Changes in Organization of the Endoplasmic Reticulum during Xenopus Oocyte Maturation and ActivationV⃞

The organization of the endoplasmic reticulum (ER) in the cortex of Xenopus oocytes was investigated during maturation and activation using a green fluorescent protein chimera, immunofluorescence, and electron microscopy. Dense clusters of ER developed on the vegetal side (the side opposite the meiotic spindle) during maturation. Small clusters appeared transiently at the time of nuclear envelope breakdown, disappeared at the time of first polar body formation, and then reappeared as larger clusters in mature eggs. The appearance of the large ER clusters was correlated with an increase in releaseability of Ca2+ by IP3. The clusters dispersed during the Ca2+ wave at activation. Possible relationships of ER structure and Ca2+ regulation are discussed.

Functional organization of the hippocampal memory system

In humans declarative or explicit memory is supported by the hippocampus and related structures of the medial temporal lobe working in concert with the cerebral cortex. This paper reviews our progress in developing an animal model for studies of cortical–hippocampal interactions in memory processing. Our findings support the view that the cortex maintains various forms of memory representation and that hippocampal structures extend the persistence and mediate the organization of these codings. Specifically, the parahippocampal region, through direct and reciprocal interconnections with the cortex, is sufficient to support the convergence and extended persistence of cortical codings. The hippocampus itself is critical to the organization cortical representations in terms of relationships among items in memory and in the flexible memory expression that is the hallmark of declarative memory.

The neural development and organization of letter recognition: Evidence from functional neuroimaging, computational modeling, and behavioral studies

Although much of the brain’s functional organization is genetically predetermined, it appears that some noninnate functions can come to depend on dedicated and segregated neural tissue. In this paper, we describe a series of experiments that have investigated the neural development and organization of one such noninnate function: letter recognition. Functional neuroimaging demonstrates that letter and digit recognition depend on different neural substrates in some literate adults. How could the processing of two stimulus categories that are distinguished solely by cultural conventions become segregated in the brain? One possibility is that correlation-based learning in the brain leads to a spatial organization in cortex that reflects the temporal and spatial clustering of letters with letters in the environment. Simulations confirm that environmental co-occurrence does indeed lead to spatial localization in a neural network that uses correlation-based learning. Furthermore, behavioral studies confirm one critical prediction of this co-occurrence hypothesis, namely, that subjects exposed to a visual environment in which letters and digits occur together rather than separately (postal workers who process letters and digits together in Canadian postal codes) do indeed show less behavioral evidence for segregated letter and digit processing.

Long-term change in the organization of inventive activity

Relying on a quantitative analysis of the patenting and assignment behavior of inventors, we highlight the evolution of institutions that encouraged trade in technology and a growing division of labor between those who invented new technologies and those who exploited them commercially over the nineteenth and early-twentieth centuries. At the heart of this change in the organization of inventive activity was a set of familiar developments which had significant consequences for the supply and demand of inventions. On the supply side, the growing complexity and capital intensity of technology raised the amount of human and physical capital required for effective invention, making it increasingly desirable for individuals involved in this activity to specialize. On the demand side, the growing competitiveness of product markets induced firms to purchase or otherwise obtain the rights to technologies developed by others. These increasing incentives to differentiate the task of invention from that of commercializing new technologies depended for their realization upon the development of markets and other types of organizational supports for trade in technology. The evidence suggests that the necessary institutions evolved first in those regions of the country where early patenting activity had already been concentrated. A self-reinforcing process whereby high rates of inventive activity encouraged the evolution of a market for technology, which in turn encouraged greater specialization and productivity at invention as individuals found it increasingly feasible to sell and license their discoveries, appears to have been operating. This market trade in technological information was an important contributor to the achievement of a high level of specialization at invention well before the rise of large-scale research laboratories in the twentieth century.

Comparative organization of cattle chromosome 5 revealed by comparative mapping by annotation and sequence similarity and radiation hybrid mapping

A whole genome cattle-hamster radiation hybrid cell panel was used to construct a map of 54 markers located on bovine chromosome 5 (BTA5). Of the 54 markers, 34 are microsatellites selected from the cattle linkage map and 20 are genes. Among the 20 mapped genes, 10 are new assignments that were made by using the comparative mapping by annotation and sequence similarity strategy. A LOD-3 radiation hybrid framework map consisting of 21 markers was constructed. The relatively low retention frequency of markers on this chromosome (19%) prevented unambiguous ordering of the other 33 markers. The length of the map is 398.7 cR, corresponding to a ratio of ≈2.8 cR5,000/cM. Type I genes were binned for comparison of gene order among cattle, humans, and mice. Multiple internal rearrangements within conserved syntenic groups were apparent upon comparison of gene order on BTA5 and HSA12 and HSA22. A similarly high number of rearrangements were observed between BTA5 and MMU6, MMU10, and MMU15. The detailed comparative map of BTA5 should facilitate identification of genes affecting economically important traits that have been mapped to this chromosome and should contribute to our understanding of mammalian chromosome evolution.

Molecular organization of the postsynaptic specialization

A specific set of molecules including glutamate receptors is targeted to the postsynaptic specialization of excitatory synapses in the brain, gathering in a structure known as the postsynaptic density (PSD). Synaptic targeting of glutamate receptors depends on interactions between the C-terminal tails of receptor subunits and specific PDZ domain-containing scaffold proteins in the PSD. These scaffold proteins assemble a specialized protein complex around each class of glutamate receptor that functions in signal transduction, cytoskeletal anchoring, and trafficking of the receptors. Among the glutamate receptor subtypes, the N-methyl-d-aspartate receptor is relatively stably integrated in the PSD, whereas the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor moves in and out of the postsynaptic membrane in highly dynamic fashion. The distinctive cell biological behaviors of N-methyl-d-aspartate and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors can be explained by their differential interactions with cytoplasmic proteins.

Extracellular Matrix Assembly in Diatoms (Bacillariophyceae)1 : III. Organization of Fucoglucuronogalactans within the Adhesive Stalks of Achnanthes longipes

Achnanthes longipes is a marine, biofouling diatom that adheres to surfaces via adhesive polymers extruded during motility or organized into structures called stalks that contain three distinct regions: the pad, shaft, and collar. Four monoclonal antibodies (AL.C1–AL.C4) and antibodies from two uncloned hybridomas (AL.E1 and AL.E2) were raised against the extracellular adhesives of A. longipes. Antibodies were screened against a hot-water-insoluble/hot-bicarbonate-soluble-fraction. The hot-water-insoluble/hot-bicarbonate-soluble fraction was fractionated to yield polymers in three size ranges: F1, ≥ 20,000,000 Mr; F2, ≅100,000 Mr; and F3, <10,000 Mr relative to dextran standards. The ≅100,000-Mr fraction consisted of highly sulfated (approximately 11%) fucoglucuronogalactans (FGGs) and low-sulfate (approximately 2%) FGGs, whereas F1 was composed of O-linked FGG (F2)-polypeptide (F3) complexes. AL.C1, AL.C2, AL.C4, AL.E1, and AL.E2 recognized carbohydrate complementary regions on FGGs, with antigenicity dependent on fucosyl-containing side chains. AL.C3 was unique in that it had a lower affinity for FGGs and did not label any portion of the shaft. Enzyme-linked immunosorbent assay and immunocytochemistry indicated that low-sulfate FGGs are expelled from pores surrounding the raphe terminus, creating the cylindrical outer layers of the shaft, and that highly sulfated FGGs are extruded from the raphe, forming the central core. Antibody-labeling patterns and other evidence indicated that the shaft central-core region is related to material exuded from the raphe during cell motility.

Modular organization of intrinsic connections associated with spectral tuning in cat auditory cortex

Many response properties in primary auditory cortex (AI) are segregated spatially and organized topographically as those in primary visual cortex. Intensive study has not revealed an intrinsic, anatomical organizing principle related to an AI functional topography. We used retrograde anatomic tracing and topographic physiologic mapping of acoustic response properties to reveal long-range (≥1.5 mm) convergent intrinsic horizontal connections between AI subregions with similar bandwidth and characteristic frequency selectivity. This suggests a modular organization for processing spectral bandwidth in AI.

High-resolution mapping of nucleoprotein complexes by site-specific protein-DNA photocrosslinking: organization of the human TBP-TFIIA-TFIIB-DNA quaternary complex.

We have used a novel site-specific protein-DNA photocrosslinking procedure to define the positions of polypeptide chains relative to promoter DNA in binary, ternary, and quaternary complexes containing human TATA-binding protein, human or yeast transcription factor IIA (TFIIA), human transcription factor IIB (TFIIB), and promoter DNA. The results indicate that TFIIA and TFIIB make more extensive interactions with promoter DNA than previously anticipated. TATA-binding protein, TFIIA, and TFIIB surround promoter DNA for two turns of DNA helix and thus may form a "cylindrical clamp" effectively topologically linked to promoter DNA. Our results have implications for the energetics, DNA-sequence-specificity, and pathway of assembly of eukaryotic transcription complexes.

Modular cis-regulatory organization of developmentally expressed genes: two genes transcribed territorially in the sea urchin embryo, and additional examples.

The cis-regulatory systems that control developmental expression of two sea urchin genes have been subjected to detailed functional analysis. Both systems are modular in organization: specific, separable fragments of the cis-regulatory DNA each containing multiple transcription factor target sites execute particular regulatory subfunctions when associated with reporter genes and introduced into the embryo. The studies summarized here were carried out on the CyIIIa gene, expressed in the embryonic aboral ectoderm and on the Endo16 gene, expressed in the embryonic vegetal plate, archenteron, and then midgut. The regulatory systems of both genes include modules that control particular aspects of temporal and spatial expression, and in both the territorial boundaries of expression depend on a combination of negative and positive functions. In both genes different regulatory modules control early and late embryonic expression. Modular cis-regulatory organization is widespread in developmentally regulated genes, and we present a tabular summary that includes many examples from mouse and Drosophila. We regard cis-regulatory modules as units of developmental transcription control, and also of evolution, in the assembly of transcription control systems.

The physical and genomic organization of microsatellites in sugar beet.

Microsatellites, tandem arrays of short (2-5 bp) nucleotide motifs, are present in high numbers in most eukaryotic genomes. We have characterized the physical distribution of microsatellites on chromosomes of sugar beet (Beta vulgaris L.). Each microsatellite sequence shows a characteristic genomic distribution and motif-dependent dispersion, with site-specific amplification on one to seven pairs of centromeres or intercalary chromosomal regions and weaker, dispersed hybridization along chromosomes. Exclusion of some microsatellites from 18S-5.8S-25S rRNA gene sites, centromeres, and intercalary sites was observed. In-gel and in situ hybridization patterns are correlated, with highly repeated restriction fragments indicating major centromeric sites of microsatellite arrays. The results have implications for genome evolution and the suitability of particular microsatellite markers for genetic mapping and genome analysis.

The organization of seismicity on fault networks.

Although models of homogeneous faults develop seismicity that has a Gutenberg-Richter distribution, this is only a transient state that is followed by events that are strongly influenced by the nature of the boundaries. Models with geometrical inhomogeneities of fracture thresholds can limit the sizes of earthquakes but now favor the characteristic earthquake model for large earthquakes. The character of the seismicity is extremely sensitive to distributions of inhomogeneities, suggesting that statistical rules for large earthquakes in one region may not be applicable to large earthquakes in another region. Model simulations on simple networks of faults with inhomogeneities of threshold develop episodes of lacunarity on all members of the network. There is no validity to the popular assumption that the average rate of slip on individual faults is a constant. Intermediate term precursory activity such as local quiescence and increases in intermediate-magnitude activity at long range are simulated well by the assumption that strong weakening of faults by injection of fluids and weakening of asperities on inhomogeneous models of fault networks is the dominant process; the heat flow paradox, the orientation of the stress field, and the low average stress drop in some earthquakes are understood in terms of the asperity model of inhomogeneous faulting.

Human plectin: organization of the gene, sequence analysis, and chromosome localization (8q24).

Plectin, a 500-kDa intermediate filament binding protein, has been proposed to provide mechanical strength to cells and tissues by acting as a cross-linking element of the cytoskeleton. To set the basis for future studies on gene regulation, tissue-specific expression, and pathological conditions involving this protein, we have cloned the human plectin gene, determined its coding sequence, and established its genomic organization. The coding sequence contains 32 exons that extend over 32 kb of the human genome. Most of the introns reside within a region encoding the globular N-terminal domain of the molecule, whereas the entire central rod domain and the entire C-terminal globular domain were found to be encoded by single exons of remarkable length, >3 kb and >6 kb, respectively. Overall, the organization of the human plectin gene was strikingly similar to that of human bullous pemphigoid antigen 1 (BPAG1), confirming that both proteins belong to the same gene family. Comparison of the deduced protein sequences for human and rat plectin revealed that they were 93% identical. By using fluorescence in situ hybridization, we have mapped the plectin gene to the long arm of chromosome 8 within the telomeric region. This gene locus (8q24) has previously been implicated in the human blistering skin disease epidermolysis bullosa simplex Ogna. Detailed knowledge of the structure of the plectin gene and its chromosome localization will aid in the elucidation of whether this or any other pathological conditions are linked to alterations in the plectin gene.

The genomic and physical organization of Ty1-copia-like sequences as a component of large genomes in Pinus elliottii var. elliottii and other gymnosperms.

A DNA sequence, TPE1, representing the internal domain of a Ty1-copia retroelement, was isolated from genomic DNA of Pinus elliottii Engelm. var. elliottii (slash pine). Genomic Southern analysis showed that this sequence, carrying partial reverse transcriptase and integrase gene sequences, is highly amplified within the genome of slash pine and part of a dispersed element >4.8 kbp. Fluorescent in situ hybridization to metaphase chromosomes shows that the element is relatively uniformly dispersed over all 12 chromosome pairs and is highly abundant in the genome. It is largely excluded from centromeric regions and intercalary chromosomal sites representing the 18S-5.8S-25S rRNA genes. Southern hybridization with specific DNA probes for the reverse transcriptase gene shows that TPE1 represents a large subgroup of heterogeneous Ty1-copia retrotransposons in Pinus species. Because no TPE1 transcription could be detected, it is most likely an inactive element--at least in needle tissue. Further evidence for inactivity was found in recombinant reverse transcriptase and integrase sequences. The distribution of TPE1 within different gymnosperms that contain Ty1-copia group retrotransposons, as shown by a PCR assay, was investigated by Southern hybridization. The TPE1 family is highly amplified and conserved in all Pinus species analyzed, showing a similar genomic organization in the three- and five-needle pine species investigated. It is also present in spruce, bald cypress (swamp cypress), and in gingko but in fewer copies and a different genomic organization.

Molecular organization of histidine-tagged biomolecules at self-assembled lipid interfaces using a novel class of chelator lipids.

In molecular biology, the expression of fusion proteins is a very useful and well-established technique for the identification and one-step purification of gene products. Even a short fused sequence of five or six histidines enables proteins to bind to an immobilized metal ion chelate complex. By synthesis of a class of chelator lipids, we have transferred this approach to the concept of self-assembly. The specific interaction and lateral organization of a fluorescent fusion molecule containing a C-terminal oligohistidine sequence was studied by film balance techniques in combination with epifluorescence microscopy. Due to the phase behavior of the various lipid mixtures used, the chelator lipids can be laterally structured, generating two-dimensional arrays of histidine-tagged biomolecules. Because of the large variety of fusion proteins already available, this concept represents a powerful technique for orientation and organization of proteins at lipid interfaces with applications in biosensing, biofunctionalization of nanostructured interfaces, two-dimensional crystallization, and studies of lipid-anchored proteins.