Removal Of Effectors Research Articles

The concept of space and competition in immune regulation.

Regulation of cell numbers and organ size in multicellular organisms is an important principle in biology. Experimental data in developmental biology indicate that there are mechanisms by which organs sense their total mass, linked to the regulation of cell size and proliferation, but not solely determined by either.1,2 Active monitoring of organ size is suggested by regeneration experiments following removal of part of an organ; this has been best illustrated by the regeneration of mammalian liver after partial hepatectomy.3 Another example is the demonstration that, following transplantation of multiple spleen fragments, the total spleen graft mass tends to reach a plateau which is in the range of variation of normal spleen weights.4 There are a multitude of mechanisms that regulate the number of cells (reviewed in ref. 1). For instance, the number of cell divisions may be predetermined, as in the nematode Caenorhabditis elegans,5 or divisions may stop after a given time interval (a mechanism for cell number control used in cardiac myocytes). Furthermore, hormones and components of signal-transduction pathways regulate growth and cell division, e.g. growth hormone6 and components of the insulin-signalling pathway.7 Another widely used principle in biological systems is competition for limiting resources, such as secretory molecules or cell-contact mechanisms. This is evident, for instance, in the control of oligodendrocyte precursors, which are regulated via the concentration of platelet-derived growth factor, whereas the number of mature oligodendrocytes is related to axon-dependent survival signals that are limited in amount and assure that the final number of oligodendrocytes is matched to the number of axons.8–10 In the mammalian immune system, size control checks maintain the number of peripheral T cells at more or less constant levels. This is not the result of a finite capacity for cell division, as it has been shown that T cells, following serial transfer into irradiated hosts, are able to divide up to 56 times in vivo, an expansion potential similar to that of colony-forming units.11 Thus, alternative mechanisms are involved in controlling the numbers of T cells in the face of ongoing new production from the thymus (at least for a certain period in development) and continuous expansion in response to antigenic stimuli. The American physiologist, Walter Cannon, introduced the term ‘homeostasis’ to describe the tendency of an organism to restore its original status in the face of unexpected disturbances.12 Permutations of this term are now widely used by immunologists referring to various response modes of T cells when the equilibrium is disturbed. The intrinsic dynamics of the immune system pose constant challenges threatening the equilibrium, and it is therefore understandable that the immune system has developed several layers of homeostatic control mechanisms.

Interleukin-2 receptor common gamma-chain signaling cytokines regulate activated T cell apoptosis in response to growth factor withdrawal: selective induction of anti-apoptotic (bcl-2, bcl-xL) but not pro-apoptotic (bax, bcl-xS) gene expression.

Cytokine deprivation from activated T cells leads to apoptosis associated with down-regulation of the bcl-2 gene product. It is not clear, however, how cytokines other than interleukin-2 (IL-2) may affect this process and regulate the involvement of other apoptosis-modulating genes. We show that a group of cytokines including IL-2 (IL-2R gamma), prevent the apoptosis of IL-2-deprived activated T cells. This rescue involves the induction of the anti-apoptosis genes bcl-2 and bcl-xL), but causes little change in expression of bax and bcl-xS, which promote apoptosis. Furthermore, the prevention of apoptosis and induction of proliferation by the common gamma chain cytokines can be dissociated. Thus, when proliferation is blocked, the common gamma chain cytokines still induce up-regulation of bcl-2 relative to bax and retard apoptosis. These cytokines can thus regulate the persistence or removal of effector T cells by coordinating the balance between genes which promote and those which inhibit apoptosis, events which are probably mediated at least in part by signals through the common gamma chain. These data also implicate inappropriate T cell apoptosis resulting from a dysfunctional common gamma-chain as part of the pathophysiological defect in patients with X-linked severe-combined immunodeficiency (SCID).

Control of amylase biosynthesis and release in the parotid gland of the rat

1. Amylase biosynthesis and release in the rat parotid were studied under various conditions. Incorporation of [(3)H]leucine into amylase, extracted from the tissue by immunoadsorbent, was measured and found to be time-dependent and totally inhibited by the protein synthesis inhibitor puromycin. 2. Adrenaline, at a concentration (10mum) that gave maximum stimulation of release, inhibited [(3)H]leucine incorporation into both total protein and amylase. This effect was reversed by phentolamine. 3. Adrenaline (1mum) and isoproterenol (10mum) stimulated biosynthesis of total protein and amylase. These effects were blocked by propranolol, as were the effects on release. Dibutyryl cyclic AMP (2mm) mimicked the effects of isoproterenol and adrenaline (1mum) on both amylase biosynthesis and release. All the above stimulatory effects on amylase biosynthesis were only observed if the tissue was pretreated with effector before pulse-labelling with [(3)H]leucine. 4. Insulin (625muunits/ml initial concentration, 150muunits/ml final concentration) stimulated incorporation of [(3)H]leucine into total protein and amylase when added to the tissue at the same time as the leucine. 5. Carbamoylcholine (10mum) decreased [(3)H]leucine incorporation into total protein and amylase when both were added to the tissue simultaneously, but this effect was prevented by removal of effector and washing the tissue before addition of [(3)H]leucine. 6. Stimulation of beta-adrenergic receptors increased both amylase release and biosynthesis, but stimulation of alpha-receptors can inhibit biosynthesis without inhibiting release. Cholinergic agents can also inhibit amylase biosynthesis, but stimulate release. Insulin at approximately physiological concentration can increase incorporation of leucine into amylase without stimulating release. The system described therefore provides an excellent model for the further investigation of the mechanisms of these diverse effects.

Studies on the quaternary structure of the threonine-sensitive aspartokinase-homoserine dehydrogenase of Escherichia coli. A proposed subunit-interaction model.

The threonine-sensitive aspartokinase-homoserine dehydrogenase of Escherichia coli can exist either as a tetramer (Mr= 360000) or a dimer, depending on the conditions. The subunit interactions of this enzyme have been investigated by reacting-enzyme sedimentation, gel filtration, and electrophoresis. Mild proteolysis with trypsin converts the native enzyme to a dimer which is smaller (Mr= 110000) than the native dimer (Mr= 175000). The new dimer fragment possesses homoserine dehydrogenase activity, sediments as a 6-S species in reacting-enzyme sedimentation but does not show aspartokinase activity. The dehydrogenase activity is lower, however, and the inhibition by threonine of this reaction has largely been lost. l-Threonine or to a lesser extent l-aspartate plus potassium ion protect aspartokinase-homoserine dehydrogenase against proteolysis probably by stabilization of the tetrameric form of the enzyme. Conditions which promote the formation of dimer (removal of effectors) increase the susceptibility to proteolysis. Various treatments cause dissociation of tetrameric aspartokinase-homoserine dehydrogenase, namely alteration of pH, removal of effectors, proteolysis, or sulfhydryl modification. Singly or in combination these treatments always resulted in the formation of dimers (6 S to 8 S in reacting enzyme sedimentation) rather than monomers. This finding strongly indicates that the dimers are formed in all cases by disruption of the same intersubunit bonding domain of the tetramer. Based upon these and other studies, we have proposed a subunit interaction model for aspartokinase-homoserine dehydrogenase. The model describes a molecule in which each of four identical subunits contains somewhat independent entities corresponding to aspartokinase and homoserine dehydrogenase activities. The native tetramer is held together solely by isologous interactions, two between aspartokinase entities and two between regions in which aspartokinase and homoserine dehydrogenase entities overlap. Dissociation by all mild treatments is explained by cleavage of the isologous interaction between aspartokinase entities resulting in loss of aspartokinase activity and production of a homoserine dehydrogenase dimeric unit.

Regulation of nitrate reductase activity by NADH and cyanide

NADH:nitrate oxidoreductase (EC 1.6.6.1) of Chlorella vulgaris is regulated by oxidation-reduction reactions. With a highly purified preparation of nitrate reductase, a second factor such as cyanide is required in addition to a reductant such as NADH to effect a rapid, reversible conversion of the active form to the inactive form. NADH and cyanide are the most effective combination yet tested, either reagent being effective at concentrations equivalent to the concentration of enzyme. Other reductants such as NADPH or dithionite may be substituted for NADH; and ferrocyanide, sulfide or hydroxylamine may be substituted for cyanide but higher concentrations of these reagents are required to effect a rapid inactivation of the enzyme. The inactive form of the enzyme is rapidly converted to the active form by the addition of ferricyanide. Regulation occurs on the nitrate-reducing moiety. Diaphorase activity is the same in the active and inactive forms. The conversion of the active form to the inactive form is prevented by nitrate, nitrite or flavins. No detectable changes in the absorption spectrum between 240 and 650 nm, or in molecular size, are associated with the inactivation-activation reaction. Both forms of the enzyme are stable after removal of effectors by gel filtration. No correlation was found between the state of activation and the state of oxidation of the cytochrome b associated with nitrate reductase.