The Chlamydomonas cell cycle is regulated by a light/dark-responsive cell-cycle switch

Cultures of the unicellular green alga Chlamydomonas reinhardii can be synchronized by light/dark cycling not only under photoautotrophic but also under mixotrophic growth conditions. We observed that cultures synchronized in the presence of acetate continue to divide synchronously for one cell-cycle period when transferred to heterotrophic growth conditions. This finding enabled us to investigate the differential effects of light on cell growth and cell division. When cells were exposed to continuous light at the beginning of the growth period they entered the division phase earlier than dark-grown cells as a consequence of an increased growth rate. Illumination at the end of the growth period, however, caused a considerable delay in cell division and an extended growth period. The light-induced delay in cell division was also observed in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of photosystem II. This finding demonstrates that cell division is directly influenced by a light/dard-responsive cell-cycle switch rather than by light/dark-dependent changes in energy metabolism. The importance of this light/dark control to the regulation of the Chlamydomonas cell cycle was investigated in comparison with other control mechanisms (size control, time control). We found that the light/dard-responsive cell-cycle switch regulates the transition from G1-to S-phase. This control mechanism is effective in cells which have attained the commitment to at least one round of DNA replication and division but have not attained the maximal cell mass which initiates cell division in the light.Abbreviations dCTP deoxycytidine 5-triphosphate - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea     

Synaptic changes in the terminals of rod photoreceptors of albino mice after partial visual cell loss induced by brief exposure to constant light

Summary Albino mice were exposed to constant light for 7 days and were then transferred to periodic light. After initial photic damage and partial cell loss, the remaining visual cells recovered and survived as a stable population. Regions of the outer nuclear layer containing 4–6 rows of nuclei were more affected than those containing 6–10 rows. Changes in the synaptic structures in the receptor terminals of these two regions were recorded after varying survival periods. Some of the rod terminals had multiple synaptic ribbons and larger numbers of horizontal cell processes and bipolar cell dendrites. The number of terminals with multiple ribbons increased during recovery in periodic light. Morphometry demonstrated that the perimeters of horizontal and bipolar cell processes within the rod terminals were significantly larger than those in age-matched control mice, especially 4 weeks after recovery; they remained significantly larger than controls after 2 and 3 months. We suggest that partial loss of rod cells within a group of cells that are synaptically related to a common bipolar or horizontal cell results in synaptic growth inside the terminals of the surviving cells.     

Effects of light and acetate on the liberation of zoospores by a mutant strain ofChlamydomonas reinhardtii

In light-dark-synchronized cultures of the unicellular green algaChlamydomonas reinhardtii, release of zoospores from the wall of the mother cell normally takes place during the second half of the dark period. The recently isolated mutant ls, however, needs light for the liberation of zoospores when grown photoautotrophically under a 12 h light-12 h dark regime. The light-induced release of zoospores was found to be prevented by addition of the photosystem-II inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Furthermore, light dependence of this process was shown to be abolished when the mutant ls was grown either photoautotrophically under a 14 h light-10 h dark regime or in the presence of acetate. Our findings indicate that the light-dependency of zoospore liberation observed in cultures of this particular mutant during photoautotrophic growth under a 12 h light-12 h dark regime might be attributed to an altered energy metabolism. The light-induced release of zoospores was found to be prevented by addition of cycloheximide or chloramphenicol, antibiotics which inhibit protein biosynthesis by cytoplasmic and organellar ribosomes, respectively. Actinomycin D, an inhibitor of RNA synthesis, however, did not affect the light-induced liberation of zoospores.Sporangia accumulate in stationary cultures of the mutant ls. Release of zoospores was observed when these sporangia were collected by centrifugation and incubated in the light after resuspension in fresh culture medium. Since liberation of zoospores was not observed after dilution of the stationary cultures with fresh culture medium, we suppose that components which interfere with the action of the sporangial autolysin are accumulated in the culture medium of the mutant ls.Abbreviation DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea     

Algesia and local responses induced by neurokinin A and substance P in human skin and temporal muscle

Neurokinin A (NKA), substance P (SP) and the two peptides combined (SP + NKA) were injected intracutaneously on the forearm and into the temporal muscle of healthy volunteers. Pain intensity, cutaneous wheal and flare responses and tenderness of the temporal muscle were quantitated. SP but not NKA induced cutaneous pain. This relates the algesic effect of SP to the specific N-terminal amino acid sequence of the peptide, not shared by NKA. NKA, however, potentiated the algesic effect of SP as SP + NKA induced a significantly prolonged cutaneous pain sensation. Both peptides induced wheals, but only SP induced flare. These results confirm previous studies relating wheal formation to the identical C-terminal amino acid sequence of the two peptides and flare reaction to the N-terminal part of SP. Injections into the temporal muscle did not cause pain or tenderness.     

Ultrastructure of neutrophilic granulopoiesis in the bone marrow of patients with chronic myeloid leukemia (CML). A morphometric study with special emphasis on azurophil (primary) and specific (secondary) granules

In seven patients with chronic myeloid leukemia (CML) and ultrastructural and morphometric study was performed on neutrophilic granulopoiesis in bone marrow trephine biopsies. Bone marrow specimens from five patients without hematological abnormalities served as controls. In stable phases of CML, abnormalities of the maturing granulocytic lineage were most conspicuously expressed by an infrequently occurring nuclear disfiguration (blebs and disturbed bridging of segments). Morphometric evaluation included the numbers of azurphil (primary) and specific (secondary) granules, the cisternal length of the endoplasmic reticulum and the area of the mitochondrial profiles. These variables could be determined in early and late myeloblasts, promyelocytes, metamyelocytes, band cells and mature polymorphonuclear granulocytes. Statistical analysis with regard to control specimens demonstrated no significant differences in the total amount of neutrophil granules or of the other cell organelles.     

Characterization of lipopolysaccharide-induced macrophage gene expression

A cDNA library from LPS-treated murine peritoneal macrophages has been screened by differential hybridization with radiolabeled cDNA from untreated and LPS-treated macrophages. Six clones hybridizing with mRNA sequences present in LPS-treated cells but not in controls were selected for further characterization. When the recombinant bacteriophage DNA from each clone was used as a probe in Northern analysis of total RNA from LPS-treated macrophages, inducible mRNA ranging from 1.45 to 6.4 kb were seen. In five of six cases, the mRNA expression was undetectable in untreated macrophage cultures. All but one clone identified mRNA that were inducible even in the presence of cycloheximide, indicating the independence of such gene expression from protein synthesis; none of the genes were superinduced by this treatment. The time course of expression differed among the individual genes. Four were induced transiently, whereas two showed stable increasing accumulation through an 8-h period after stimulation. In addition, four of the genes were seen within 30 min of stimulation, whereas two were seen only after 2 to 4 h. Two genes were induced only by treatment with LPS, whereas four were also induced in response to other agents, including IFN-gamma, macrophage CSF, and PMA. The insert sequences from these recombinant clones did not hybridize with a set of cDNA encoding other inducible gene products, including TNF, IL-1, ornithine decarboxylase, c-myc, c-fos, JE, or KC. Thus, these six cDNA appear to encode inducible macrophage genes that are distinct from one another as well as from a selection of previously described early genes. Although their functional identity remains indeterminate, they may encode previously described early proteins induced in macrophages treated with LPS.     

Effect of the specific cholecystokinin-receptor antagonist loxiglumide on bombesin stimulated pancreatic enzyme secretion in man.

We have investigated the effects of the specific cholecystokinin (CCK) receptor antagonist loxiglumide on basal and bombesin stimulated pancreatic enzyme secretion, bilirubin output and plasma CCK release in six healthy subjects. The data were compared with those obtained in control experiments where saline was infused instead of loxiglumide. Basal amylase output (4.7 +/- 0.8 kU/45 min), trypsin output (2.9 +/- 0.8 kU/45 min) and bilirubin output (7.7 +/- 2.8 mmol/45 min) gradually declined during infusion of loxiglumide to values of 1.3 +/- 0.3 kU/45 min, 0.5 +/- 0.1 kU/45 min and 0.4 +/- 0.0 mmol/45 min, respectively, reaching statistical significance (P less than 0.05) in the 30 to 45-min period after the start of the loxiglumide infusion. In the control experiments saline infusion failed to influence basal amylase, trypsin and bilirubin output, while bombesin stimulated amylase output from 4.7 +/- 0.8 kU/45 min to 25.1 +/- 5.1 kU/45 min (P less than 0.05), trypsin output from 2.9 +/- 0.8 kU/45 min to 11.6 +/- 2.0 kU/45 min (P less than 0.05) and bilirubin output from 7.7 +/- 2.8 mmol/45 min to 68.0 +/- 16.0 mmol/45 min (P less than 0.05). Loxiglumide failed to significantly influence bombesin stimulated amylase output (36.7 +/- 9.0 kU/45 min) and trypsin output (8.3 +/- 2.9 kU/45 min), but almost abolished bilirubin output (9.7 +/- 3.6 mmol/45 min) (P less than 0.05). Basal plasma CCK (2.4 +/- 0.1 pM) was not significantly influenced by loxiglumide (2.4 +/- 0.2 pM).(ABSTRACT TRUNCATED AT 250 WORDS)     

Modified structure and kinetics of cytochrome-c oxidase in fibroblasts from patients with Leigh syndrome

In this study we compared the properties of cytochrome-c oxidase (COX) in cultured fibroblasts from two patients with Leigh Syndrome with COX from control fibroblasts. The fibroblasts from patients showed decreased growth reates and elevated lactate production. COX activity of patients fibroblasts was about 25% of control. Kinetic studies with isolated mitochondria showed a higher K_m for cytochrome c and a markedly reduced molecular turnover of COX from patients, indicating a different structure of the enzyme. A biphasic change of COX activity was obtained by titration of dodecylmaltoside solubilized mitochondria from control fibroblasts with increasing concentrations of anions. With patient mitochondria we found only the inhibiting phase of COX activity and, in contrast to control mitochondria, irreversible inhibition of COX activity by guanidinium chloride. ELISA titrations with monoclonal antibodies to subunit II, IV, Vab, VIac and VIIab indicated a normal amount of mitochondrial coded subunit II, but a reduced amound of nuclear coded subunits. The data indicate incompletely assembled nuclear coded subunits of COX from patient fibroblasts.     

Androgen receptor heterogeneity in LNCaP cells is caused by a hormone independent phosphorylation step

Androgen receptor synthesis and modification were studied in the human LNCaP cell line. Immunoblotting showed that the androgen receptor migrated as a closely spaced 110–112 kDa doublet on SDS-PAGE gels. Most of the receptor protein is present in the higher molecular mass form. Labelling experiments with [³⁵S]methionine showed that the androgen receptor is synthesized as a single 110 kDa protein which is rapidly converted to a 112 kDa protein. Upon alkaline phosphatase treatment a gradual elimination of the 112 kDa isoform with a concomitant increase of the 110 kDa isoform was seen, indicating that the observed 110 to 112 kDa upshift reflects androgen receptor phosphorylation. Furthermore, it is shown that both isoforms can bind hormone and undergo a hormone dependent transformation to a tight nuclear binding form, indicating that the 110 to 112 kDa conversion is not an obligatory step for hormone binding or receptor transformation.