the most important negative regulator of the cell-survival signalling pathway initiated by phosphatidylinositol 3-kinase (PI3K). Although PTEN is mutated or deleted in many tumours, deregulation of the PI3K-PTEN network also occurs through other mechanisms. Crosstalk between the PI3K pathways and other tumorigenic signalling pathways, such as those that involve Ras, p53, TOR (target of rapamycin) or DJ1, can contribute to this deregulation. How does the PI3K pathway integrate signals from numerous sources, and how can this information be used in the rational design of cancer therapies?

Animals↗

PubMed2005

Detection of functional single-nucleotide polymorphisms that affect apoptosis.

Human EBV-transformed B lymphocyte cell lines (LCLs) were used to measure the apoptotic response of individuals to gamma radiation. The responses form a normal distribution around a median of 35.5% apoptosis with a range of 12-58%. This heterogeneous response has a genetic basis. LCLs from Caucasian donors and African American donors form distinct distributions of apoptotic response; all of the 11 LCLs comprising the lowest responding group (exhibiting between 12-20% apoptosis) are from Caucasian donors. The assay is capable of detecting significant effects of SNPs in two genes, MDM2 and AKT1, whose products are involved in controlling the p53 pathway and cellular response to DNA damage, suggesting that these data and this assay can be used to identify novel SNPs in other genes whose products impact the cellular response to radiation. Finally, the LCLs in the lowest apoptotic response group have the highest concentration of AKT1 protein and all harbor a haplotype in AKT1 that is present in Caucasians but absent in African Americans.

Apoptosis↗

PubMed2005

A plausible model for the digital response of p53 to DNA damage.

Recent observations show that the single-cell response of p53 to ionizing radiation (IR) is "digital" in that it is the number of oscillations rather than the amplitude of p53 that shows dependence on the radiation dose. We present a model of this phenomenon. In our model, double-strand break (DSB) sites induced by IR interact with a limiting pool of DNA repair proteins, forming DSB-protein complexes at DNA damage foci. The persisting complexes are sensed by ataxia telangiectasia mutated (ATM), a protein kinase that activates p53 once it is phosphorylated by DNA damage. The ATM-sensing module switches on or off the downstream p53 oscillator, consisting of a feedback loop formed by p53 and its negative regulator, Mdm2. In agreement with experiments, our simulations show that by assuming stochasticity in the initial number of DSBs and the DNA repair process, p53 and Mdm2 exhibit a coordinated oscillatory dynamics upon IR stimulation in single cells, with a stochastic number of oscillations whose mean increases with IR dose. The damped oscillations previously observed in cell populations can be explained as the aggregate behavior of single cells.

Ataxia Telangiectasia Mutated Proteins↗

PubMed2005

The coordinate regulation of the p53 and mTOR pathways in cells.

Cell growth and proliferation requires an intricate coordination between the stimulatory signals arising from nutrients and growth factors and the inhibitory signals arising from intracellular and extracellular stresses. Alteration of the coordination often causes cancer. In mammals, the mTOR (mammalian target of rapamycin) protein kinase is the central node in nutrient and growth factor signaling, and p53 plays a critical role in sensing genotoxic and other stresses. The results presented here demonstrate that activation of p53 inhibits mTOR activity and regulates its downstream targets, including autophagy, a tumor suppression process. Moreover, the mechanisms by which p53 regulates mTOR involves AMP kinase activation and requires the tuberous sclerosis (TSC) 1/TSC2 complex, both of which respond to energy deprivation in cells. In addition, glucose starvation not only signals to shut down mTOR, but also results in the transient phosphorylation of the p53 protein. Thus, p53 and mTOR signaling machineries can cross-talk and coordinately regulate cell growth, proliferation, and death.

AMP-Activated Protein Kinases↗

PubMed2005

A chromatin-associated and transcriptionally inactive p53-Mdm2 complex occurs in mdm2 SNP309 homozygous cells.

In cancer cells, the function of the tumor suppressor protein p53 is usually blocked. Impairment of the p53 pathway results in tumor cells with endogenous overexpression of Mdm2 via a naturally occurring single nucleotide polymorphism (SNP) in the mdm2 gene at position 309. Here we report that in mdm2 SNP309 cells, inactivation of p53 results in a chromatin-associated Mdm2-p53 complex without clearance of p53 by protein degradation. Nuclear accumulation of p53 protein in mdm2 SNP309 cells results after 6 h of camptothecin, etoposide, or mitomycin C treatment, with the p53 protein phosphorylated at Ser15. Chromatin immunoprecipitation demonstrated p53 and Mdm2 bound to p53 responsive elements. Interestingly, although the p53 protein was able to bind to DNA, quantitative PCR showed compromised transcription of endogenous target genes. Additionally, exogenously introduced p53 was incapable of activating transcription from p53 responsive elements in SNP309 cells, confirming the trans-acting nature of the inhibitor. Inhibition of Mdm2 by siRNA resulted in transcriptional activation of these p53 targets. Our data suggest that overproduction of Mdm2, resulting from a naturally occurring SNP, inhibits chromatin-bound p53 from activating the transcription of its target genes.

Antineoplastic Agents↗

PubMed2005

The p53 pathway: positive and negative feedback loops.

The p53 pathway responds to stresses that can disrupt the fidelity of DNA replication and cell division. A stress signal is transmitted to the p53 protein by post-translational modifications. This results in the activation of the p53 protein as a transcription factor that initiates a program of cell cycle arrest, cellular senescence or apoptosis. The transcriptional network of p53-responsive genes produces proteins that interact with a large number of other signal transduction pathways in the cell and a number of positive and negative autoregulatory feedback loops act upon the p53 response. There are at least seven negative and three positive feedback loops described here, and of these, six act through the MDM-2 protein to regulate p53 activity. The p53 circuit communicates with the Wnt-beta-catenin, IGF-1-AKT, Rb-E2F, p38 MAP kinase, cyclin-cdk, p14/19 ARF pathways and the cyclin G-PP2A, and p73 gene products. There are at least three different ubiquitin ligases that can regulate p53 in an autoregulatory manner: MDM-2, Cop-1 and Pirh-2. The meaning of this redundancy and the relative activity of each of these feedback loops in different cell types or stages of development remains to be elucidated. The interconnections between signal transduction pathways will play a central role in our understanding of cancer.

Amino Acid Sequence↗

PubMed2005

Comparison of the protein-protein interfaces in the p53-DNA crystal structures: towards elucidation of the biological interface.

p53, the tumor suppressor protein, functions as a dimer of dimers. However, how the tetramer binds to the DNA is still an open question. In the crystal structure, three copies of the p53 monomers (containing chains A, B, and C) were crystallized with the DNA-consensus element. Although the structure provides crucial data on the p53-DNA contacts, the active oligomeric state is unclear because the two dimeric (A-B and B-C) interfaces present in the crystal cannot both exist in the tetramer. Here, we address the question of which of these two dimeric interfaces may be more biologically relevant. We analyze the sequence and structural properties of the p53-p53 dimeric interfaces and carry out extensive molecular dynamics simulations of the crystal structures of the human and mouse p53 dimers. We find that the A-B interface residues are more conserved than those of the B-C. Molecular dynamics simulations show that the A-B interface can provide a stable DNA-binding motif in the dimeric state, unlike B-C. Our results indicate that the interface between chains A-B in the p53-DNA complex constitutes a better candidate for a stable biological interface, whereas the B-C interface is more likely to be due to crystal packing. Thus, they have significant implications toward our understanding of DNA binding by p53 as well as p53-mediated interactions with other proteins.

Amino Acid Motifs↗

PubMed2005

In the quest for stable rescuing mutants of p53: computational mutagenesis of flexible loop L1.

p53 is a protein with marginal stability. Its transcriptional functions are often inactivated by single missense mutations, shown to be associated with half of all human cancers. Here, we aim to design stable functional p53 mutants. We target loop L1, one of the most mobile structural motifs in the p53 core domain (p53C). Specifically, we selected Ser116 in the middle of loop L1 and mutated it to 14 other amino acids. All resulting mutants were subjected to molecular dynamics simulations, revealing a wide spectrum of stabilities. Among these, mutant S116M displayed a remarkable stability, with a structural deviation comparable to that of the experimental quadruple mutant M133L/V203A/N239Y/N268D that is thermodynamically more stable than that of the wild type by 2.6 kcal/mol. Structural analysis showed that the high stability of the S116M mutant was indeed due to the preservation of the p53C loop L1 conformation and the reduction of mobility in that region. The differential stabilities conferred by the single mutations are rationalized based on the geometries and chemical properties of the side chains introduced into this site. Linearity (i.e., nonbranched), moderate size, and balanced hydrophobic and hydrophilic properties of the side chain are crucial to the stabilizing effect of the residue substitutions.

Amino Acid Substitution↗

PubMed2005

The contribution of the Trp/Met/Phe residues to physical interactions of p53 with cellular proteins.

Dynamic molecular interaction networks underlie biological phenomena. Among the many genes which are involved, p53 plays a central role in networks controlling cellular life and death. It not only operates as a tumor suppressor, but also helps regulate hundreds of genes in response to various types of stress. To accomplish these functions as a guardian of the genome, p53 interacts extensively with both nucleic acids and proteins. This paper examines the physical interfaces of the p53 protein with cellular proteins. Previously, in the analysis of the structures of protein-protein complexes, we have observed that amino acids Trp, Met and Phe are important for protein-protein interactions in general. Here we show that these residues are critical for the many functions of p53. Several clusters of the Trp/Met/Phe residues are involved in the p53 protein-protein interactions. Phe19/Trp23 in the TA1 region extensively binds to the transcriptional factors and the MDM2 protein. Trp53/Phe54 in the TA2 region is crucial for transactivation and DNA replication. Met243 in the core domain interacts with 53BP1, 53BP2 and Rad 51 proteins. Met384/Phe385 in the C-terminal region interacts with the S100B protein and the Bromodomain of the CBP protein. Thus, these residues may assist in elucidating the p53 interactions when structural data are not available.

Biophysics↗

PubMed2005

A robust meta-classification strategy for cancer diagnosis from gene expression data.

One of the major challenges in cancer diagnosis from microarray data is to develop robust classification models which are independent of the analysis techniques used and can combine data from different laboratories. We propose a meta-classification scheme which uses a robust multivariate gene selection procedure and integrates the results of several machine learning tools trained on raw and pattern data. We validate our method by applying it to distinguish diffuse large B-cell lymphoma (DLBCL) from follicular lymphoma (FL) on two independent datasets: the HuGeneFL Affmetrixy dataset of Shipp et al. (www. genome.wi.mit.du/MPR /lymphoma) and the Hu95Av2 Affymetrix dataset (DallaFavera's laboratory, Columbia University). Our meta-classification technique achieves higher predictive accuracies than each of the individual classifiers trained on the same dataset and is robust against various data perturbations. We also find that combinations of p53 responsive genes (e.g., p53, PLK1 and CDK2) are highly predictive of the phenotype.

Algorithms↗

PubMed2005

A relative-entropy algorithm for genomic fingerprinting captures host-phage similarities.

The degeneracy of codons allows a multitude of possible sequences to code for the same protein. Hidden within the particular choice of sequence for each organism are over 100 previously undiscovered biologically significant, short oligonucleotides (length, 2 to 7 nucleotides). We present an information-theoretic algorithm that finds these novel signals. Applying this algorithm to the 209 sequenced bacterial genomes in the NCBI database, we determine a set of oligonucleotides for each bacterium which uniquely characterizes the organism. Some of these signals have known biological functions, like restriction enzyme binding sites, but most are new. An accompanying scoring algorithm is introduced that accurately (92%) places sequences of 100 kb with their correct species among the choice of hundreds. This algorithm also does far better than previous methods at relating phage genomes to their bacterial hosts, suggesting that the lists of oligonucleotides are "genomic fingerprints" that encode information about the effects of the cellular environment on DNA sequence. Our approach provides a novel basis for phylogeny and is potentially ideally suited for classifying the short DNA fragments obtained by environmental shotgun sequencing. The methods developed here can be readily extended to other problems in bioinformatics.

Algorithms↗