TAK1 kinase switches cell fate from apoptosis to necrosis following TNF stimulation

TNF activates three distinct intracellular signaling cascades leading to cell survival, caspase-8–mediated apoptosis, or receptor interacting protein kinase 3 (RIPK3)–dependent necrosis, also called necroptosis. Depending on the cellular context, one of these pathways is activated upon TNF challenge. When caspase-8 is activated, it drives the apoptosis cascade and blocks RIPK3-dependent necrosis. Here we report the biological event switching to activate necrosis over apoptosis. TAK1 kinase is normally transiently activated upon TNF stimulation. We found that prolonged and hyperactivation of TAK1 induced phosphorylation and activation of RIPK3, leading to necrosis without caspase activation. In addition, we also demonstrated that activation of RIPK1 and RIPK3 promoted TAK1 activation, suggesting a positive feedforward loop of RIPK1, RIPK3, and TAK1. Conversely, ablation of TAK1 caused caspase-dependent apoptosis, in which Ripk3 deletion did not block cell death either in vivo or in vitro. Our results reveal that TAK1 activation drives RIPK3-dependent necrosis and inhibits apoptosis. TAK1 acts as a switch between apoptosis and necrosis.     

Immunocytochemical detection of triphosphoinositide in vestibular hair cells

Triphosphoinositide (TPI), an aminoglycoside receptor and a possible regulator of cationic permeation through its ability to bind with Ca++, was localized by the protein-A gold technique in vestibular sensory epithelia using an antibody highly specific to TPI. TPI was detected on the stereocilia, kinocilia, and cuticular plate of hair cells, and in the reticular membrane of supporting cells. The cilia of hair cells are damaged by aminoglycosides at a relatively early stage of toxicity. Ca++-regulated bioactivity in this area is probably involved.     

Synthesis and transport of the organic matrix of the spicules in the gorgonian Leptogorgia virgulata (Lamarck) (Coelenterata: Gorgonacea)

Summary The sequence of the synthesis and transport of the organic matrix of spicules has been elucidated in the gorgonian Leptogorgia virgulata by use of ³H-aspartic acid as the tracer in electron-microscopic autoradiography. The entire process of matrix synthesis and transport takes approximately 2 h. It seems that the protein moiety of the organic matrix is synthesized in the RER prior to 5 min following the initial 10 min incubation in the tracer. At the 5 min chase the label is moving from the RER to the Golgi complexes where the carbohydrate moiety of the matrix is presumed to be synthesized. At the 5 to 15 min chases the label is transported out of the Golgi complexes via Golgi vesicles. This phase continues for 30 min. From 60 to 120 min the ³H-aspartic acid moves to the spicules. After 120 min the majority of the label has moved into the spicules. Silver grain counts over both multivesicular and electron-dense bodies remain at relatively low and constant levels over 4 h indicating that neither organelle is involved in the synthesis and transport of the organic matrix.Contribution No 512; Belle W. Baruch Institute for Marine Biology and Coastal Research, University of South Carolina, Columbia, South Carolina 29208, USA     

Binding sites of an aminoglycoside in the cochlea examined by immunocytochemistry

Summary To localize the binding sites of aminoglycosides in the cochlea, immunocytochemistry was used with the antibody to gentamicin and the protein-A/gold complex. We found that the main binding sites were the stereocilia, the cuticular plates of hair cells, the head plates of Deiters' cells, cell filaments and the cones of pillar cells, tectorial membranes, basilar membranes, the matrix of the spiral limbus, plasma membranes, mitochondria, and the chromatin of various kinds of cells. Triphosphoinositide and acidic glycosaminoglycans are the two most likely candidates for the cause of binding activity.     

Butyrate-binding protein from rat and mouse liver

A novel component which specifically binds butyrate was found in rat and mouse liver. This component, termed butyrate binding protein (BBP), is localized in the cytosolic fraction and exhibits protein characteristics, such as heat- and protease-sensitivity. The size of BBP was found to be 7.6S, while it was converted to subunits of 45,000--48,000 dalton by treatment with sodium dodecyl sulfate. The dissociation constant of the binding of BBP with butyrate was 2.22 X 10(-6) M in the standard assay. 30-Fold purification of BBP was achieved by batch-wise adsorption and elution from CM-cellulose and hydroxylapatite column chromatography. BBP is clearly distinguishable from the fatty acid-binding protein found previously on the basis of its size and binding specificity.     

Effect of larval brain extracts derived from three swallowtail butterflies,Papilio helenus L., P. machaon L. and P. memnon L., on seasonal morph development of P. xuthus L. (Lepidoptera: Papilionidae)

Adults of the three papilionid butterflies, Papilio helenus L., Papilio machaon L. and Papilio memnon L., exhibit seasonal diphenism comprising spring and summer morphs. To elucidate the physiological mechanism underlying seasonal morph development in papilionid butterflies, we investigated whether a cerebral factor showing summer‐morph‐producing hormone (SMPH) activity is present in the brain of three Papilio species using an assay system with chilled male short‐day pupae of P. xuthus L. When 2% NaCl extracts derived from 20 larval brains of the three species were injected into abdomens of chilled male short‐day pupae of P. xuthus, all recipients destined to develop into spring‐morph adults developed into summer‐ and intermediate‐morph adults. On the other hand, all recipients injected with distilled water as a control developed into spring‐morph adults. These results indicate that a cerebral factor showing SMPH activity is present in the larval brain of the three Papilio species. Additionally, all recipients injected with 2% NaCl extracts derived from 20 adult brains of Bombyx mori L. also developed into summer‐ and intermediate‐morph adults. The results revealed that SMPH or a cerebral factor showing SMPH activity is widely distributed among lepidopteran insects.     

H-Ferritin Is Preferentially Incorporated by Human Erythroid Cells through Transferrin Receptor 1 in a Threshold-Dependent Manner

Ferritin is an iron-storage protein composed of different ratios of 24 light (L) and heavy (H) subunits. The serum level of ferritin is a clinical marker of the body’s iron level. Transferrin receptor (TFR)1 is the receptor not only for transferrin but also for H-ferritin, but how it binds two different ligands and the blood cell types that preferentially incorporate H-ferritin remain unknown. To address these questions, we investigated hematopoietic cell-specific ferritin uptake by flow cytometry. Alexa Fluor 488-labeled H-ferritin was preferentially incorporated by erythroid cells among various hematopoietic cell lines examined, and was almost exclusively incorporated by bone marrow erythroblasts among human primary hematopoietic cells of various lineages. H-ferritin uptake by erythroid cells was strongly inhibited by unlabeled H-ferritin but was only partially inhibited by a large excess of holo-transferrin. On the other hand, internalization of labeled holo-transferrin by these cells was not inhibited by H-ferritin. Chinese hamster ovary cells lacking functional endogenous TFR1 but expressing human TFR1 with a mutated RGD sequence, which is required for transferrin binding, efficiently incorporated H-ferritin, indicating that TFR1 has distinct binding sites for H-ferritin and holo-transferrin. H-ferritin uptake by these cells required a threshold level of cell surface TFR1 expression, whereas there was no threshold for holo-transferrin uptake. The requirement for a threshold level of TFR1 expression can explain why among primary human hematopoietic cells, only erythroblasts efficiently take up H-ferritin.