A reassessment of the telomere hypothesis of senescence

According to the telomere hypothesis of senescence, the telomeric shortening that accompanies the replication of normal somatic cells acts as the mitotic clock that eventually results in their permanent exit from the cell cycle. Although evidence consistent with the telomere hypothesis continues to accumulate, on the basis of recent findings it is suggested that instead of a single clock mechanism there are multiple inducers of senescence.     

Rb function in the apoptosis and senescence of non-neuronal and neuronal cells: role in oncogenesis

Regulators of the cell cycle machinery play a major role in modulating a variety of cellular phenomena including proliferation, quiescence, differentiation, senescence and apoptosis. Studies in the past decade have clearly established a role for the retinoblastoma tumor suppressor protein, Rb, and its primary downstream target E2F1, in the above processes. While the role of the Rb protein in the regulation of cell cycle progression has been analyzed in great detail, its potential roles in apoptosis as well as senescence are relatively less studied. It has become increasingly clear that the anti-apoptotic functions of Rb contribute significantly to the genesis and progression of tumors. This is especially relevant in neuronal systems, since terminally differentiated neurons do not proliferate; therefore the normal anti-proliferative functions of Rb in neurons are not very dominant. This chapter describes the current thoughts on the role of Rb function in the apoptosis and senescence of cells, both of neuronal and non-neuronal origin. Recent studies have also addressed how Rb function is differentially modulated by proliferative and apoptotic signals received at the cell surface, though both lead to Rb inactivation. The contribution of Rb to inducing cellular senescence has been long recognized, but the underlying molecular mechanisms are being elucidated only recently; the contribution of this function of Rb to tumor suppression remains to be understood in detail. It can be expected that an understanding of Rb function in cellular apoptosis and senescence will enhance our ability to develop novel agents and strategies to combat cancer.     

Population cycles in microtines: The senescence hypothesis

Summary The cause of population cycles in microtines (voles and lemmings) remains an enigma. I propose a new solution to this problem based on a crucial feature of microtine biology, shifts in age structure, that has been ignored until now. Empirical evidence indicates that age structure must shift markedly towards older animals during declines because of three characteristics of the previous peak year: a shortened breeding season, total replacement of the breeding population from peak to decline and density-dependent social inhibition of maturation of young. Declines become inevitable as populations composed of older animals survive and reproduce poorly because of the effects of senescence, possibly interacting with the experiences of peak density and I present both theoretical and empirical evidence for this hypothesis. Although a variety of physiological systems deteriorate with aging, I focus on a crucial one — the inability of older animals to effectively maintain homeostasis in the face of environmental challenges because of a progressive deterioration in the endocrine feedback mechanisms involved in the hippocampal—hypothalamic—pituitary—adrenal axis. Microtine populations will not exhibit cycles where age structure shifts are prevented owing to extrinsic factors such as intense predation. Six testable predictions are made that can falsify this hypothesis.     

Regulation and execution of molecular disassembly and catabolism during senescence

Senescence is a highly orchestrated developmental stage in the life cycle of plants. The onset of senescence is tightly controlled by signaling cascades that initiate changes in gene expression and the synthesis of new proteins. This complement of new proteins includes hydrolytic enzymes capable of executing catabolism of macromolecules, which in turn sets in motion disassembly of membrane molecular matrices, leading to loss of cell function and, ultimately, complete breakdown of cellular ultrastructure. A distinguishing feature of senescence that sets it apart from other types of programmed cell death is the recovery of carbon and nitrogen from the dying tissue and their translocation to growing parts of the plant such as developing seeds. For this to be accomplished, the initiation of senescence and its execution have to be meticulously regulated. For example, the initiation of membrane disassembly has to be intricately linked with the recruitment of nutrients because their ensuing translocation out of the senescing tissue requires functional membranes. Molecular mechanisms underlying this linkage and its integration with the catabolism of macromolecules in senescing tissues are addressed.     

A methodological study for the quantification and the control of the physico-chemical processes occurring during cell fixation

Summary Fixation in a traditional sense means the immersion of biological material into a chemical fluid. For permanent preservation the fixative is always offered (1) in excess of the cell sample, and the process of fixation is influenced by (2) chemical impurities of the fixative fluid.Both factors influence the succeeding dyeing of cells. In order to avoid these uncontrolled criteria, a new technology for controlled cell fixation has been developed, whereby freshly prepared formaldehyde and methanol gas in an inert gas-flow of helium was applied to thin membranes by aid of a capillary flow-in technique.The instrumental equipment consists of (1) an ultra-high vacuum flowapparatus with a total-pressure measuring unit, (2) a gas-supply device, (3) a mass spectrometer including a pump system, and (4) a Teflon and/or glass-gas chamber for the treatment of synthetic (Hostaphan foils) or biological membranes (mesenterium) with formaldehyde as the fixative gas.The amount of offered, adsorbed, absorbed, diffused, and desorbed fixative gas could be absolutely estimated after the saturation of the membranes with an on-line operating inert mass spectrometer of the Omegatron type.The gas treatment of the Hostaphan foils with formaldehyde showed that nearly all adsorbed gas molecules could be desorbed. In contrast to native membranes the greatest proportion of the gas molecules adhered to the biological surface, and only a small quantity were desorbable. Physisorption or physisorption and chemisorption occured depending on the adsorber surface property.A monolayer of formaldehyde of 5·10¹⁴ to 1·10¹⁵ molecules per 10¹⁶ Ų surface area can be postulated on the basis of these preliminary results. This value corresponds to a mass of about 5·10^–8g CH₂O. It resulted in an area-coverage ratio of CH₂O molecules per cell of 10⁹:1.The membrane surface facing the gas side always amounted to 1 cm². A fixative gas concentration of 10⁶ molecules/cm³, and therefore a degree of coverage of <1/1000 monolayer can be estimated absolutely. For a precise determination of the degree of fixation, further experiments and the evaluation of additional physico-chemical parameters are necessary.     

Regulation of homeostasis and oncogenesis in the intestinal epithelium by Ras

Much of our current state of knowledge pertaining to the mechanisms controlling intestinal epithelial homeostasis derives from epidemiological, molecular genetic, cell biological, and biochemical studies of signaling pathways that are dysregulated during the process of colorectal tumorigenesis. Activating mutations in members of the RAS oncoprotein family play an important role in the progression of colorectal cancer (CRC) and, by extension, intestinal epithelial homeostasis. Mutations in K-RAS account for 90% of the RAS mutations found in CRC. As such, the study of RAS protein function in the intestinal epithelium is largely encompassed by the study of K-RAS function in CRC. In this review, we summarize the data available from genetically defined in vitro and in vivo models of CRC that aim to characterize the oncogenic properties of mutationally activated K-RAS. These studies paint a complex picture of a multi-functional oncoprotein that engages an array of downstream signaling pathways to influence cellular behaviors that are both pro- and anti-tumorigenic. While the complexity of K-RAS biology has thus far prevented a comprehensive understanding of its oncogenic properties, the work to date lays a foundation for the development of new therapeutic strategies to treat K-RAS mutant CRC.     

Regulation of ethylene biosynthesis during senescence of oat leaf segments

The evolution of endogenous ethylene, the conversion of 1-aminocylopropane-1-car-boxylic acid (ACC) to ethylene and the amounts of ACC (free and conjugated) have been followed during the senescence of oat ( Avena sativa L. cv. Victory) leaf segments. During the first three days of incubation of leaf segments in darkness, endogenous ethylene evolution and ACC-dependent ethylene production displayed a close relationship, both showing an increase followed by a decrease to the basal rate. However, unlike ethylene production, the level of ACC increased during the five days of incubation in the dark without any decline. It is concluded that ACC synthesis does not limit ethylene production, at least in the last stages of leaf senescence when ethylene production markedly decreased. The level of conjugated ACC increased and reached a plateau already at the first day of incubation. Yet, at the progressive stages of senescence, when the level af ACC gradually increased, no further conjugation of ACC could be detected. Thus, conjugation of ACC cannot account for ethylene drop at the last stages of oat leaf senescence.     

A mathematical model of the processes of mitochondrial death and reproduction during body irradiation

A method of construction of mathematical model of processes of death and reproduction of mitochondria based on morphometric measurements is proposed. Using the model the time of mitochondria death and reproduction during ionized irradiation of the organism is identified.     

Changes occurring during aging and senescence in a submerged aquatic angiosperm (Potamogeton pectinatus)

Changes occurring during aging and senescence of leaves of a submerged aquatic angiosperm ( Potamogeton pectinatus L.) were studied. Total chlorophyll and chlorophylls a and b were maximal in mature, and minimal in old leaves. The chlorophyll a to b ratio was highest in mature leaves. During senescence, the chlorophyll content and the ratio of chlorophyll a to b decreased. The content of DNA, RNA, protein and dry weight, and the activity of alkaline pyrophosphatase decreased while free amino acids, the activity of protease, RNase and acid pyrophosphatase, and the ratio of acid to alkaline pyrophosphatase activity increased during aging and senescence. Kinetin (0.23 m M ) deferred leaf senescence by delaying the loss of chlorophyll, protein, nucleic acids and dry weight, and reducing the rise in free amino acids, the activity of protease, RNase and acid pyrophosphatase and the ratio of acid to alkaline pyrophosphatase activity; while both 0.69 m M ethrel and 0.075 m M ABA hastened senescence. Kinetin pretreatment for an optimum period (12 h) followed by ethrel or ABA treatment partially erased the senescence-promoting effect of the latter. But treatments in a reverse order markedly reduced the delaying effect of kinetin on senescence.     

Regulation of enzyme activity by enzyme orientation: A hypothesis

In addition to substrate binding sites, many enzymes must possess supersubstrate binding sites that regulate attachment and orientation of the enzyme toward the matrix (micelle, membrane) in which the substrate molecules are embedded, the supersubstrate.     

Regulation of mitochondrial respiration in senescence

The ADP-stimulated (State 3) respiration of myocardial mitochondria with glutamate-malate, glutamate-pyruvate, palmitylcarnitine and β-hydroxybutyrate as substrates declined in rats after the age of 20 months. There was no significant decline in pyruvate-malate, α-oxoglutarate, palmityl-CoA, succinate and ascorbate cytochrome c oxidation. Skeletal muscle mitochondria from senescent animals showed a similar decline in glutamate-malate oxidation but not in palmityl-CoA, palmitylcarnitine, succinate and ascorbate-cytochrome c oxidation. The controlled oxidation with ADP-limiting (State 4) and the ADP/O ratio were not affected. The results indicate an alteration in the subtle regulatory capacity for mitochondrial oxidation in senescent rats. It is suggested that the alteration may be in certain anion transport and associated functions across the mitochondrial membrane or dehydrogenase activity.     

A hypothesis for the role of dithiol-disulfide interchange in solute transport and energy-transducing processes

We have recently shown that the physical mechanism for delta approximately mu H+-driven changes in the Km for three different transport systems is an oxidation-reduction reaction involving a dithiol-disulfide interconversion [Robillard, G.T. and Konings, W.N. (1981) Biochemistry, 20, 5025-5032; Konings, W.N. and Robillard, G.T. (1982) Proc. Natl Acad. Sci. USA, in the press]. Based on the similarities between the data from these three systems and published data from other systems, we now propose that dithiol-disulfide interchange may play a general role in membrane-related processes such as transport, energy transduction and hormone-receptor interactions. We propose that the affinities of the substrate-binding sites are regulated by a dithiol and a disulfide situated at different depths in the membrane. In addition we propose that the oxidation states of these two redox centers are coupled by dithiol-disulfide interchange such that, when one is oxidized, the other is reduced. Since a transmembrane electrical potential, delta psi, or a pH gradient, delta pH, can alter the redox state, it can change the affinity of the substrate-binding sites. The delta approximately mu H+-induced changes in affinity are sufficient to drive active transport (symport or antiport) and energy-transducing processes. A similar mechanism can be applied to transport systems driven by phosphorylated enzyme intermediates instead of delta approximately mu H+. Changes of the redox potential in a given compartment during metabolism could also control the affinity of ligand binding even in the absence of a delta approximately mu H+. The ligand-binding affinities of facilitated diffusion transport systems and receptor proteins may be regulated in this manner.