Structural disorder serves as a weak signal for intracellular protein degradation.

Targeted turnover of proteins is a key element in the regulation of practically all basic cellular processes. The underlying physicochemical and/or sequential signals, however, are not fully understood. This issue is particularly pertinent in light of the recent recognition that intrinsically unstructured/disordered proteins, common in eukaryotic cells, are extremely susceptible to proteolytic degradation in vitro. The in vivo half-lives of proteins were determined recently in a high-throughput study encompassing the entire yeast proteome; here we examine whether these half-lives correlate with the presence of classical degradation motifs (PEST region, destruction-box, KEN-box, or the N-terminal residue) or with various physicochemical characteristics, such as the size of the protein, the degree of structural disorder, or the presence of low-complexity regions. Our principal finding is that, in general, the half-life of a protein does not depend on the presence of degradation signals within its sequence, even of ubiquitination sites, but correlates mainly with the length of its polypeptide chain and with various measures of structural disorder. Two distinct modes of involvement of disorder in degradation are proposed. Susceptibility to degradation of longer proteins, containing larger numbers of residues in conformational disorder, suggests an extensive function, whereby the effect of disorder can be ascribed to its mere physical presence. However, after normalization for protein length, the only signal that correlates with half-life is disorder, which indicates that it also acts in an intensive manner, that is, as a specific signal, perhaps in conjunction with the recognition of classical degradation motifs. The significance of correlation is rather low; thus protein degradation is not determined by a single characteristic, but is a multi-factorial process that shows large protein-to-protein variations. Protein disorder, nevertheless, plays a key signalling role in many cases.

SPINE bioinformatics and data-management aspects of high-throughput structural biology.

SPINE (Structural Proteomics In Europe) was established in 2002 as an integrated research project to develop new methods and technologies for high-throughput structural biology. Development areas were broken down into workpackages and this article gives an overview of ongoing activity in the bioinformatics workpackage. Developments cover target selection, target registration, wet and dry laboratory data management and structure annotation as they pertain to high-throughput studies. Some individual projects and developments are discussed in detail, while those that are covered elsewhere in this issue are treated more briefly. In particular, this overview focuses on the infrastructure of the software that allows the experimentalist to move projects through different areas that are crucial to high-throughput studies, leading to the collation of large data sets which are managed and eventually archived and/or deposited.

AutoDep: a web-based system for deposition and validation of macromolecular structural -information.

This paper describes the design and full implementation of a new concept in data deposition and validation: AutoDep (copyright Brookhaven Science Associates LLC). AutoDep changes the traditional procedure for data acceptance and validation of the primary databases into an interactive depositor-driven operation which almost eliminates the delay between the acceptance of the data and its public release. The system takes full advantage of the knowledge and expertise of the experimenters, rather than relying on the database curators for the complete and accurate description of the structural experiment and its results. AutoDep, developed by the Protein Data Bank at Brookhaven National Laboratory (BNL) as a flexible and portable system, has already been adopted by other primary databases and implemented on different platforms/operating systems. AutoDep was introduced at BNL in 1996 [see Manning (1996), Protein Data Bank Quart. Newslett. 77, 2 (ftp://ftp.rcsb. org/pub/pdb/doc/newsletters/bnl/newsletter96jul/newslttr+ ++.txt); Manning (1996), Protein Data Bank Quart. Newslett. 78, 2 (ftp://ftp. rcsb.org/pub/pdb/doc/newsletters/bnl/newsletter96oct/+ ++newslttr.txt)].

Automated analysis of interatomic contacts in proteins.

MOTIVATION: New software has been designed to assist the molecular biologist in understanding the structural consequences of modifying a ligand and/or protein. RESULTS: Tools are described for the analysis of ligand-protein contacts (LPC software) and contacts of structural units (CSU software) such as helices, sheets, strands and residues. Our approach is based on a detailed analysis of interatomic contacts and interface complementarity. For any ligand or structural unit, these software automatically: (i) calculate the solvent-accessible surface of every atom; (ii) determine the contacting residues and type of interaction they undergo (hydrophobic-hydrophobic, aromatic-aromatic, etc.); (iii) indicate all putative hydrogen bonds. LPC software further predicts changes in binding strength following chemical modification of the ligand. AVAILABILITY: Both LPC and CSU can be accessed through the PDB and are integrated in the 3DB Atlas page of all PDB files. For any given file, the tools can also be accessed at http://www.pdb.bnl. gov/pdb-bin/lpc?PDB_ID= and http://www.pdb.bnl. gov/pdb-bin/csu?PDB_ID= with the four-letter PDB code added at the end in each case. Finally, LPC and CSU can be accessed at: http://sgedg.weizmann.ac.il/lpc and http://sgedg.weizmann.ac.il/csu.

The protein data bank. Bridging the gap between the sequence and 3D structure world.

The protein data bank (PDB), at Brookhaven National Laboratory, is a database containing information on experimentally determined three-dimensional structures of proteins, nucleic acids, and other biological macromolecules, with approximately 9000 entries. The PDB has a 27-year history of service to a global community of researchers, educators, and students in a wide variety of scientific disciplines. Data are easily submitted via PDB's WWW-based tool AutoDep, in either PDB or mmCIF format, and are most conveniently examined via PDB's WWW-based tool 3DB Browser. Collaborative centers have been, and continue to be, established worldwide to assist in data deposition, archiving, and distribution.

GeneCards: a novel functional genomics compendium with automated data mining and query reformulation support.

MOTIVATION: Modern biology is shifting from the 'one gene one postdoc' approach to genomic analyses that include the simultaneous monitoring of thousands of genes. The importance of efficient access to concise and integrated biomedical information to support data analysis and decision making is therefore increasing rapidly, in both academic and industrial research. However, knowledge discovery in the widely scattered resources relevant for biomedical research is often a cumbersome and non-trivial task, one that requires a significant amount of training and effort. RESULTS: To develop a model for a new type of topic-specific overview resource that provides efficient access to distributed information, we designed a database called 'GeneCards'. It is a freely accessible Web resource that offers one hypertext 'card' for each of the more than 7000 human genes that currently have an approved gene symbol published by the HUGO/GDB nomenclature committee. The presented information aims at giving immediate insight into current knowledge about the respective gene, including a focus on its functions in health and disease. It is compiled by Perl scripts that automatically extract relevant information from several databases, including SWISS-PROT, OMIM, Genatlas and GDB. Analyses of the interactions of users with the Web interface of GeneCards triggered development of easy-to-scan displays optimized for human browsing. Also, we developed algorithms that offer 'ready-to-click' query reformulation support, to facilitate information retrieval and exploration. Many of the long-term users turn to GeneCards to quickly access information about the function of very large sets of genes, for example in the realm of large-scale expression studies using 'DNA chip' technology or two-dimensional protein electrophoresis. AVAILABILITY: Freely available at http://bioinformatics.weizmann.ac.il/cards/ CONTACT: cards@bioinformatics.weizmann.ac.il

Protein Data Bank (PDB): database of three-dimensional structural information of biological macromolecules.

The Protein Data Bank (PDB) at Brookhaven National Laboratory, is a database containing experimentally determined three-dimensional structures of proteins, nucleic acids and other biological macromolecules, with approximately 8000 entries. Data are easily submitted via PDB's WWW-based tool AutoDep, in either mmCIF or PDB format, and are most conveniently examined via PDB's WWW-based tool 3DB Browser.

Rapid access to biomedical knowledge with GeneCards and HotMolecBase: implications for the electrophoretic analysis of large sets of gene products.

Rapid access to well-organized information about gene products is important for many studies that simultaneously monitor large sets of those factors, for example with electrophoretic methods. HotMolecBase and GeneCards, Internet resources that may be accessed from our Bioinformatics homepage at http://bioinfo.weizmann.ac.il/, have been designed to address similar problems. GeneCards presents semi-automatically collected information about all approved human genes and their products (with a focus on cellular functions and medical aspects), and offers a new kind of knowledge navigation guidance system that interactively guides the information-seeking scientist to relevant information. On the other hand, HotMolecBase is a collection of more extensive hypertext fact sheets about a small set of medically interesting molecules (mainly proteins) that are regarded as especially promising targets for drug development. Together, both resources may help scientists world-wide to find their way in the growing labyrinth of biomedical information on the World Wide Web. In the present article, we want to explain how these resources may be used by researchers who want to access information related to particular spots on two-dimensional electrophoresis gels.

Computers in biology: the future is today.

Which are the applications of microcomputers in biology today? Some areas where the work with microcomputers is becoming increasingly important are considered: laboratory applications, event simulation and word processing. Almost all laboratory computer applications can be described as one of the following functions: 1. control of experiments, including timing and synchronizing external voltages; 2. data acquisition, usually through digital conversion of analog electrical signals; 3. data storage and, 4. data analysis. Event simulation is considered both as a research tool and as an important element in the educational area. Word processing and the automatic creation of literature references lists is considered as an ignored role of the microcomputers in the laboratory field. What about the influence of biology in computer technology? As specialized magazines say, many laboratories of biotechnologist are working hard to build a molecular computer. That is, an artificially designed ultramicroscopic machine built of proteins, nucleic acids, metals and non-metals in a planned arrangement. And this is not the end. The latest application able to expand our horizon in the biological field may be starting to be used at this moment.

A dual modulatory effect of progesterone on the LHRH-induced LH release.

The role of progesterone on the release of LH induced by 25 or 50 ng of LHRH was studied in proestrus rats in which spontaneous preovulatory release of LH was prevented by sodium pentobarbitone. After the s.c. administration of progesterone (5 mg) at 18.00 h of diestrus day 2 or at 12.00 h of proestrus, serum LH was not detectable at 17.15 h of proestrus. Injections of 25 or 50 ng of LHRH at 17.00 h of proestrus induced a dose response release of LH 15 min after. However, the LH response to LHRH administration increased significantly when progesterone was injected at 12.00 h of proestrus. The potentiating effect of progesterone seems to be exerted at pituitary level. The effect of LHRH and the enhanced response of the pituitary after progesterone treatment was prevented by the administration of the antiestrogen Tamoxifen in diestrus day 2. The release of LH induced by 50 ng of LHRH on proestrus day was blocked by the previous injection of progesterone on diestrus day 2. The inhibition was maintained even though a second dose of progesterone was given at 12.00 h of proestrus. The simultaneous administration of estrogen and progesterone on diestrus day 2 did not prevent the inhibitory effect of progesterone. It is concluded that the facilitatory or inhibitory effect of progesterone on the release of LH induced by LHRH is dependent upon the previous sensitization of the pituitary to estrogen.

Participation of the hippocampus in the facilitatory effect of an exteroceptive stimulus on milk ejection.

The auditory exteroceptive stimulus emanating from a lactating rat and the litter while suckling was used to facilitate milk ejection in another "induced" mother suckled by her own litter. Sectioning of the tractus corticohypothalamicus medialis (TCM) or the columns of the fornix, prevented the facilitatory effect of the exteroceptive stimulus on milk ejection. Sham operated induced mothers gave significantly more milk than noninduced rats. Electrolytic lesioning of the stria terminalis did not affect normal milk ejection nor the response to the exteroceptive stimulus. It is proposed that the hippocampus through the TCM may facilitate the suckling-induced milk ejection when an appropiate exteroceptive stimulus is applied. Neither the lesion of the stria terminalis nor the section of the fornix or the TCM seems to alter the normal suckling-induced milk ejection. A general modulatory role of the limbic system is described.

Inhibition of milk ejection by a visual stimulus in lactating rats: implication of the pineal gland.

The effect of visual and auditory stimuli on milk ejection during suckling was studied in normal and pinealectomized lactating rats. The photic and auditory stimuli were applied to each mother for 10 s every 20 s during the 30 min suckling period. Both stimuli inhibited milk ejection without altering the nursing behavior. In mothers kept in complete darkness or in which the visual stimulus shone continuously during the suckling period, milk ejection was not affected. The inhibition of milk ejection is therefore produced by the light on-off sequence. In lactating rats exposed to the stimulus during 3 consecutive days, a significant inhibition of milk ejection was obtained each day. A normal milk-ejection response occurred in both non-stimulated pinealectomized and in stimulated pinealectomized lactating rats. Pinealectomy did not prevent the inhibitory effect of the sound stimulus. Treatment with methysergide prevented the inhibition of milk ejection induced by the visual stimulus but did not prevent the inhibitory effect of the auditory stimulus. It seems that the pineal gland mediates an inhibitory visual reflex acting on oxytocin release and milk ejection.

Responses of supraoptic neurones in the intact and deafferented rat hypothalamus to injections of hypertonic sodium chloride.

1. Recordings were made from a total of fifty-three neurones in the supraoptic nuclei of four groups of rats: intact rats, animals in which the hypothalamus had been partly denervated by anteriorly or posteriorly placed semicircular cuts, and rats with a totally deafferented hypothalamus. 2. When first encountered, cells from intact animals fired at a mean rate of 5.08 +/- 0.78 spikes/sec, those from posteriorly isolated hypothalami at 3.93 +/- 0.63 spikes/sec, those from the anteriorly isolated hypothalami at 2.05 +/- 0.83 spikes/sec, and those from totally isolated hypothalami at 0.99 +/- 0.46 spikes/sec. 3. When stimulated osmotically by an intraperitoneal injection of ml. 1.5 M-NaCl, eight out of eight cells in intact rats showed a significant increase in firing rate between 20 and 30 min after the injection. Six out of nine cells in posteriorly isolated hypothalami showed significant but smaller responses. No increase in firing rate could be detected in seven cells from totally isolated hypothalami or from eight cells in hypothalami partly isolated by anterior cuts. 4. The results imply that under the conditions of these experiments by the spontaneous activity of the supraoptic nucleus in intact animals was maintained by an extrahypothalamic excitatory input, that partial hypothalamic isolation reduced its intensity, possibly by unmasking an inhibitory input, and that total isolation reduced it to an even greater extent. Osmotic activation of supraoptic cells was only possible when the anterior connexions of the hypothalamus were intact. Thus the cerebral osmo-receptors for vasopressin release may be situated outside the supraoptic nuclei.