COLONY DEVELOPMENT IN EUDORINA ELEGANS (CHLOROPHYTA,VOLVOCALES)1

A detailed ultrastructure study was made of cell division and colony development in Eudorina elegans Ehrenberg. At the onset of cell division and prior to nuclear division the nucleus moved from the cell center to the cell surface. During nuclear division the nuclear membrane remained intact, except for openings occurring at the nuclear poles. The spindle microtubules appeared to arise from a MTOC-like (microtubule organizing centers) structure, while centrioles were absent from the nuclear poles. Following telophase, daughter nuclei formed which were separated by several distinct bands of endoplasmic reticulum. Cytokinesis occurred with formation of a cleavage furrow, associated with a typical phycoplast band of microtubules. However, cytokinesis was incomplete, resulting in formation of cytoplasmic bridges between the plakeal cells. Upon completion of up to five successive cell divisions, the plakea underwent inversion, which appeared to involve the production of colonial envelope material and rearrangement of cytoplasmic bridges. A new hypothesis concerning inversion is postulated based on these observations.     

Functional analysis of human T cell subsets defined by monoclonal antibodies. V. Suppressor cells within the activated OKT4+ population belong to a distinct subset

In the present report, we characterize a monoclonal antibody directed at a surface differentiation antigen on human T cells. The monoclonal antibody, OKT17, recognizes a cell surface antigen present on the majority of resting normal peripheral T cells. In contrast, OKT17 is unreactive with normal B cells, B cell lines, T cell lines, or SIg+ CLL. Interestingly, after activation, the antigen recognized by OKT17 is lost from a subset of OKT4+ cells. We took advantage of this finding to explore further the functional heterogeneity within activated OKT4+ cells. Evidence was obtained that the PWM-activated OKT4+ subset remaining after depletion of OKT17-reactive T cells (OKT4+ 17-) contains radiosensitive helperr cells but is devoid of suppressor cells. In contrast, the activated OKT4+ 17+ population contains potent radiosensitive suppressor cells as well as radioresistant helpe cells. Taken together, these studies suggest that the OKT17 monoclonal antibody can differentiate two functionally mature, activated OKT4+ human T cells: OKT4+ OKT17+ radiosensitive suppressor cells and OKT4+ 17- radiosensitive helper cells.     

Cyanide-insensitive and Cyanide-sensitive O(2) Uptake in Wheat: I. GRADIENT-PURIFIED MITOCHONDRIA

The mitochondrial fraction isolated from durum wheat seedlings by differential centrifugation demonstrated antimycin A- or cyanide-insensitive O₂ uptake. Further purification of this initial mitochondrial pellet using a linear Percoll (Pharmacia) density gradient separated the mitochondria into two bands of physiologically distinct activity. Based on the usual mitochondrial respiratory criteria of ADP/O and respiratory control values, these fractions were qualitatively similar to the crude pellet. However, we observed no antimycin A-insensitive O₂ uptake in either gradient band. Antimycin A-insensitive O₂ consumption could be restored to the upper gradient band of mitochondria by the addition of linoleic acid. This activity was inhibited either by salicylhydroxamic acid or propyl gallate, a known lipoxygenase inhibitor. Likewise, addition of linoleic acid to the crude mitochondrial pellet elicited a 4- to 5-fold increase in O₂ uptake. This O₂ consumption was insensitive to antimycin A and cyanide but was inhibited by either propyl gallate or salicylhydroxamic acid. Electron microscopic examination revealed that only the lower gradient band contained contamination-free mitochondria, which, in turn, lacked ability to oxidize linoleic acid. Antimycin A-insensitive O₂ consumption in the differential centrifugation fraction from germinating durum wheat seedlings decreased over 64 hours of development.     

Vasotocin, melatonin and narcolepsy: Possible involvement of the pineal gland in its patho-physiological mechanism

Both the pineal nonapeptide hormone arginine vasotocin (AVT) (2.5 μg) administered intra-nasally and the pineal indole melatonin (50 mg) administered intravenously to three male narcoleptics (two with auxiliary symptoms and one with sleep attacks only), dramatically increased the amount of REM sleep and decreased REM sleep latency. The duration of the sleep onset REM periods in the two narcoleptics with auxiliary symptoms increased by more than 100 percent after AVT and melatonin administration. In the narcoleptic with sleep attacks only both AVT and melatonin induced REM periods at sleep onset. The hypothesis is advanced that narcolepsy represents an impairment of the melatonin-AVT control in the induction and circadian organization of REM sleep associated with an immaturity of REM triggering centers.     

Insulin degradation by adipose tissue. Studies at several levels of cellular organization.

A systematic study of the degradation of physiological concentrations of 125I-labelled insulin was performed in intact fat-pads, isolated adipocytes and subcellular fractions of isolated adipocytes. The findings indicate that insulin is rapidly degraded to low-molecular-weight peptides and/or amino acids by the intact tissue and isolated cells. Of the total insulin-degradation products present after incubation with an intact fat-pad, 94% is recovered in the medium, indicating that these products are not retained by the cells or tissue. The plasma membranes do not degrade insulin significantly in the absence of reduced glutathione, and over 99% of the cellular degradative capacity is found in the postmicrosomal supernatant (cytosol). The cytosol degrades insulin to several labelled fragments that are intermediate in size between insulin and insulin A chain, as well as to the low-molecular-weight tissue degradation products. Inclusion of plasma membranes with cytosol accelerates the cleavage of the intermediate fragments to the size of the small products seen with the intact tissue. However, plasma membranes do not increase the initial step in the degradation of insulin when incubated with cytosol, suggesting that the insulin receptor is not involved with the direct cleavage of insulin. This study supports the hypothesis that the bulk of insulin degradation occurs in the adipocyte cytosol, where intermediate-sized fragments are generated and rapidly cleaved to smaller products by the plasma membrane and quickly released into the surrounding medium.     

Glutamine release from hindlimb and uptake by kidney in the acutely acidotic rat.

The arteriovenous difference (release) for glutamine across the hindlimb increases significantly during acute HCl-induced acidosis. This additional amount release by muscle tissue can account for the extra glutamine taken up by the kidney.     

A general multistate model for the analysis of hydrogen-exchange kinetics

In proton nmr, the chemical exchange rates of slowly exchanging labile hydrogens (with lifetimes in the range ～ 10 msec – ～ 1 sec) of peptides, proteins, and nucleic acids can be measured in H₂O by a combination of two separate experiments: (1) the transfer of solvent saturation and (2) saturation-recovery experiments. When these molecules exist in a dynamic equilibrium among different conformations, the experiments cannot be analyzed in a straightforward manner to derive the intrinsic exchange rates. In the present study we have derived analytical expressions for the above two experiments on a biomolecule under certain limiting conditions: (1) the extreme low-motility limit, where each of the conformational transitions is much slower than the corresponding hydrogen exchange rate with the solvent; (2) the high-motility limit (EX₂ mechanism), which is the opposite extreme of the previous limit; and (3) the low-motility limit (EX₁ mechanism), which is a mixture of limits (1) and (2), i.e., for some of the conformations, the exchange rate with the solvent is much faster than their conformational transition rates, while for the remaining conformations the reverse situation is realized. The results may be considered as a generalization to an arbitrary number of states of the two-state model treated by Hvidt. Equations have also been derived that are applicable to the iostope exchange method of measuring very slow exchange rates (with life-times of the order of minutes and longer) in biomolecules. The saturation recovery experiments performed in H₂O on the active pentapeptide fragment of thymopoietin serve to illustrate the high-motility limit. The theoretical formulation presented in this study can be easily adapted to other double-resonance techniques and also to situations where the kinetics of an arbitrary system existing in a multistate equilibrium are of interest.     

Some invariant properties of IgE-mediated basophil activation and desensitization.

We investigate certain general properties of antigen induced degranulation of sensitized basophils by analyzing two types of experiments: Experiments in which we expose basophils to two antigens sequentially and then determine the fraction of histamine released; and experiments in which we obtain time-dependent release and desensitization curves. To analyze the latter type of experiments we introduce a new way to plot release and desensitization data that depends on the nature of the interactions of histamine-containing units (histamine quanta) with themselves or the cells degranulation apparatus, but not on any specific properties of the antigen. From our analysis we conclude that: 1) A fraction of histamine within a population of basophils is nonreleasable by antigenic stimulation. 2) When a basophil degranulates the initial release of histamine appears to inhibit subsequent release. 3) The rate of histamine release is proportional to the amount of releasable histamine remaining in the cells when the amount remaining is small, as expected if release of histamine granules is a stochastic process. 4) There is no dependence of desensitization on the extracellular calcium concentration.     

Structural changes in simian virus 40 chromatin as probed by restriction endonucleases.

The structure of simian virus 40 (SV40) chromatin was probed by treatment with single- and multiple-site bacterial restriction endonucleases. Approximately the same fraction of the chromatin DNA was cleaved by each of three different single-site endonucleases, indicating that the nucleosomes do not have unique positions with regard to specific nucleotide sequences within the population of chromatin molecules. However, the extent of digestion was found to be strongly influenced by salt concentration. At 100 mM NaCl-5 mM MgCl2, only about 20% of the simian virus 40 (SV40) DNA I in chromatin was converted to linear SV40 DNA III. In contrast, at lower concentrations of NaCl (0.05 or 0.01 M), an additional 20 to 30% of the DNA was cleaved. These results suggest that at 100 mM NaCl only the DNA between nucleosomes was accessible to the restriction enzymes, whereas at the lower salt concentrations, DNA within the nucleosome regions became available for cleavage. Surprisingly, when SV40 chromatin was digested with multiple-site restriction enzymes, less than 2% of the DNA was digested to limit digest fragment, whereas only a small fraction (9 to 15%) received two or more cuts. Instead, the principal digest fragment was full-length linear SV40 DNA III. The failure to generate limit digest fragments was not a consequence of reduced enzyme activity in the reaction mixtures or of histone exchange. When the position of the principal cleavage site was mapped after HpaI digestion, it was found that this site was not unique. Nevertheless, all sites wree not cleaved with equal probability. An additional finding was that SV40 chromatin containing nicked-circular DNA II produced by random nicking of DNA I was also resistant to digestion by restriction enzymes. These results suggest that the initial cut which causes relaxation of topological constraint in SV40 chromatin DNA imparts resistance to further digestion by restriction enzymes. We propose that this may be accomplished by either "winding" of the internucleosomal DNA into the body of the nucleosome, or as suggested by others, by successive right-hand rotation of nucleosomes.