The occurrence of fimbriae on a N 2 2 -fixing cyanobacterium which occurs in lichen symbiosis

The Nostoc cyanobiont of the lichen Peltigera canina when grown on N₂ possesses, in the motile stage, discrete unbranched non-flagellar appendages (fimbriae or pili). These arise from the host cell surface in a peritrichous manner, have an axial hole, are 7.0 ±0.3 nm in diameter and are up to 3 m long. They do not haemagglutinate guinea pig red blood corpuscles and differ from the major fimbrial types reported for Gram-negative heterotrophic bacteria and from sex pili. They may be involved in motility and specificity in symbiotic cyanobacteria.     

Antigenic Properties of Bacteriophage φ29 Structural Proteins

Serological methods and electron microscopy were used to study the structural proteins of the small Bacillus subtilis bacteriophage phi29. This virus has a large number of fibers attached at both ends of its prolate head. A complex neck assembly is comprised of 12 symmetrically arranged appendages as the outer component. Head fibers, neck appendages, and the head surface bind anti-phi29 antibodies. Immune sera absorbed with defective lysates of suppressor-sensitive (sus) mutants have been used to determine the genetic control of neck appendages production. Studies on the serum-blocking power of lysates defective in different tail components showed that appendages contain the main serum-blocking protein. This finding suggests an essential role of the neck appendages in phage adsorption or DNA injection.     

Diversity of archaeal type IV pilin-like structures

Bacterial type IV pili perform important functions in such disparate biological processes as surface adhesion, cell–cell interactions, autoaggregation, conjugation, and twitching motility. Unlike bacteria, archaea use a type IV pilus related structure to drive swimming motility. While this unique flagellum is the best-studied example of an archaeal IV pilus-like structure, recent in silico, in vivo and structural analyses have revealed a highly diverse set of archaeal non-flagellar type IV pilus-like structures. Accumulating evidence suggests that these structures play important diverse roles in archaea.     

Characterization of a Novel <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> Phenotype Possessing Multiple Appendages Attached to a Parasporal Body

Bacillus thuringiensis is a bacterium best known for its production of crystal-like bodies comprised of one or more Cry-proteins, which can be toxic to insects, nematodes or cancer cells. Although strains of B. thuringiensis have occasionally been observed with filamentous appendages attached to their spores, appendages in association with their parasporal bodies are extremely rare. Herein we report the characterization of Bt1-88, a bacterial strain isolated from the Caribbean that produces a spore–crystal complex containing six long appendages, each comprised of numerous thinner filaments approximately 10 nm in diameter and 2.5 μm in length. Each of the multi-filament appendages was attached to a single, small parasporal body located at one end of the bacterial spore. Biochemical tests, 16S rDNA gene sequencing, and the identification of two Cry proteins by partial protein sequencing (putatively Cry1A and Cry2A), unambiguously identified Bt1-88 as a strain of B. thuringiensis. Bt1-88 represents the second reported strain of B. thuringiensis possessing a parasporal body/appendage phenotype characterized by one or more long appendages, comprised of numerous filaments in association with a parasporal body. This finding suggests that Bt1-88 is a member of a new phenotypic class of B. thuringiensis, in which the parasporal body may perform a novel structural role through its association with multi-filament appendages.     

Cytoskeleton Structure and Composition in Choanoflagellates

The structure and composition of the cytoskeleton has been studied in Monosiga ovata (Protozoa: Order Choanofiagellida Kent 1880) using a combination of methods in association with light and electron microscopy. Supplementary observations are included for Desmarella moniliformis. The basal body of the single anterior flagellum is subtended proximally and at right angles by a second, non-flagellar basal body. The edges of the two basal bodies are connected by a fibrillar bridge. A long, narrow, striated, fibrillar rootlet extends posteriorly from the lower edge of the non-flagellar basal body towards the Golgi apparatus. It is associated throughout most of its length with the surface of a flattened sac. Rootlet microtubules pass radially from a ring of electron dense material which encircles the distal end of the flagellar basal body. These microtubules extend outwards for about one-third of the length of the cell. Within each collar tentacle is a longitudinal bundle of microfilaments composed of actin as illustrated by rhodamine-phalloidin staining for fluorescence microscopy. The base of each microfilament bundle is associated with one or more rootlet microtubules by fine fibrillar bridges. The attachment between microtubules and tentacle microfilaments is further demonstrated by their coordinated displacement when the cytoskeleton becomes dislodged. The role of the cytoskeleton in maintaining the position of the collar tentacles during interphase and cell division is discussed.     

Larval leg integrity is maintained by Distal-less and is required for proper timing of metamorphosis in the flour beetle, Tribolium castaneum

The dramatic transformation from a larva to an adult must be accompanied by a coordinated activity of genes and hormones that enable an orchestrated transformation from larval to pupal/adult tissues. The maintenance of larval appendages and their subsequent transformation to appendages in holometabolous insects remains elusive at the developmental genetic level. Here the role of a key appendage patterning gene Distal-less (Dll) was examined in mid- to late-larval stages of the flour beetle, Tribolium castaneum. During late larval development, Dll was expressed in appendages in a similar manner as previously reported for the tobacco hornworm, Manduca sexta. Removal of this late Dll expression resulted in disruption of adult appendage patterning. Intriguingly, earlier removal resulted in dramatic loss of structural integrity and identity of larval appendages. A large amount of variability in appendage morphology was observed following Dll dsRNA injection, unlike larvae injected with dachshund dsRNA. These Dll dsRNA-injected larvae underwent numerous supernumerary molts, which could be terminated with injection of either JH methyltransferase or Methoprene-tolerant dsRNA. Apparently, the partial dedifferentiation of the appendages in these larvae acts to maintain high JH and, hence, prevents metamorphosis.     

The Expression of Beta ({beta}) Keratins in the Epidermal Appendages of Reptiles and Birds

The integuments of extant vertebrates display a variety of epidermalappendages whose patterns, morphology and terminal differentiation(epidermal keratins) depend upon interactions between ectodermal(epidermis) and mesodermal (dermis) tissues. In reptiles andbirds, appendage morphogenesis precedes terminal differentiation.Studies have demonstrated that appendage morphogenesis influencesthe expression of the appendage specific keratin genes. However,little is known about the nature of the structural genes expressedby the epidermal appendages of reptiles. How pattern formationand/or appendage morphogenesis influence terminal differentiationof reptilian appendages is not known. The epidermal appendages of reptiles and birds are characterizedby the presence of both alpha () and beta (ß) typekeratin proteins. Studies have focused on the genes of avianß keratins because they are the major structural proteinsof feathers. The occurrence of ß keratin proteinsin the scales and claws of both birds and reptiles and theirimmunological cross-reactivity suggest that the genes for reptilianß keratins may be homologous with those of birds.In bird appendages, the ß keratins are the productsof a large family of homologous genes. Specific members of thisgene family are expressed during the development of each appendage.Recent sequence analyses of feather ß keratins, fromdifferent orders of birds, demonstrate that there is more diversityat the DNA level than was implied by earlier protein sequencingstudies. Immunological techniques show that the same antibodies thatreact with the epidermal ß keratins of the chicken(Gallus domesticus) react with the epidermal ß keratinsof American alligators (Alligator mississippiensis). Furthermore,a peptide sequence (20 amino acids) from an alligator claw ßkeratin is similar to a highly conserved region of avian claw,scale, feather, and feather-like ß keratins. Theseobservations suggest that the ß keratin genes of avianepidermal appendages have homologues in the American alligator.Understanding the origin and evolution of the ß keratingene families in reptiles and birds will undoubtedly add toour understanding of the evolution of skin appendages such asscales and feathers.     

Fine structure of the elasmobranch renal tubule: neck and proximal segments of the little skate

This is the first in a series of studies that examines the renal tubular ultrastructure of elasmobranch fish. Each subdivision of the neck segment and proximal segment of the renal tubule of the little skate (Raja erinacea) has been investigated using electron microscopy of thin sections and freeze-fracture replicas. Flagellar cells, characterized by long, wavy, flagellar ribbons, were observed in both nephron segments. They were found predominantly in the first subdivision of the neck segment, which suggests that propulsion of the glomerular filtrate is a primary function of this part of the renal tubule. In the non-flagellar cells of the neck segment (subdivisions I and II), there were bundles of microfilaments, a few apical cell projections, and, in subdivision II, numerous autophagosomes. In the proximal segment, the non-flagellar cells varied in size, being low in subdivision I, cuboidal in II, tall columnar in III, and again low in IV. Apical cell projections were low and scattered in subdivisions I and IV and were highest in III where the basolateral plasma membrane was extremely amplified by cytoplasmic projections. Furthermore, in these cells the mitochondria were numerous with an extensive matrix and short cristae. A network of tubules of the endoplasmic reticulum characterized the apical region of the non-flagellar cells in subdivisions I, II, and IV. In the late part of subdivision II and the early part of III, the cells were characterized by numerous coated pits and vesicles, large subluminal vacuoles, and basally located dense bodies, all of which are structures involved in receptor-mediated endocytosis. Freeze-fracture replicas revealed gap junctions restricted to the cells of the first three subdivisions of the proximal segment. The zonulae occludentes were not different in the neck and proximal segments, being composed of several strands, suggesting a moderately leaky paracellular pathway.     

MODULARITY AND RATES OF EVOLUTIONARY CHANGE IN A POWER‐AMPLIFIED PREY CAPTURE SYSTEM

The dynamic interplay among structure, function, and phylogeny form a classic triad of influences on the patterns and processes of biological diversification. Although these dynamics are widely recognized as important, quantitative analyses of their interactions have infrequently been applied to biomechanical systems. Here we analyze these factors using a fundamental biomechanical mechanism: power amplification. Power‐amplified systems use springs and latches to generate extremely fast and powerful movements. This study focuses specifically on the power amplification mechanism in the fast raptorial appendages of mantis shrimp (Crustacea: Stomatopoda). Using geometric morphometric and phylogenetic comparative analyses, we measured evolutionary modularity and rates of morphological evolution of the raptorial appendage's biomechanical components. We found that “smashers” (hammer‐shaped raptorial appendages) exhibit lower modularity and 10‐fold slower rates of morphological change when compared to non‐smashers (spear‐shaped or undifferentiated appendages). The morphological and biomechanical integration of this system at a macroevolutionary scale and the presence of variable rates of evolution reveal a balance between structural constraints, functional variation, and the “roles of development and genetics” in evolutionary diversification.     

New TRAP1 and Hsp90 chaperone inhibitors with cationic components: Preliminary studies on mitochondrial targeting

TRAP1 (Hsp75) is the mitochondrial paralog of the Hsp90 molecular chaperone family. Due to structural similarity among Hsp90 chaperones, a potential strategy to induce apoptosis through mitochondrial TRAP1 ATPase inhibition has been envisaged and a series of compounds has been developed by binding the simple pharmacophoric core of known Hsp90 inhibitors with various appendages bearing a permanent cationic head, or a basic group highly ionizable at physiologic pH. Cationic appendages were selected as vehicles to deliver drugs to mitochondria. Indeed, masses of new derivatives were evidenced to accumulate in the mitochondrial fraction from colon carcinoma cells and a compound in the series, with a guanidine appendage, demonstrated good activity in inhibiting recombinant TRAP1 ATPase and cell growth and in inducing apoptotic cell death in colon carcinoma cells.     

Central core in the spinal grey matter.

Golgi--Kopsch impregnated spinal cord sections of adult cats revealed a regular orientation of the dendrites in the intermediate region forming a circular or elliptical grey matter. The orientation of the dendrites is strictly transverse in the central core. A new interpretation of the spinal grey matter is proposed by viewing the dorsal, lateral and ventral horns (or columns) as crest-like appendages attached to the rod-like central core. A structural and possibly functional similarity between the central core and the brain stem reticular formation is emphasized.     

Type III secretion: a secretory pathway serving both motility and virulence (review)

'Type III secretion' (T3S) refers to a secretion pathway that is common to the flagellae of eubacteria and the injectisomes of some gram-negative bacteria. Flagellae are rotary nanomachines allowing motility but they contain a built-in secretion apparatus that exports their own distal components to the distal end of the growing structure where they polymerize. In some cases they have been shown to export non-flagellar proteins. Injectisomes are transkingdom communication apparatuses allowing bacteria docked at the surface of a eukaryotic cell membrane to inject effector proteins across the two bacterial membranes and the eukaryotic cell membrane. Both nanomachines share a similar basal body embedded in the two bacterial membranes, topped either by a hook and a filament or by a stiff short needle. Both appear to be assembled in the same fashion. They recognize their substrate by a loose N-terminal peptide signal and the help of individual chaperones of a new type.     

Membrane tubules attach Salmonella Typhimurium to eukaryotic cells and bacteria

Using scanning electron microscopy techniques we measured the diameter of adhesive tubular appendages of Salmonella enterica serovar S. Typhimurium. The appendages interconnected bacteria in biofilms grown on gallstones or coverslips, or attached bacteria to host cells (human neutrophils). The tubular appendage diameter of bacteria of virulent flagellated C53 strain varied between 60 and 70 nm, thus considerably exceeding in size of flagella or pili. Nonflagellated bacteria of mutant SJW 880 strain in biofilms grown on gallstones or coverslips were also interconnected by 60-90-nm tubular appendages. Transmission electron microscopy studies of thin sections of S. Typhimurium biofilms grown on agar or coverslips revealed numerous fragments of membrane tubular and vesicular structures between bacteria of both flagellated and nonflagellated strains. The membrane structures had the same diameter as tubular appendages observed by scanning electron microscopy, indicating that tubular appendages might represent membrane tubules (tethers). Previously, we have shown that neutrophils can contact cells and bacteria over distance via membrane tubulovesicular extensions (TVE) (cytonemes). The present electron microscopy study revealed the similarities in size and behavior of bacterial tubular appendages and neutrophil TVE. Our data support the hypothesis that bacteria establish long-range adhesive interactions via membrane tubules.     

Coiled-coils in type III secretion systems: structural flexibility, disorder and biological implications

Recent structural studies and analyses of microbial genomes have consolidated the understanding of the structural and functional versatility of coiled-coil domains in proteins from bacterial type III secretion systems (T3SS). Such domains consist of two or more α-helices forming a bundle structure. The occurrence of coiled-coils in T3SS is considerably higher than the average predicted occurrence in prokaryotic proteomes. T3SS proteins comprising coiled-coil domains are frequently characterized by an increased structural flexibility, which may vary from localized structural disorder to the establishment of molten globule-like state. The propensity for coiled-coil formation and structural disorder are frequently essential requirements for various T3SS functions, including the establishment of protein–protein interaction networks and the polymerization of extracellular components of T3SS appendages. Possible correlations between the frequently observed N-terminal structural disorder of effectors and the T3SS secretion signal are discussed. The results for T3SS are also compared with other Gram-negative secretory systems.     

Segmentation and tagmosis in Chelicerata

Patterns of segmentation and tagmosis are reviewed for Chelicerata. Depending on the outgroup, chelicerate origins are either among taxa with an anterior tagma of six somites, or taxa in which the appendages of somite I became increasingly raptorial. All Chelicerata have appendage I as a chelate or clasp-knife chelicera. The basic trend has obviously been to consolidate food-gathering and walking limbs as a prosoma and respiratory appendages on the opisthosoma. However, the boundary of the prosoma is debatable in that some taxa have functionally incorporated somite VII and/or its appendages into the prosoma. Euchelicerata can be defined on having plate-like opisthosomal appendages, further modified within Arachnida. Total somite counts for Chelicerata range from a maximum of nineteen in groups like Scorpiones and the extinct Eurypterida down to seven in modern Pycnogonida. Mites may also show reduced somite counts, but reconstructing segmentation in these animals remains challenging. Several innovations relating to tagmosis or the appendages borne on particular somites are summarised here as putative apomorphies of individual higher taxa. We also present our observations within the concept of pseudotagma, whereby the true tagmata – the prosoma and opisthosoma – can be defined on a fundamental change in the limb series while pseudotagmata, such as the cephalosoma/proterosoma, are expressed as divisions in sclerites covering the body without an accompanying change in the appendages.     

Bioinformatics, genomics and evolution of non-flagellar type-III secretion systems: a Darwinian perspective

We review the biology of non-flagellar type-III secretion systems from a Darwinian perspective, highlighting the themes of evolution, conservation, variation and decay. The presence of these systems in environmental organisms such as Myxococcus, Desulfovibrio and Verrucomicrobium hints at roles beyond virulence. We review newly discovered sequence homologies (e.g., YopN/TyeA and SepL). We discuss synapomorphies that might be useful in formulating a taxonomy of type-III secretion. The problem of information overload is likely to be ameliorated by launch of a web site devoted to the comparative biology of type-III secretion ().     

The insect upper lip (labrum) is a nonsegmental appendage-like structure

SUMMARY The insect upper lip—the labrum—is a lobe-like structure anterior to the mouth opening. Whether the labrum represents a fused pair of segmental appendages or evolved independently is heavily debated. Here, we identify additional similarities of the regulatory gene network active in labrum and trunk appendages. However, we do not find a labral parasegment boundary and we show that labral Tc-Dll expression is independent of Tc-wg and Tc-hh signals. In contrast, Tc-Dll expression in all trunk appendages does require these signals. Finally, we identify crucial differences between the location of the labrum and trunk appendages: the labrum develops in median rather than lateral tissues and is part of an anterior nonsegmental tissue marked by and dependent on Tc-six3 activity. To reconcile these seeming contradictory results, we propose that the genetic network evolved in either labrum or trunk appendages and became redeployed at a novel location to form the other structure.     

Oxylipin covered ascospores of Eremothecium coryli

Eremothecium coryli is known to produce intriguing spindle-shaped ascospores with long and thin whip-like appendages. Here, ultra structural studies using scanning electron microscopy, indicate that these appendages serve to coil around themselves and around ascospores causing spore aggregation. Furthermore, using immunofluorescence confocal laser scanning microscopy it was found that hydrophobic 3-hydroxy oxylipins cover the surfaces of these ascospores. Using gas chromatography–mass spectrometry, only the oxylipin 3-hydroxy 9:1 (a monounsaturated fatty acid containing a hydroxyl group on carbon 3) could be identified. Sequential digital imaging suggests that oxylipin-coated spindle-shaped ascospores are released from enclosed asci probably by protruding through an already disintegrating ascus wall.     

Characterization of spore appendages from Bacillus cereus strains

AIMS: Further characterization and comparison of spore appendages from Bacillus cereus strains. METHODS AND RESULTS: Appendages were isolated from 10 B. cereus strains from the food industry and food-borne outbreaks. The appendage proteins were dissolved in sample buffer containing 2% SDS and 5% mercaptoethanol at 100 degrees C, and subjected to SDS-PAGE. None of the appendages showed identical protein patterns. Western blots, using antibodies raised against a 3.5 kDa appendage protein, showed that the majority of the appendage proteins reacted with the antibody. Removal of the appendages by sonic treatment of the spores did not alter their heat resistance. The appendages were digested by proteinase K, pepsin, and the enzymes in the detergent Paradigm 10, but not by trypsin or chymotrypsin. Spore adhesion to stainless steel was scarcely affected by removal of the appendages. Digestion of adhered intact spores (with appendages) with Paradigm 10 showed a high degree of variation. CONCLUSIONS: Spore appendages from B. cereus are complex proteinaceous structures that differ among strains. SIGNIFICANCE AND IMPACT OF THE STUDY: Information about spore appendages and their involvement in spore adhesion is crucial for improving cleaning methods used for control of bacterial spores in the food industry.