Prototheca filamenta: A fungus not an alga

The genusPrototheca is composed of colorless algae. Plastids, a characteristic feature of colorless algae, are absent fromP. filamenta cells. The growth pattern seen with light and scanning electron microscopes, and the ultrastructure observed with the transmission electron microscope suggest thatP. filamenta is a fungus. Consequently we recommend the removal ofP. filamenta from the genusPrototheca and subsequent investigation of this organism to determine its correct taxonomic placement among the fungi.     

Extracellular glycoprotein specific for Saccharomyces sensu stricto

Abstract Using affinity-purified rabbit polyclonal antibodies against an extracellular mannoprotein (gp400) of Saccharomyces cerevisiae , the presence of immunohomologic proteins with similar electrophoretic mobility was shown in the culture medium of S. bayanus, S. paradoxus and S. pastorianus . Cross-reactive bands with different electrophoretic behaviour were observed for S. dairensis, S. exiguus, S. kluyveri, S. unisporus and also for the species moved from Sacchromyces to Arxiozyma, Kluyveromyces, Pachytichospora, Torulaspora and Zygosaccharomyces , in contrast to ascosporous yeasts of other genera in which these proteins were not found.     

Morphology and molecular phylogeny of a new hypotrich ciliate,Anteholosticha songi nov. spec., and an American population of Holosticha pullaster (Müller, 1773) Foissner et al., 1991 (Ciliophora,Hypotrichia)

A new urostylid ciliate, Anteholosticha songi nov. spec., isolated from forest soil in Tibet, and an American population of Holosticha pullaster (Müller, 1773) Foissner et al., 1991, isolated from a freshwater pond in the USA, are investigated in terms of their morphology, ontogenesis, and molecular biology. Anteholosticha songi nov. spec. is characterized by a slender to ellipsoidal body measuring 160–205 × 40–55 μm in vivo; rod-shaped yellowish cortical granules arranged in irregular short rows; four dorsal kineties; adoral zone consisting of 35–40 membranelles; three frontal, one buccal, one parabuccal, two frontoterminal, two pretransverse, and four to six transverse cirri and 14–25 midventral pairs; 12–22 ellipsoidal macronuclear nodules longitudinally arranged in pairs left of cell mid-line. Supplemental information on morphogenesis in Holosticha pullaster is also presented. The phylogenetic relationship of Anteholosticha and Holosticha inferred from SSU rDNA sequence data are concordant with previous studies and showing that Holosticha is monophyletic whereas Anteholosticha is polyphyletic and should be split into two or more genera.     

Resumption of growth of heat inactivated embryonic epiphyses by grafting

Summary This communication describes a new experimental model for the study of the role of the extracellular matrix (ECM) in morphogenesis. In a preceding paper (Markson et al. 1991) we demonstrated that isolated epiphyses from femora of 6-day-old chick embryos grow during the first days in organ culture almost as well as their intact counterparts. Heating femora for 1 h at 45.2° C caused complete cessation of growth and proteoglycan biosynthesis. When the cut surface of a heat-inactivated (HI) epiphysis was brought into apposition with the cut surface of a live epiphysis and the attached pair placed in organ culture, the HI epiphysis began to grow and reached almost the same size as its live partner. The different possible interpretations of this finding are discussed. When a HI epiphysis of a certain shape (from humerus) is attached to a live epiphysis of a very different shape (from femur) and the attached pair is cultured for 6–7 days, the typical resumption of growth can be observed and the HI epiphysis that doubled or tripled its size retains its original characteristic form. The possibility that the existing infrastructure of the cartilaginous ECM directs the pattern of deposition of newly synthesized ECM by the chondroblasts is discussed, and it is suggested that stretch-activated channels participate in a process by which cells sense the topography of their ECM.Offprint requests to: F. Doljanski     

Siliceous structures and silicification in flagellated protists

Summary Flagellated protists produce a diverse range of siliceous structures, such as internal and external skeletons, scales, spines, bristles, cell walls, cyst walls, and loricae. The different groups of silica-depositing flagellates, i.e., chrysophytes/synurophytes, choanoflagellates, dinoflagellates, ebriids, silicoflagellates, thaumatomastigids, and the genusPetasaria are reviewed. Brief mention is also given to those algal groups in which silicification is uncommon and rare (i.e., chlorophytes, euglenophytes, haptophytes/prymnesiophytes, xanthophytes/tribophytes), but in which silicified structures nevertheless occur in few flagellate genera. Special attention is given to aspects of morphology and development of the different siliceous structures as well as on aspects of systematics and taxonomy.     

Developmental effects of monensin on Drosophila melanogaster

Extracellular matrix and membrane proteins and their correct secretion probably are key elements in morphogenesis and differentiation in Drosophila. In this study, we have analysed the effects of monensin, a Na⁺-H⁺-ionophore which blocks normal secretion, applied during cellular blastoderm formation on further development. Normal cell morphology and intercellular contacts are lost and the extracellular matrix becomes disorganized. Gastrulation is blocked and abnormal foldings can be observed. Cuticle phenotypes showed different degrees of ventral, dorsal, head and posterior defects. The results are discussed in the context of what is known about membrane and extracellular matrix proteins in Drosophila.     

Techniques for assessing 3-D cell–matrix mechanical interactions in vitro and in vivo

Cellular interactions with extracellular matrices (ECM) through the application of mechanical forces mediate numerous biological processes including developmental morphogenesis, wound healing and cancer metastasis. They also play a key role in the cellular repopulation and/or remodeling of engineered tissues and organs. While 2-D studies can provide important insights into many aspects of cellular mechanobiology, cells reside within 3-D ECMs in vivo, and matrix structure and dimensionality have been shown to impact cell morphology, protein organization and mechanical behavior. Global measurements of cell-induced compaction of 3-D collagen matrices can provide important insights into the regulation of overall cell contractility by various cytokines and signaling pathways. However, to understand how the mechanics of cell spreading, migration, contraction and matrix remodeling are regulated at the molecular level, these processes must also be studied in individual cells. Here we review the evolution and application of techniques for imaging and assessing local cell–matrix mechanical interactions in 3-D culture models, tissue explants and living animals.     

Growth and proteoglycan metabolism of chick embryonic cartilaginous long bone rudiments and of isolated epiphyses

Summary Turnover of extracellular matrix (ECM) proteoglycans was studied in chick cartilaginous femur rudiments grown in organ culture. Femora from six-day-old embryos showed nearly normal growth rates during the first few days in culture. By labeling the rudiment with ³⁵S-sulfate or ¹⁴C-glucosamine, it was demonstrated that the cartilaginous ECM undergoes rapid turnover. It was also found that the metabolic fate of the proteoglycans is to be released as macromolecules into the culture medium. When a rudiment was cut to obtain two epiphyses it was observed that each part grows and synthesizes proteoglycans at nearly normal rates, which indicates that the isolated epiphyses, like the whole rudiment, behave as autonomous systems. We suggest that the turnover of ECM components is part of the continuous remodelling process rudiments undergo during their growth and development. In order to study cell-ECM interaction in morphogenesis, we made an attempt to prepare an intact cell-free ECM. Epiphyses were heated at 45.2° C for 1 h. The treatment caused complete cessation of growth and biosynthesis. When the cut surface of a live epiphysis was brought into apposition to a heat-treated epiphysis and the attached pair placed in organ culture, it was found that the heat-treated epiphysis begins to grow and reaches almost the same size as its live counterpart. We discuss the possible advantage of this new experimental system for studies on the role of ECM in morphogenesis.     

Effect of gossypol on the ultrastructure of rat spermatozoa

Summary Gossypol was found to induce sterility in male rats when administered orally. A reduction in the number of spermatozoa in the epididymis from the gossypol-treated rats was observed when compared to the control animals. An examination of the spermatozoa from the treated rats showed the following ultrastructural modifications: disorganization of the mitochondrial sheath and missing cell membrane from the middle piece, broken cell membrane and missing members of both outer fibers and inner microtubules of the principal piece, and broken cell membrane of the sperm head. Serial mating experiments proved that gossypol-treated males were indeed sterile. The results suggest that gossypol at low concentrations is able to affect the motility of spermatozoa, thus contributing to its contraceptive action.     

The morphology of the juxtaglomerular apparatus in the toad,Bufo bufo

Summary In a light microscopic study the course of the tubule in the kidney of the toadBufo bufo was studied. The distal tubule returning to the glomerulus of its origin appears to enclose the afferent arteriole. In that area, from which a three dimensional graphic reconstruction is made, there is an intimate contact between tubular and vascular wall. The latter contains granulated media cells. In the part of the tubule adjacent to the afferent arteriole an accumulation of nuclei is present. It is suggested that this structure is similar to the macula densa of the mammalian juxtaglomerular apparatus. The functional significance of a stricture in the tubule distally from the macula densa-like structure is discussed.The authors wish to thank Mrs. Ineke van de Mee-Wienen and Miss Ans Rouwenhorst for their technical assistance and Mr. J. J. M. de Bekker for the realization of the graphic reconstruction.     

Extracellular matrix control of mammary gland morphogenesis and tumorigenesis: insights from imaging

The extracellular matrix (ECM), once thought to solely provide physical support to a tissue, is a key component of a cell’s microenvironment responsible for directing cell fate and maintaining tissue specificity. It stands to reason, then, that changes in the ECM itself or in how signals from the ECM are presented to or interpreted by cells can disrupt tissue organization; the latter is a necessary step for malignant progression. In this review, we elaborate on this concept using the mammary gland as an example. We describe how the ECM directs mammary gland formation and function, and discuss how a cell’s inability to interpret these signals—whether as a result of genetic insults or physicochemical alterations in the ECM—disorganizes the gland and promotes malignancy. By restoring context and forcing cells to properly interpret these native signals, aberrant behavior can be quelled and organization re-established. Traditional imaging approaches have been a key complement to the standard biochemical, molecular, and cell biology approaches used in these studies. Utilizing imaging modalities with enhanced spatial resolution in live tissues may uncover additional means by which the ECM regulates tissue structure, on different length scales, through its pericellular organization (short-scale) and by biasing morphogenic and morphostatic gradients (long-scale). Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Sid4: A secreted vertebrate immunoglobulin protein with roles in zebrafish embryogenesis

The small members of the immunoglobulin superfamily (IGSF) are a molecularly diverse group of proteins composed solely of immunoglobulin domains. They may be secreted or tethered to the cell mebrane via GPI linkages and are proposed to have important functions in vivo. However, very few small IGSFs have been functionally characterized. During an ongoing in situ hybridization analysis of expressed sequence tags in zebrafish we identified secreted immunoglobulin domain 4 (sid4), a gene encoding a soluble vertebrate protein composed solely of four immunoglobulin domains. Throughout development, sid4 is expressed in regions of the embryo undergoing active cell division and migration. Functional analysis using morpholino antisense oligonucleotides demonstrates that timing of gene expression is normal in morphants, but these embryos are smaller and exhibit defects in epiboly and patterning of axial and prechordal mesoderm. Analyses of chordin, pax2, krox20, and dlx2 expression in morphants demonstrate that early brain patterning is normal but later organization of hindbrain neurons and development of cranial neural crest are perturbed. Levels of apoptosis in morphants were normal prior to 90% epiboly, but were elevated after 10 h post-fertilization (hpf). Apoptosis does not account for early patterning defects of axial mesoderm, but likely contributes to overall reduction in embryo size. Phylogenetic analysis demonstrates that Sid4 is strikingly similar to the fibronectin binding Ig domains of Perlecan/HSPG2. Overall, our data demonstrate a fundamental role for sid4, possibly as a co-factor in extracellular matrix (ECM) interactions, in processes underlying tissue patterning and organogenesis in a vertebrate.     

Invertebrate Models of Kallmann Syndrome: Molecular Pathogenesis and New Disease Genes

Kallmann Syndrome is a heritable disorder characterized by congenital anosmia, hypogonadotropic hypogonadism and, less frequently, by other symptoms. The X-linked form of this syndrome is caused by mutations affecting the KAL1 gene that codes for the extracellular protein anosmin-1. Investigation of KAL1 function in mice has been hampered by the fact that the murine ortholog has not been identified. Thus studies performed in other animal models have contributed significantly to an understanding of the function of KAL1. In this review, the main results obtained using the two invertebrate models, the nematode worm Caenorhabditis elegans and the fruit fly Drosophila melanogaster, are illustrated and the contribution provided by them to the elucidation of the molecular pathogenesis of Kallmann Syndrome is discussed in detail. Structure-function dissection studies performed in these two animal models have shown how the different domains of anosmin-1 carry out specific functions, also suggesting a novel intramolecular regulation mechanism among the different domains of the protein. The model that emerges is one in which anosmin-1 plays different roles in different tissues, interacting with different components of the extracellular matrix. We also describe how the genetic approach in C. elegans has allowed the discovery of the genes involved in KAL1-heparan sulfate proteoglycans interactions and the identification of HS6ST1 as a new disease gene.