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Biomedical subjects

S Krauss

Publications and source records attributed to S Krauss.

At least 19 recordsLinked to original sources

Origin and molecular changes associated with emergence of a highly pathogenic H5N2 influenza virus in Mexico.

In October of 1993, there was decreased egg production and increased mortality among Mexican chickens, in association with serologic evidence of an H5N2 influenza virus. First isolated from chickens in May of 1994, after spreading widely in the country, the virus caused only a mild respiratory syndrome in specific pathogen-free chickens. Because eradication of the virus by destruction of infected birds posed major obstacles to the poultry industry in Mexico, we were able to conduct a "field experiment" to determine the fate of an avirulent virus after repeated cycles of replication in millions of chickens. By the end of 1994, the virus had mutated to contain a highly cleavable hemagglutinin (HA), but remained only mildly pathogenic in chickens. Within months, however, it had become lethal in poultry. Nucleotide sequence analysis of the HA cleavage site of the original avirulent strain revealed R-E-T-R, typical of avirulent viruses and unlike the K-K-K-R sequence characterizing viruses responsible for the 1983 outbreak in poultry in the United States. Both mildly and highly pathogenic isolates contained insertions and a substitution of basic residues in the HA connecting peptide, R-K-R-K-T-R, which made the HA highly cleavable in trypsin-free chicken embryo fibroblasts. Phylogenetic analysis of the HA of H5 avian influenza viruses, including the Mexican isolates, indicated that the epidemic virus had originated from the introduction of a single virus of the North American lineage into Mexican chickens. This sequence of events demonstrates, apparently for the first time, the stepwise acquisition of virulence by an avian influenza virus in nature.

Amino Acid Sequence

Recombinant CD4-IgE, a novel hybrid molecule, inducing basophils to respond to human immunodeficiency virus (HIV) and HIV-infected target cells.

Basophils and mast cells, as the main effector cells in IgE-mediated type I hypersensitivity, are involved in the elimination of parasites and, according to recent findings, may also play an important role in the defense against bacterial and viral infections. Using a genetic engineering approach we wanted to redirect this potent IgE-mediated defense system against intruding human immune deficiency virus. We constructed a recombinant CD4-IgE molecule, consisting of the two N-terminal domains of CD4 and the CH2-4 domains of the IgE heavy chain, thus providing the IgE with specificity for the gp120 of human immunodeficiency virus (HIV). The binding properties of hybrid CD4-IgE to the high-affinity receptor for IgE (Fc epsilon RI) on basophils as well as to the low-affinity receptor (Fc epsilon RII or CD23) for IgE on lymphoid cells were found to be similar to those of native IgE. At the same time, the CD4 domains of the recombinant molecule retained the gp120 binding specificity with an affinity similar to that of the native CD4. By functional tests, we demonstrated that CD4-IgE armed basophils can be triggered by free HIV and by HIV-infected cells to release their mediators. We further show that HIV-triggered basophils lead to a decreased replication of HIV in susceptible T cells. We, therefore, conclude that the type I hypersensitivity effector cells can be engaged in the elimination of HIV-infected cells, at least in vitro. Because of the strong binding of the CD4-IgE construct to the Fc epsilon RI, we assume that CD4-IgE has a short t1/2 in serum, but may similarly to IgE exhibit prolonged resident time on basophils and mast cells, which are located close to mucosal surfaces or in the connective tissue. Thus CD4-IgE could play an important role in the elimination of HIV also in vivo.

Basophils

cDNA sequence of zebrafish (Brachydanio rerio) translation elongation factor-1 alpha: molecular phylogeny of eukaryotes based on elongation factor-1 alpha protein sequences.

We have isolated and determined the nucleotide sequence of a cDNA clone containing the complete coding region for elongation factor-1 alpha (EF-1 alpha) from an embryonic zebrafish cDNA library. A secondary structure model based on all known EF-1 alpha and EF-Tu protein sequences is presented and the presence of conserved putative protein kinase C phosphorylation sites in loop regions of eukaryotic EF-1 alpha is demonstrated. Using distance matrix and maximum parsimony methods we constructed multi-kingdom phylogenetic trees containing 22 different eukaryotic sequences. Strikingly, both tree constructions show Fungi to be the closest relative of Animalia among eukaryotic kingdoms. A 12 amino acid stretch present in all animal and fungal sequences known to date was found to be absent from all plant, protist an archaebacterial EF-1 alpha sequences suggesting that this sequence was inserted following the separation of plants from the lineage leading to fungi and animals. In contrast to our results, molecular phylogenies based on small subunit ribosomal RNA sequences as well as other protein sequences have failed to yield consistent results regarding the branching order among the kingdoms Plantae, Fungi and Animalia. The slow evolutionary rate and universal occurrence of EF-1 alpha (EF-Tu in eubacteria) makes this protein a particularly interesting tool for probing distant evolutionary relationships.

Animals

Heat-loss response to a thermal challenge in seasonal affective disorder.

This study extends earlier findings of poorly facilitated postexercise heat loss during the winter in seasonal affective disorder (SAD). While depressed in the winter, 19 SAD subjects exhibited a significantly impaired postexercise heat loss relative to 10 control subjects. During the summer while euthymic, SAD subjects did not significantly differ from control subjects in postexercise heat loss. Since thermoregulatory heat loss is a highly dopamine-dependent process, these results support earlier findings of poorly facilitated dopamine availability in SAD during the winter and suggest a centrally mediated effect of light in SAD.

Adult

Regulatory gene expression boundaries demarcate sites of neuronal differentiation in the embryonic zebrafish forebrain.

During development of the zebrafish forebrain, a simple scaffold of axon pathways is pioneered by a small number of neurons. We show that boundaries of expression domains of members of the eph, forkhead, pax, and wnt gene families correlate with the positions at which these neurons differentiate and extend axons. Analysis of genetically or experimentally altered forebrains indicates that if a boundary is maintained, there is appropriate neural differentiation with respect to the boundary. Conversely, in the absence of a boundary, there is concomitant disruption of neural patterning. We also show that a strip of cells within the dorsal diencephalon shares features with ventral midline cells. This strip of cells fails to develop in mutant fish in which specification of the ventral CNS is disrupted, suggesting that its development may be regulated by the same inductive pathways that pattern the ventral midline.

Animals

The hedgehog gene family in Drosophila and vertebrate development.

The segment polarity gene hedgehog plays a central role in cell patterning during embryonic and post-embryonic development of the dipteran, Drosophila melanogaster. Recent studies have identified a family of hedgehog related genes in vertebrates; one of these, Sonic hedgehog is implicated in positional signalling processes that show interesting similarities with those controlled by its Drosophila homologue.

Amino Acid Sequence

A functionally conserved homolog of the Drosophila segment polarity gene hh is expressed in tissues with polarizing activity in zebrafish embryos.

The segment polarity gene hedgehog (hh) encodes a novel signaling protein that mediates local cell-cell interactions in the developing Drosophila embryo. Here we describe the existence of an hh-related gene family in the zebrafish, Brachydanio rerio. One of these genes, sonic hedgehog (shh), is expressed in the notochord, floor plate, and posterior fin mesoderm, tissues associated with polarizing activities in various vertebrate embryos. The pattern of shh expression in zebra-fish mutants affecting axial structures, together with the consequences of its ectopic expression in normal embryos, is consistent with a role for shh in floor plate induction. By expressing shh in transgenic Drosophila embryos, we also demonstrate a strong functional conservation between the fish and fly hh genes.

Amino Acid Sequence

Zebrafish pou[c]: a divergent POU family gene ubiquitously expressed during embryogenesis.

We report the isolation and characterization of cDNA for a novel zebrafish (Brachyodanio rerio) POU domain gene, pou[c], which is ubiquitously expressed during embryonic development. This gene encodes a 610 amino acids long protein with a 149 amino acid POU domain ending only 8 residues before the C terminus. The 453 amino acids long region N-terminal to the POU domain contains several features typical of transcriptional activation domains such as an acidic region with a putative amphipathic alpha-helix, a glutamine-rich region, and short threonine- and/or serine-rich regions. Comparison of the POU domain of pou[c] to other known POU sequences clearly show that pou[c] has the most divergent POU domain sequence reported to date. Thus, we suggest that pou[c] should be placed as the presently sole member of a new, sixth class of POU proteins. DNA-binding studies revealed that pou[c] is not an octamer-binding transcription factor like the Oct proteins described from mammals, chicken and Xenopus. Rather, pou[c] binds with high affinity to the TAATGARAT motif found in the promoters of the herpes simplex virus immediate early genes and to degenerate octamer-TAATGA motifs. Circular permutation analyses also show that pou[c] induces DNA bending upon sequence-specific binding.

Amino Acid Sequence

Genomic sequence and embryonic expression of the zebrafish homeobox gene hox-3.4.

We have analyzed the genomic DNA sequence and embryonic expression pattern of the zebrafish hox-3.4 gene. Two exons, encoding a protein with a total size of 232 amino acids, were identified within a 3.5 kbp genomic region. Besides the homeodomain, which is identical to that of the human HOX3D and mouse Hox-3.4 genes, the first 58 residues of the N-terminal domain in the predicted Hox-3.4 protein share 48% sequence identity with the gene product of the human cognate. Some of the N-terminal sequence elements are also conserved relative to the two other members of the Hox-1.3/Hox-2.1/Hox-3.4 paralogy group. In addition, the paralogous genes share a significant degree of sequence identity in non-coding regions. This conservation is particularly evident in the promoter regions of the cognates hox-3.4, Hox-3.4, and HOX3D, where a 180 bp TATA-box-containing element with a 60% identity is located. This is in agreement with the previous finding that the HOX3D promoter region contains response elements for other Hox proteins and retinoids. Also with respect to embryonic expression, the zebrafish hox-3.4 gene is very similar to its mammalian counterparts. Within the central nervous system of 16, 24, and 48 h embryos, hox-3.4 transcripts were detected throughout most of the spinal cord from a boundary at the posterior end of the hindbrain. In 16 h embryos the hox-3.4 gene is also active within a restricted region of the tailbud.

Amino Acid Sequence

The monoclonal CD4 antibody M-T413 inhibits cellular infection with human immunodeficiency virus after viral attachment to the cell membrane: an approach to postexposure prophylaxis.

Infectious cellular uptake of human immunodeficiency virus (HIV) is initiated by a complex sequence of interactions between the viral envelope gp120/gp41 complex and the cellular CD4 receptor resulting in the exposure of a hydrophobic region of gp41 that mediates the irreversible fusion of the virus with the cell membrane. Here we show that viral penetration into a susceptible cell can be inhibited by the high-affinity monoclonal CD4 antibody (CD4 mAb) M-T413 even when it is added as late as 30-120 min after the initial contact of virus with the cell membrane. Inhibition of infection was assessed by monitoring cultures for 34 days after exposure to virus using four different methods simultaneously, including detection of viral DNA by PCR. The interval during which HIV remains sensitive to postbinding neutralization by CD4 mAb depends on strain of virus and type of target cell. Preparations of recombinant soluble CD4 (and the immunoadhesin CD4-IgG1) were much less efficient when compared with mAb M-T413, particularly in blocking infection by fresh HIV-1 isolates. Also cellular transmission of HIV, as determined by syncytia formation within 24 hr, was prevented by mAb M-T413 when added within 45 min of contact of infected H9 cells with uninfected C8166 cells. Together with the favorable clinical experience obtained with CD4 mAbs as immunomodulatory drugs, these data suggest that infusion of CD4 mAb M-T413 may be a therapeutic modus for immediate prophylactic intervention after occupational exposure to HIV and for prevention of intrapartum mother-to-infant HIV transmission.

Animals

Zebrafish pax[b] is involved in the formation of the midbrain-hindbrain boundary.

Among the genes thought to be involved in patterning the nervous system are a family of developmentally regulated paired box-containing (Pax) genes. Mutations in some of these Pax genes lead to severe developmental abnormalities. Zebrafish pax[b](pax[zf-b]) is a member of the Pax gene family that is expressed in the presumptive posterior midbrain from the end of gastrulation and, at later stages, in other localized regions of the developing embryo. Here we show that injection of antibodies raised against the pax[b] protein causes a localized malformation at the midbrain-hindbrain boundary. In situ hybridizations demonstrate that antibody injection causes downregulation of pax[b] transcripts in the posterior midbrain and alteration of wnt-1 and eng-2 expression in this area. The data demonstrate an involvement of pax[b] in the formation of the midbrain-hindbrain junction.

Animals

The paired domain-containing nuclear factor pax[b] is expressed in specific commissural interneurons in zebrafish embryos.

The zebrafish paired box (Pax) genes are expressed in the early neural tube and are thought to be transcription factors that regulate the differentiation of cells in the central nervous system (CNS). The protein product of one of these Pax genes, pax[b], is detectable as a nuclear antigen in all the regions of the embryo that transcribe the gene including the posterior midbrain, the nephritic primordium, the Wolffian duct, the optic stalk, and, in specific neurons, in the hindbrain and spinal cord. The timing and pattern of axonal outgrowth by the early pax[b]-positive neurons suggest that they are the commissural secondary ascending (CoSA) interneurons in the spinal cord; the primary commissural interneurons (MiD2c and MiD3c) in hindbrain rhombomeres mi2 and mi3; and a previously unclassified set of commissural interneurons that we termed the commissural caudalrhombomere ascending (CoCaA) interneurons in the caudal hindbrain. In contrast, the Mauthner interneurons do not express pax[b] early in development. Shortly after the appearance of the first pax[b]-positive interneurons, additional nuclei adjacent to the first pax[b]-positive interneurons become pax[b] positive. This pattern of expression suggests that the pax[b] protein may be involved in determining the identity of specific commissural interneurons.

3,3'-Diaminobenzidine

Structure and early embryonic expression of the zebrafish engrailed-2 gene.

The Drosophila homeobox gene engrailed (en) is needed for correct embryonic development, and related sequences are active during vertebrate embryogenesis. Here we report the protein coding sequence and embryonic expression pattern of the zebrafish engrailed-2 gene (eng-2) which is directly homologous to En-2 in mice and Xenopus. The predicted zebrafish Eng-2 protein shares 65% overall identity to its Xenopus counterpart. In addition to the highly conserved homeodomain region, sequence conservation is present within three short stretches in the N-terminal region. The embryonic expression of the eng-2 gene was analysed by in situ hybridization to whole-mount embryos and tissue sections. Transcripts are first detected in two lateral bands at the 10-h stage, when epiboly is completed. Within the next 2 h of development, these two bands migrate and fuse at the midline. By the time the neural keel becomes visible (11-12 h), a transverse stripe of eng-2 expressing cells is seen at the presumptive midbrain-hindbrain boundary. Later this stripe becomes significantly compressed along the AP axis, and in 24-h embryos eng-2 transcripts are detected mainly in the posterior midbrain. In the hindbrain, eng-2 expression seems restricted to the primordium of the cerebellum. A second site of activity was observed in each somite where specific myotomal cells, the muscle pioneers, express eng-2. Our observations are discussed in relation to early regionalization of the central nervous system (CNS) and the generation of morphological borders.

Amino Acid Sequence

Expression of four zebrafish wnt-related genes during embryogenesis.

The wnt gene family codes for a group of cysteine-rich, secreted proteins, which are differentially expressed in the developing embryo and are possibly involved in cellular communication. Here, we describe the polymerase chain reaction based cloning and embryonic expression patterns of four zebrafish wnt-related sequences; wnt[a], wnt[b], wnt[c] and wnt[d]. One of these genes, wnt[a], is a potential homologue of murine Wnt-3, while the other three genes most likely represent new members of the vertebrate wnt gene family. In zebrafish embryos, transcripts of wnt[a] are confined to the dorsal diencephalon, the dorsal midbrain, the rhombic lips and the dorsal portions of the spinal cord. wnt[b] is expressed in the tail bud and at considerably lower levels in the mesoderm of the head. wnt[c] transcripts are present within the diencephalon and the posterior midbrain whereas wnt[d] shows a surprisingly similar expression pattern to zebrafish wnt-1. By analogy to wnt-1, it is likely that the members of the zebrafish wnt gene family play an important role in cell-to-cell signalling during pattern formation in the neural tube and the tail bud.

Amino Acid Sequence

Expression pattern of zebrafish pax genes suggests a role in early brain regionalization.

In vertebrates the developing hindbrain is organized in segmental units. These units provide the primary grid for differentiation and axonal outgrowth. In the more anterior regions of the brain, however, the subdivisions remain more controversial. Cellular and molecular studies of the embryonic brain in lower vertebrates such as the zebrafish, Brachydanio rerio, may reveal remnants of such subdivisions. We have isolated complementary DNA clones for two zebrafish pax genes related to Drosophila and mouse paired-box-containing segmentation genes. The expression of these two genes is confined to specific regions in the embryonic forebrain and midbrain. Strikingly, the borders of expression of the two pax genes coincide with morphological landmarks corresponding to the primary axon tracts that are generated in the embryonic brain a few hours after the initiation of expression of these genes.

Amino Acid Sequence