The role actin filaments play in providing the characteristic curved form of Drosophila bristles

Drosophila bristles display a precise orientation and curvature. An asymmetric extension of the socket cell overlies the newly emerging bristle rudiment to provide direction for bristle elongation, a process thought to be orchestrated by the nerve dendrite lying between these cells. Scanning electron microscopic analysis of individual bristles showed that curvature is planar and far greater near the bristle base. Correlated with this, as development proceeds the pupa gradually recedes from the inner pupal case (an extracellular layer that encloses the pupa) leading to less bristle curvature along the shaft. We propose that the inner pupal case induces elongating bristles to bend when they contact this barrier. During elongation the actin cytoskeleton locks in this curvature by grafting together the overlapping modules that comprise the long filament bundles. Because the bristle is curved, the actin bundles on the superior side must be longer than those on the inferior side. This is accomplished during grafting by greater elongation of superior side modules. Poor actin cross-bridging in mutant bristles results in altered curvature. Thus, the pattern of bristle curvature is a product of both extrinsic factors-the socket cell and the inner pupal case--and intrinsic factors--actin cytoskeleton assembly.     

Crystal types and their distribution in the bark of African genistoid legumes (Fabaceae tribes Sophoreae,Podalyrieae, Crotalarieae and Genisteae)

The occurrence and distribution of seven crystal types in 114 bark samples from 25 genera and 91 species, representing all four tribes of African genistoid legumes, are reported. The seven types are prismatic, druse (including irregular crystal clusters), navicular (including truncated navicular, here reported for the first time), spherical crystal cluster, styloid, crystal sand and acicular crystals in sheaf‐like aggregates. Unlike most studies, the elemental composition of the crystals was examined using X‐ray microanalyses. With the exception of acicular crystals, all crystals showed the typical peaks of calcium (sometimes accompanied by small amounts of magnesium). Acicular crystals in sheaf‐like aggregates were composed only of carbon and oxygen, indicating that they are organic and precipitate during drying or after fixation in alcohol. These crystals are found only in the two early‐diverging lineages of Podalyrieae (Cyclopia and Virgilia+Calpurnia clades), consistent with the phylogenetic pattern in the tribe (indicating a secondary loss). Navicular crystals are restricted to Podalyrieae and Crotalarieae. Prismatic crystals in bark are proposed to be the ancestral condition, with multiple losses (or reversals) in Podalyrieae, Crotalarieae and Genisteae. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 178 , 620–632.     

Actin filaments, stereocilia, and hair cells of the bird cochlea. IV. How the actin filaments become organized in developing stereocilia and in the cuticular plate

In 8-day-old embryos stereocilia can be identified on the hair cells of the chick cochlea; within each is a small population of actin filaments which extend from the tip of the stereocilium to the apical cytoplasm of the cell. These filaments are not ordered in a regular way, however, and tend to be found near the lateral margins of the stereocilia with large spaces between adjacent filaments. By 9 days the spaces between adjacent filaments are reduced and there are regions where the crossover points of adjacent actin helices are in register even though in cross section the actin filaments do not lie on a regular lattice. By 10-11 days the actin filaments become progressively more crossbridged together and we can recognize in longitudinal section horizontal stripes caused by the periodicity of the crossbridges. In transverse section the filaments begin to lie on a hexagonal lattice. Each stereocilium, however, contains less than 100 actin filaments. Evidence is presented that once crossbridging is maximal and the filaments hexagonally packed (Days 11-12), the stereocilia increase in width by the orderly addition of actin filaments to the lateral margins of the existing filament bundle so that by Day 16 we find up to 400 filaments all packed on a hexagonal lattice. Thus there are two stages in bundle formation. In the first a small number of filaments condense into a hexagonally packed, crosslinked bundle. In the second, the bundle increases in diameter by addition of filaments to the periphery of the bundle in a process akin to crystal growth. From observations on the elongation of filaments in the rootlets and stereocilia, we conclude that rootlets grow by addition of subunits at the nonpreferred end while stereocilia elongate by addition to the preferred end. What makes this interesting is that these two modes of addition occur at different developmental times.     

The Epidermal Cell Structure of the Secondary Pollen Presenter in Vangueria infausta (Rubiaceae: Vanguerieae) Suggests a Functional Association with Protruding Onci in Pollen Grains

Secondary pollen presentation is a well-known phenomenon in the Rubiaceae with particularly conspicuous pollen presenters occurring in the tribe Vanguerieae. These knob-like structures are formed by a modification of the upper portion of the style and stigma, together known as the stylar head complex. In the flower bud and shortly before anthesis, the anthers surrounding the stylar head complex dehisce and release pollen grains which adhere to the pollen presenter. The epidermal cells of the pollen presenter facing the anthers are radially elongated with a characteristic wall thickening encircling the anticlinal walls of each cell towards the distal end. These cells were studied in the pollen presenter of Vangueria infausta using electron and light microscopy in conjunction with histochemical tests and immunohistochemical methods. Other prominent thickenings of the cell wall were also observed on the distal and proximal walls. All these thickenings were found to be rich in pectin and possibly xyloglucan. The terms “thickenings of Igersheim” and “bands of Igersheim” are proposed to refer, respectively, to these wall structures in general and those encircling the anticlinal walls of each cell near the distal end. The epidermal cells have an intricate ultrastructure with an abundance of organelles, including smooth and rough endoplasmic reticulum, Golgi apparatus, mitochondria and secretory vesicles. This indicates that these cells are likely to have an active physiological role. The pollen grains possess prominent protruding onci and observations were made on their structure and development. Walls of the protruding onci are also rich in pectin. Pectins are hydrophilic and known to be involved in the dehydration and rehydration of pollen grains. We hypothesise that the thickenings of Igersheim, as well as the protruding onci of the pollen grains, are functionally associated and part of the adaptive syndrome of secondary pollen presentation, at least in the Vanguerieae.     

Two phenotypically distinct populations of T cells have suppressor capabilities simultaneously in the maintenance phase of immunologic enhancement

The events leading to immunologic enhancement in LEW rats immunized actively with Brown Norway (BN) rat spleen cells and passively with LEW anti-BN hyperimmune serum 11 and 10 days before receiving (LEW X BN)F1 cardiac allografts, respectively, have been studied. Cellular suppressor mechanisms developing during the induction phase of this phenomenon have recently been shown to be mediated by W3/25+ T cells in an antigen-specific manner. The present study suggests that the late maintenance phase of immunologic enhancement is mediated in vivo by simultaneously present separate donor-specific T cell subpopulations of W3/25+ and OX8+ phenotypes. Splenocyte subsets from grafted recipients greater than 100 days after transplantation were adoptively transferred into unmodified syngeneic LEW rats that received a specific test allograft 24 hr later, or into B recipients bearing indefinitely surviving heart grafts. Test graft survival was prolonged significantly in the first group and not altered in the second. Indeed, nonoverlapping W3/25+ and OX8+ cell fractions were separately responsible for suppression. However, when suppressor activity was tested in vitro in a three-component coculture mixed lymphocyte reaction, no suppression by T cells was obtained; this lack of correlation between in vivo and in vitro results has also been noted by other investigators in different systems. Thus, in the maintenance phase of actively and passively induced immunologic enhancement, interplay between two phenotypically distinct T cells with suppressor characteristics, but not putative cell-surface blocking factors, seems to prevent development of an alloreactive response and mediate host unresponsiveness.     

A new species of Polemanniopsis (Apiaceae) from Namibia

A new species of woody Apiaceae, Polemanniopsis namibensis B-E. Van Wyk, A. Burke & C. Mannheimer, is described. The species is allied to Polemanniopsis marlothii but differs in the much smaller habit of (up to 0.6 m vs 1.5–4 m in Polemanniopsis marlothii), the white or greyish colour of the twigs (brown in Polemanniopsis marlothii), the 3-foliolate leaves (predominantly 5-foliate in Polemanniopsis marlothii), the smaller inflorescences with persistent involucral bracts (caducous in Polemanniopsis marlothii) and anatomical details of the leaves. The species is endemic to Namibia and is known only from a localised area in the southern Namib desert near Lüderitz, where it is relatively common.     

Evidence for the presence of suppressor T lymphocytes in animals treated with cyclosporin A

Mice sensitized with alloantigens and treated with cyclosporin A (CsA) were incapable of generating antigen-specific cytolytic lymphocytes (CL). Lymphocytes from these CsA-treated animals could not be reactivated upon exposure to the same alloantigens in mixed lymphocyte culture (MLC), whereas their response to a third-party antigen remained intact, suggesting a long-lasting and specific effect of CsA. After being irradiated, these lymphocytes from CsA-treated animals were added to normal MLC and were shown to prevent normal lymphocytes from becoming cytolytic in a dose-dependent and antigen-nonspecific fashion. These suppressor cells were not detected in mice receiving CsA only, indicating that CsA did not induce but rather permitted the expression of suppressor cells possibly generated by allosensitization. The suppressor cells appeared to be T lymphocytes, because treatment with anti-Thy-1.2 antibody and C abrogated their suppressive activity. The present results suggest that activation and/or sparing of suppressor cells by CsA may account for the long-lasting unresponsiveness seen in CsA-treated animals.     

Antibodies designed as effective cancer vaccines

Antigen/antibody complexes can efficiently target antigen presenting cells to allow stimulation of the cellular immune response. Due to the difficulty of manufacture and their inherent instability complexes have proved inefficient cancer vaccines. However, anti-idiotypic antibodies mimicking antigens have been shown to stimulate both antibody and T cell responses. The latter are due to T cell mimotopes expressed within the complementarity-determining regions (CDRs) of antibodies that are efficiently presented to dendritic cells in vivo. Based on this observation we have designed a DNA vaccine platform called ImmunoBody™, where cytotoxic T lymphocyte (CTL) and helper T cell epitopes replace CDR regions within the framework of a human IgG1 antibody. The ImmunoBody™ expression system has a number of design features which allow for rapid production of a wide range of vaccines. The CDR regions of the heavy and light chain have been engineered to contain unique restriction endonuclease sites, which can be easily opened, and oligonucleotides encoding the T cell epitopes inserted. The variable and constant regions of the ImmunoBody™ are also flanked by restriction sites, which permit easy exchange of other IgG subtypes. Here we show a range of T cell epitopes can be inserted into the ImmunoBody™ vector and upon immunization these T cell epitopes are efficiently processed and presented to stimulate high frequency helper and CTL responses capable of anti-tumor activity.Key words: DNA vaccines, cancer vaccines, melanoma, CTL, helper T cells     

Actin filament turnover regulated by cross-linking accounts for the size, shape, location, and number of actin bundles in Drosophila bristles

Drosophila bristle cells are shaped during growth by longitudinal bundles of cross-linked actin filaments attached to the plasma membrane. We used confocal and electron microscopy to examine actin bundle structure and found that during bristle elongation, snarls of uncross-linked actin filaments and small internal bundles also form in the shaft cytoplasm only to disappear within 4 min. Thus, formation and later removal of actin filaments are prominent features of growing bristles. These transient snarls and internal bundles can be stabilized by culturing elongating bristles with jasplakinolide, a membrane-permeant inhibitor of actin filament depolymerization, resulting in enormous numbers of internal bundles and uncross-linked filaments. Examination of bundle disassembly in mutant bristles shows that plasma membrane association and cross-bridging adjacent actin filaments together inhibits depolymerization. Thus, highly cross-bridged and membrane-bound actin filaments turn over slowly and persist, whereas poorly cross-linked filaments turnover more rapidly. We argue that the selection of stable bundles relative to poorly cross-bridged filaments can account for the size, shape, number, and location of the longitudinal actin bundles in bristles. As a result, filament turnover plays an important role in regulating cytoskeleton assembly and consequently cell shape.