Munc13-1 is essential for fusion competence of glutamatergic synaptic vesicles.

Neurotransmitter release at synapses between nerve cells is mediated by calcium-triggered exocytotic fusion of synaptic vesicles. Before fusion, vesicles dock at the presynaptic release site where they mature to a fusion-competent state. Here we identify Munc13-1, a brain-specific presynaptic phorbol ester receptor, as an essential protein for synaptic vesicle maturation. We show that glutamatergic hippocampal neurons from mice lacking Munc13-1 form ultrastructurally normal synapses whose synaptic-vesicle cycle is arrested at the maturation step. Transmitter release from mutant synapses cannot be triggered by action potentials, calcium-ionophores or hypertonic sucrose solution. In contrast, release evoked by alpha-latrotoxin is indistinguishable from wild-type controls, indicating that the toxin can bypass Munc13-1-mediated vesicle maturation. A small subpopulation of synapses of any given glutamatergic neuron as well as all synapses of GABA (gamma-aminobutyric acid)-containing neurons are unaffected by Munc13-1 loss, demonstrating the existence of multiple and transmitter-specific synaptic vesicle maturation processes in synapses.     

Primary structure of human T-cell receptor alpha-chain

The T-cell receptor has been studied intensely over the past 10 years in an effort to understand the molecular basis for major histocompatibility complex (MHC) restricted antigen recognition. The use of anti-receptor monoclonal antibodies to isolate and characterize the receptor from human and murine T-cell clones has shown that the protein consists of two disulphide-linked glycopeptides, alpha and beta, distinct from known immunoglobulin light and heavy chains. Like immunoglobulin light and heavy chains, however, both the alpha- and beta-chains are composed of variable and constant regions. Molecular cloning has revealed that the beta-chain is evolutionarily related to immunoglobulins, and is encoded in separate V (variable), D (diversity), J (joining) and C (constant) segments that are rearranged in T cells to produce a functional gene. We report here cDNA clones encoding the alpha-chain of the receptor of the human T-cell leukaemia line HPB-MLT. Using these cDNA probes, we find that expression of alpha-chain mRNA and rearrangement of an alpha-chain V-gene segment occur only in T cells. The protein sequence predicted by these cDNAs is homologous to T-cell receptor beta-chains and to immunoglobulin heavy and light chains, particularly in the V and J segments.     

Chemische Kopplung von Hexokinase aus Hefe an Sephadex

Summary Yeast hexokinase was coupled to Sephadex by means of cyanogen bromide. The activity of the fixed enzyme was 2% of that before coupling. The activity of the noncoupled part of the enzyme was also decreased to the same degree. This decrease cannot be caused by the coupling reaction itself. The Michaelis-constants and the pH-activity curve of the coupled enzyme do not differ significantly from that of the free enzyme.     

Reverse self-splicing of group II intron RNAs in vitro.

Group II introns, which are classed together on the basis of a conserved secondary structure, are found in organellar genes of lower eukaryotes and plants. Like introns in nuclear pre-messenger RNA, they are excised by a two-step splicing reaction to generate branched circular RNAs, the so-called lariats. A remarkable feature of group II introns is their self-splicing activity in vitro. In the absence of a nucleotide cofactor, the intron RNAs catalyse two successive transesterification reactions which lead to autocatalytic excision of the lariat IVS from pre-mRNA and concomitantly to exon ligation. By virtue of its ability to specifically bind the 5' exon, the intron can also catalyse such reactions on exogenous RNA substrates. This sequence-specific attachment could enable group II introns to integrate into unrelated RNAs by reverse splicing, in a process similar to that described for the self-splicing Tetrahymena group I intron. Here we report that group II lariat IVS can indeed reintegrate itself into an RNA composed of the ligated exons in vitro. This occurs by a process of self-splicing that completely reverses both transesterification steps of the forward reaction: it involves a transition of the 2'-5' phosphodiester bond of the lariat RNA into the 3'-5' bond of the reconstituted 5' splice junction.     

Eight glacial cycles from an Antarctic ice core

The Antarctic Vostok ice core provided compelling evidence of the nature of climate, and of climate feedbacks, over the past 420,000 years. Marine records suggest that the amplitude of climate variability was smaller before that time, but such records are often poorly resolved. Moreover, it is not possible to infer the abundance of greenhouse gases in the atmosphere from marine records. Here we report the recovery of a deep ice core from Dome C, Antarctica, that provides a climate record for the past 740,000 years. For the four most recent glacial cycles, the data agree well with the record from Vostok. The earlier period, between 740,000 and 430,000 years ago, was characterized by less pronounced warmth in interglacial periods in Antarctica, but a higher proportion of each cycle was spent in the warm mode. The transition from glacial to interglacial conditions about 430,000 years ago (Termination V) resembles the transition into the present interglacial period in terms of the magnitude of change in temperatures and greenhouse gases, but there are significant differences in the patterns of change. The interglacial stage following Termination V was exceptionally long--28,000 years compared to, for example, the 12,000 years recorded so far in the present interglacial period. Given the similarities between this earlier warm period and today, our results may imply that without human intervention, a climate similar to the present one would extend well into the future.     

Fehlermöglichkeiten bei der Bestimmung von Metabolitgehalten in Hefezellen

Summary The exact assay of metabolites like NAD, ATP and other glycolytic substances in intact yeast cells greatly depends on the method of cell extraction. Reproducible and high levels can be obtained after prolonged incubation at high temperatures or by freezing and thawing of the denatureted cellular material in perchloric acid, thus eliminating possible errors due to different recoveries in the cell-free extracts.     

Testing the speed of 'spooky action at a distance'

Correlations are generally described by one of two mechanisms: either a first event influences a second one by sending information encoded in bosons or other physical carriers, or the correlated events have some common causes in their shared history. Quantum physics predicts an entirely different kind of cause for some correlations, named entanglement. This reveals itself in correlations that violate Bell inequalities (implying that they cannot be described by common causes) between space-like separated events (implying that they cannot be described by classical communication). Many Bell tests have been performed, and loopholes related to locality and detection have been closed in several independent experiments. It is still possible that a first event could influence a second, but the speed of this hypothetical influence (Einstein's 'spooky action at a distance') would need to be defined in some universal privileged reference frame and be greater than the speed of light. Here we put stringent experimental bounds on the speed of all such hypothetical influences. We performed a Bell test over more than 24 hours between two villages separated by 18 km and approximately east-west oriented, with the source located precisely in the middle. We continuously observed two-photon interferences well above the Bell inequality threshold. Taking advantage of the Earth's rotation, the configuration of our experiment allowed us to determine, for any hypothetically privileged frame, a lower bound for the speed of the influence. For example, if such a privileged reference frame exists and is such that the Earth's speed in this frame is less than 10(-3) times that of the speed of light, then the speed of the influence would have to exceed that of light by at least four orders of magnitude.     

Resident pulmonary lymphocytes expressing the gamma/delta T-cell receptor

The biological role of cells bearing the gamma delta T-cell antigen receptor (TCR) is as yet unclear. Although there are indications that some gamma delta+ cells can mediate cytotoxicity, their antigen-related functions have not yet been defined. In the mouse, gamma delta+ cells constitute 1-3% of T cells in lymphoid organs. Intestinal intraepithelial lymphocytes (IELs) and dendritic epidermal cells (DECs) also appear to carry the gamma delta TCR. The strategic locations of DECs and IELs have led to the suggestion that gamma delta+ cells could constitute a first line of defence in the vicinity of large surfaces of contact with the environment. We report here that an estimated 8-20% of resident pulmonary lymphocytes (RPLs) are CD3+ alpha beta TCR-, and presumably gamma delta TCR+. Furthermore, mice exposed to aerosols containing a Mycobacterium tuberculosis extract have an increased number of activated CD3+ alpha beta-TCR- pulmonary T cells which can be propagated in vitro.