Antimicrobial activity of synthetic bactenecin

Bactenecin is an antimicrobial peptide isolated from bovine neutrophils. Bactenecin was synthesized by solid-phase peptide synthesis and renatured to a fully disulfide bonded form. The peptide inhibits the growth of Escherichia coli and Staphylococcus aureus at the same concentration reported for the peptide purified from bovine neutrophils. Bactenecin inhibits the growth of other medically important bacteria and yeast, and it kills the fungus Trichophyton rubrum. Acetylation and amidation of the amino- and carboxy-termini of bactenecin do not change its potency, while replacement of its two cysteine residues with serine decreases the potency.     

Reclustering the cluster flies (Diptera: Oestroidea,Polleniidae)

A phylogenetic analysis of selected oestroid taxa based on 66 morphological traits and sequences from three nuclear protein‐coding genes (CAD, MAC, MCS) resolved the composition and phylogenetic position of the former subfamily Polleniinae of the Calliphoridae – here resurrected at family rank as Polleniidae Brauer & Bergenstamm, 1889 stat. rev. Six species are transferred from the family Rhinophoridae to the Polleniidae: the Palaearctic genus Alvamaja Rognes, along with its single species Alvamaja chlorometallica Rognes, and five Afrotropical species comprising the carinata‐group formerly in the genus Phyto Robineau‐Desvoidy but here assigned to genus Morinia Robineau‐Desvoidy, i.e. M. carinata (Pape, 1987) comb.n. , M. lactineala (Pape, 1997) comb.n. , M. longirostris (Crosskey, 1977) comb.n. , M. royi (Pape, 1997) comb.n. and M. stuckenbergi (Crosskey, 1977) comb.n. The Polleniidae are monophyletic and, in agreement with most recent phylogenetic reconstructions, sister to the Tachinidae. The female of A. chlorometallica and a new species of Morinia of the carinata‐group (Morinia tsitsikamma sp.n. from South Africa) are described. This published work has been registered in ZooBank, http://zoobank.org/urn:lsid:zoobank.org:pub:25B0C220‐DEE4‐4B0C‐88EA‐35FDE298EBC5 .     

Characterization of the cDNA and gene for mouse tumour necrosis factor alpha converting enzyme (TACE/ADAM17) and its location to mouse chromosome 12 and human chromosome 2p25.

Numerous proteins are cleaved or "shed" from their membrane-bound form. One such protein, tumour necrosis factor alpha (TNF-alpha), is synthesized as a type 2 transmembrane protein. Recently, a human protease responsible for this shedding, the TNF-alpha converting enzyme (TACE/ADAM17), was isolated. TACE/ADAM17 is a member of the adamalysin class of zinc-binding metalloproteases or ADAM (a disintegrin and metalloprotease). We report the isolation and characterization of the mouse TACE/ADAM17 cDNA and gene. Mouse TACE/ADAM17 has a 92% amino-acid identity with the human protein and was ubiquitously expressed. A recombinant form of the protease is found to cleave a peptide representing the cleavage site of precursor mouse TNF-alpha. An alternatively spliced form of mouse TACE/ADAM17 was found that would produce a soluble protein. The gene for TACE/ADAM17 is approximately 50 kb and contains 19 exons. Chromosomal mapping places TACE/ADAM17 on mouse chromosome 12 and human chromosome 2p25.     

Blockade of EphA receptor tyrosine kinase activation inhibits vascular endothelial cell growth factor-induced angiogenesis

Angiogenesis is a multistep process involving a diverse array of molecular signals. Ligands for receptor tyrosine kinases (RTKs) have emerged as critical mediators of angiogenesis. Three families of ligands, vascular endothelial cell growth factors (VEGFs), angiopoietins, and ephrins, act via RTKs expressed in endothelial cells. Recent evidence indicates that VEGF cooperates with angiopoietins to regulate vascular remodeling and angiogenesis in both embryogenesis and tumor neovascularization. However, the relationship between VEGF and ephrins remains unclear. Here we show that interaction between EphA RTKs and ephrinA ligands is necessary for induction of maximal neovascularization by VEGF. EphA2 RTK is activated by VEGF through induction of ephrinA1 ligand. A soluble EphA2-Fc receptor inhibits VEGF-, but not basic fibroblast growth factor-induced endothelial cell survival, migration, sprouting, and corneal angiogenesis. As an independent, but complementary approach, EphA2 antisense oligonucleotides inhibited endothelial expression of EphA2 receptor and suppressed ephrinA1- and VEGF-induced cell migration. Taken together, these data indicate an essential role for EphA receptor activation in VEGF-dependent angiogenesis and suggest a potential new target for therapeutic intervention in pathogenic angiogenesis.     

Tonotopic gradients of Eph family proteins in the chick nucleus laminaris during synaptogenesis

Topographically precise projections are established early in neural development. One such topographically organized network is the auditory brainstem. In the chick, the auditory nerve transmits auditory information from the cochlea to nucleus magnocellularis (NM). NM in turn innervates nucleus laminaris (NL) bilaterally. These projections preserve the tonotopy established at the level of the cochlea. We have begun to examine the expression of Eph family proteins during the formation of these connections. Optical density measurements were used to describe gradients of Eph proteins along the tonotopic axis of NL in the neuropil, the somata, and the NM axons innervating NL at embryonic day 10, when synaptic connections from NM to NL are established. At E10-11, NL dorsal neuropil expresses EphA4 at a higher concentration in regions encoding high frequency sounds, decreasing in concentration monotonically toward the low frequency (caudolateral) end. In the somata, both EphA4 and ephrin-B2 are concentrated at the high frequency end of the nucleus. These tonotopic gradients disappear between E13 and E15, and expression of these molecules is completely downregulated by hatching. The E10-11 patterns run counter to an apparent gradient in dendrite density, as indicated by microtubule associated protein 2 (MAP2) immunolabeling. Finally, ephrin-B2 is also expressed in a gradient in tissue ventral to the NL neuropil. Our findings thus suggest a possible conserved mechanism for establishing topographic projections in diverse sensory systems. These results of this study provide a basis for the functional examination of the role of Eph proteins in the formation of tonotopic maps in the brainstem.     

Functional analysis of the domain structure of tumor necrosis factor-alpha converting enzyme

Many membrane-bound proteins, including cytokines, receptors, and growth factors, are proteolytically cleaved to release a soluble form of their extracellular domain. The tumor necrosis factor (TNF)-alpha converting enzyme (TACE/ADAM-17) is a transmembrane metalloproteinase responsible for the proteolytic release or "shedding" of several cell-surface proteins, including TNF and p75 TNFR. We established a TACE-reconstitution system using TACE-deficient cells co-transfected with TACE and substrate cDNAs to study TACE function and regulation. Using the TACE-reconstitution system, we identified two additional substrates of TACE, interleukin (IL)-1R-II and p55 TNFR. Using truncations and chimeric constructs of TACE and another ADAM family member, ADAM-10, we studied the function of the different domains of TACE in three shedding activities. We found that TACE must be expressed with its membrane-anchoring domain for phorbol ester-stimulated shedding of TNF, p75 TNFR, and IL-1R-II, but that the cytoplasmic domain is not required for the shedding of these substrates. The catalytic domain of ADAM-10 could not be functionally substituted for that of TACE. IL-1R-II shedding required the cysteine-rich domain of TACE as well as the catalytic domain, whereas TNF and p75 TNFR shedding required only the tethered TACE catalytic domain.