p53, the Cellular Gatekeeper for Growth and Division

mTOR Signaling in Growth, Metabolism, and Disease.

In all eukaryotes, the target of rapamycin (TOR) signalling pathway couples energy and nutrient abundance to the execution of cell growth and division, owing to the ability of TOR protein kinase to simultaneously sense energy, nutrients and stress and, in metazoans, growth factors. Mammalian TOR complex 1 (mTORC1) and mTORC2 exert their actions by regulating other important kinases, such as S6 kinase (S6K) and Akt. In the past few years, a significant advance in our understanding of the regulation and functions of mTOR has revealed the crucial involvement of this signalling pathway in the onset and progression of diabetes, cancer and ageing.

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mTOR: from growth signal integration to cancer, diabetes and ageing

Blinded by the Light: The Growing Complexity of p53

Publisher Summary This chapter discusses the role of plasminogen activators in various biological processes. In specific, it describes two types of plasminogen activators—namely, the urokinase-type plasminogen activator (u-PA) and the tissue-type plasminogen activator (t-PA), which are essentially different gene products. The amino acid sequences of these activators and nucleotide sequences of the corresponding cDNAs have largely been determined, and the cDNAs have been cloned using recombinant techniques. A variety of enzymatic as well as immunological assay and detection methods have also been developed that allows a precise quantification of the activators, a distinction between u-PA and t-PA, determination of whether an activator is present in its active or zymogen form, analysis of the kinetics of different steps of the cascade reaction, and immunocytochemical identification of u-PA and t-PA in tissue sections. Much of the studies on plasminogen activators and cancer has been guided by the hypothesis that proteolysis of the components of extracellular matrix, initiated by the release of plasminogen activator from the cancer cells, plays a decisive role for the degradation of normal tissue, and thereby for invasive growth and metastases.

Plasminogen activators, tissue degradation, and cancer.

We report that the expression of murine or human mutant p53 proteins in cells with no endogenous p53 proteins confers new or additional phenotypes upon these cells. Mutant p53 proteins expressed in cell lines lacking p53 resulted in either enhanced tumorigenic potential in nude mice ((10)3 cells) or enhanced plating efficiency in agar cell culture (human SAOS-2 cells). Also, mutant human p53 alleles, unlike the wild-type p53 protein, could also enhance the expression of a test gene regulated by the multi-drug resistance enhancer-promoter element. These data demonstrate a gain of function associated with p53 mutations in addition to the loss of function shown previously to be associated with mutations in this tumour suppressor gene.

Gain of function mutations in p53.

The insulin-like growth factor 1 (IGF-1)-AKT-mTOR pathways sense the availability of nutrients and mitogens and respond by signaling for cell growth and division. The p53 pathway senses a variety of stress signals which will reduce the fidelity of cell growth and division, and responds by initiating cell cycle arrest, senescence, or apoptosis. This study explores four p53-regulated gene products, the beta1 and beta2 subunits of the AMPK, which are shown for the first time to be regulated by the p53 protein, TSC2, PTEN, and IGF-BP3, each of which negatively regulates the IGF-1-AKT-mTOR pathways after stress. These gene products are shown to be expressed under p53 control in a cell type and tissue-specific fashion with the TSC2 and PTEN proteins being coordinately regulated in those tissues that use insulin-dependent energy metabolism (skeletal muscle, heart, white fat, liver, and kidney). In addition, these genes are regulated by p53 in a stress signal-specific fashion. The mTOR pathway also communicates with the p53 pathway. After glucose starvation of mouse embryo fibroblasts, AMPK phosphorylates the p53 protein but does not activate any of the p53 responses. Upon glucose starvation of E1A-transformed mouse embryo fibroblasts, a p53-mediated apoptosis ensues. Thus, there is a great deal of communication between the p53 pathway and the IGF-1-AKT and mTOR pathways.

https://cancerres.aacrjournals.org/content/canres/67/7/3043.full.pdf

The Regulation of AMPK β1, TSC2, and PTEN Expression by p53: Stress, Cell and Tissue Specificity, and the Role of These Gene Products in Modulating the IGF-1-AKT-mTOR Pathways

Cancer is a disease of aging. The accumulation of mutations in individual cells over a lifetime is thought to be the reason. In this work, we explored an additional hypothesis: could p53 function decline with age, which would contribute to an enhanced mutation frequency and tumorigenesis in the aging process? The efficiency of the p53 response to γ-irradiation was found to decline significantly in various tissues of aging mice from several inbred strains, including lower p53 transcriptional activity and p53-dependent apoptosis. This decline resulted from a decreased stabilization of the p53 protein after stress. The function of the Ataxia-telangiectasia mutated (ATM) kinase declined significantly with age, which may then be responsible for the decline of the p53 response to radiation. Declining p53 responses to other stresses were also observed in the cultured splenocytes from aging mice. Interestingly, the time of onset of this decreased p53 response correlated with the life span of mice; mice that live longer delay their onset of decreased p53 activity with time. These results suggest an enhanced fixation of mutations in older individuals because of the declining fidelity of p53-mediated apoptosis or senescence in response to stress, and they suggest a plausible explanation for the correlation between tumorigenesis and the aging process.

https://www.pnas.org/content/pnas/104/42/16633.full.pdf

Declining p53 function in the aging process: A possible mechanism for the increased tumor incidence in older populations

Purified simian virus 40 (SV40) virions, grown in primary African green monkey kidney cells labeled prior to infection with 3 H-thymidine, contain a variable quantity of 3 H-labeled deoxyribonucleic acid (DNA). This DNA is resistant to deoxyribonuclease, sediments at 250 S , and is enclosed in a particle that can be precipitated with SV40-specific antiserum. DNA-DNA hybridization experiments demonstrate that this 3 H-labeled component in purified SV40 virions is cellular DNA. When this 3 H-labeled DNA is released from purified virus with sodium dodecyl sulfate, it has an average sedimentation constant of 14 S . Sedimentation through neutral and alkaline sucrose gradients shows that this 14 S DNA is composed of a collection of different sizes of DNA molecules that sediment between 11 and 15 S . As a result of this size heterogeneity, SV40 virions containing cellular DNA (pseudovirions) have a variable DNA to capsid protein ratio and exhibit a spectrum of buoyant densities in a CsCl equilibrium gradient. Pseudovirions are enriched, relative to true virions, on the lighter density side of infectious SV40 virus banded to equilibrium in a CsCl gradient. Little or no cellular DNA was found in purified SV40 virus preparations grown in BSC-1 or CV-1 cells.

Deoxyribonucleic acid replication in simian virus 40-infected cells. II. Detection and characterization of simian virus 40 pseudovirions.

Embryoid bodies are produced when a transplantable testicular teratoma from strain 129 mice is serially passaged in the peritoneal cavity of these mice. These bodies are roughly spherical containing two morphologically distinct cell types. Scanning and transmission electron microscopy have been employed to show that the endodermal cells of embryoid bodies, like those of the mouse embryo, are on the outer surface and have highly convoluted surfaces containing numerous microvilli-like projections. The inner, embryonal carcinoma cells, are the pluripotent stem cells of this tumor.



Intracisternal A-type particles have been observed in electron micrographs and are almost exclusively located in the endodermal cells of the embryoid bodies. The A-type complement-fixing antigen has been identified in extracts prepared from this tumor.



When embryoid bodies are placed in culture and allowed to attach to the surface of a petri dish, a large number of new morphologically distinct cell types appear. Attachment to the petri dish surface is required for the generation of these new cell types. Cells of similar morphology in culture, display a distinctly “clonal” distribution on the petri dish surface.

Angelika K. Teresky

Papers

Gain of function mutations in p53.

The Regulation of AMPK β1, TSC2, and PTEN Expression by p53: Stress, Cell and Tissue Specificity, and the Role of These Gene Products in Modulating the IGF-1-AKT-mTOR Pathways

Declining p53 function in the aging process: A possible mechanism for the increased tumor incidence in older populations

Deoxyribonucleic acid replication in simian virus 40-infected cells. II. Detection and characterization of simian virus 40 pseudovirions.

Morphological criteria for the in vitro differentiation of embryoid bodies produced by a transplantable teratoma of mice