Maintenance Of Cell Shape Research Articles

Possible role of a choline-containing teichoic acid in the maintenance of normal cell shape and physiology in Streptococcus oralis.

Streptococcus oralis ATCC 35037 took up radioactively labeled choline from growth medium. Most of the choline (80 to 90%) was incorporated into the cell wall teichoic acid, and about 10% was localized in the plasma membrane. While cells grew in choline-free medium, they did so at slow rates and produced cell walls with greatly reduced amounts of phosphate and no detectable choline. Cells grown in choline-free medium had grossly abnormal shape and size. Both biochemical and morphological abnormalities were reversible by addition of choline to the medium.

Morphogenesis in Drosophila requires nonmuscle myosin heavy chain function.

We provide the first link between a known molecular motor and morphogenesis, the fundamental process of cell shape changes and movements that characterizes development throughout phylogeny. By reverse genetics, we generate mutations in the Drosophila conventional nonmuscle myosin (myosin II) heavy chain gene and show that this gene is essential. We demonstrate that these mutations are allelic to previously identified, recessive, embryonic-lethal zipper mutations and thereby identify nonmuscle myosin heavy chain as the zipper gene product. Embryos that lack functional myosin display defects in dorsal closure, head involution, and axon patterning. Analysis of cell morphology and myosin localization during dorsal closure in wild-type and homozygous mutant embryos demonstrates a key role for myosin in the maintenance of cell shape and suggests a model for the involvement of myosin in cell sheet movement during development. Our experiments, in conjunction with the observation that cytokinesis also requires myosin, suggest that the processes of cell shape change in morphogenesis and cell division are intimately and mechanistically related.

Sequence encoding ribosomal protein L33 of Lactococcus lactis

A cloned fragment from Lactococcus lactis chromosome encoding the L33 ribosomal protein was sequenced. Two incomplete open reading frames (ORFs) were also found: the upstream ORF shows similarity to the tetracycline-resistance protein (Tet) of Bacillus stearothermophilus, and the downstream ORF shows homology to a protein of Bacillus subtilis participating in sporulation (SpoVE), and to proteins of Escherichia coli involved in cell division (FtsW) and the maintenance of cell shape (RodA).

Centractin is an actin homologue associated with the centrosome

Actin is one of the most ubiquitous, abundant and well-conserved proteins of eukaryotes, participating in many crucial cellular processes including the maintenance of cell shape, motility and cell division. Actins from the most divergent sources still share amino-acid identities in excess of 70% (ref. 3). This may well explain why low-abundance homologues of actin have been difficult to isolate. Genes encoding distant relatives of actin in budding and fisson yeast have now been cloned. We report here the discovery of a vertebrate actin-like protein, which we name centractin. A full-length complementary DNA clone was isolated whose sequence reveals amino-acid identities with actin of over 50%, increasing to more than 70% when conservative amino-acid changes are considered. Northern analysis and western blotting indicate a ubiquitous tissue and species distribution. Morphological and biochemical criteria show that centractin is associated with centrosomes.

Contractile proteins in podocytes: immunocytochemical localization of actin and alpha-actinin in normal and nephrotic rat kidneys

Actin and alpha-actinin immunoreactive sites have been localized at the electron microscope level by the protein A-gold immunocytochemical technique in podocytes of normal and nephrotic rat renal tissues. In normal renal glomeruli, fibrillar networks located in the core of foot processes or bundles of microfilaments interconnecting them were found to be labelled for these two cytoskeletal proteins. On the other hand, in nephrotic renal glomeruli, concomitant with the loss of podocytic foot processes a reorganization of the podocytic cytoskeleton and a concentration of some of its elements into thick uniform bands was observed. Actin and alpha-actinin were revealed in these bands. Control experiments confirmed the specificity of the labelling obtained. Our results suggest that normal podocytes contain an actin-based contractile system that might contribute to the maintenance of the particular cell shape of these cells and that the rearrangement of the podocytic cytoskeleton occurring in the nephrotic syndrome might account for the changes in the foot processes and contribute to the alteration in glomerular function.

Cytochemical evidence of an organized microtubular cytoskeleton in Xenopus laevis oocytes: Involvement in the segregation of mitochondrial populations

An organized microtubular cytoskeleton was discovered in the cytoplasm of Xenopus laevis oocytes. The microtubules were observed in 10- to 30-micron cryostat sections by indirect immunoperoxidase labeling using an antibody to tubulin. A gradual extraction of cells with a nonionic detergent was essential for good penetration of the antibody into the cells. In the cytoplasm of all previtellogenic oocytes, a dense network of criss-crossed long microtubules was associated in a basket-like structure surrounding the mitochondrial mass. At the beginning of vitellogenesis, the network meshes enlarged, while clusters of mitochondria migrated, in close association with microtubule bundles. At the beginning of vitellogenesis, the reorganization of the microtubular network, mostly in the vegetal hemisphere, occurred during the segregation of the mitochondrial populations. Reorganization is characterized by (1) a temporary enlargement of the network and close association of mitochondrial clusters with microtubular bundles, and (2) a progressive organization of a ring-shaped microtubular structure in the crown elaboration area. It is hypothesized that these modifications of the microtubular cytoskeleton contribute to the maintenance of cell shape and the polarized organization of the cell.

Effect of the anti-microtubule drug oryzalin on growth and differentiation of the parasitic protozoan Leishmania mexicana

Abstract The parasitic protozoan Leishmania mexicana differentiates from a non-motile intracellular amastigote in the mammalian macrophage phagolysosome into a motile, extracellular promastigote in the insect vector gut. This development program has been accomplished in vitro, thus providing a useful model for studying changes in the cytoskeleton during cell differentiation. The role of microtubules in leishmania differentiation was demonstrated by using the dinitroaniline herbicide oryzalin, which inhibited both leishmania proliferation and differentiation; 25 μ M oryzalin reduced promastigote division by over 95%. Interestingly, at a sublethal dose (5 μ M), promastigotes became round and multifla-gellated but remained motile. At 50 μ M oryzalin, the number of intracellular amastigotes decreased by 50%. However, leishmania differentiation seemed to be the most drug-sensitive stage: there was a 60% reduction in amastigote-to-promastigote differentiation at 0.5 μ M oryzalin. The specific action of oryzalin on leishmania microtubules was verified by its inhibition of in vitro polymerization of leishmania microtubules, but not control mammalian microtubules (from rat brain). These findings indicate that microtubules play a major role in leishmania proliferation, maintenance of cell shape, and cytodifferentiation.

The cell envelope of the hyperthermophilic archaebacterium Pyrobaculum organotrophum consists of two regularly arrayed protein layers: three‐dimensional structure of the outer layer

The cell envelope of the hyperthermophilic sulphur-reducing archaebacterium Pyrobaculum organotrophum H10 was found to be composed of two distinct hexagonally arranged crystalline protein arrays. Electron microscopic analysis of freeze-etched cells and isolated envelopes in conjunction with image processing showed that the inner layer (lattice centre-to-centre spacing 27.9 nm) is essentially identical to the protein array of Pyrobaculum islandicum GEO3, a complex, rigid structure implicated in the maintenance of cell shape. The outer layer has clear p6 symmetry and a lattice spacing of 20.6 nm. Its three-dimensional structure was reconstructed from a negative stain tilt series of an intact double-layered envelope using Fourier filtration to separate the desired information from the other lattices present. The outer layer is a unique, porous network of blocklike dimers disposed around six-fold axes, and exhibits minimal asymmetry between its inner and outer faces. It appears to be rather loosely associated with the outer surface of the inner layer. In most H10 envelopes, the inner layer is orientated with one base vector exactly perpendicular to the long axis of the cell, so that the cylindrical portion is composed of a series of parallel cell-girdling hoops of hexameric morphological units. All the other known Pyrobaculum strains were found to have a GEO3-type envelope structure, consisting of a single rigid protein array and a fibrous capsule. Although H10 does not possess a capsule, fibrils appear to be sandwiched between the two protein layers.

Steroid-induced epithelial-fibroblastic conversion associated with syndecan suppression in S115 mouse mammary tumor cells.

Cell-matrix interactions play an important role in the maintenance of cell shape, supposed to be mediated by the anchorage of cellular cytoskeleton to extracellular matrix via matrix receptors. In this work the expression of one of the known matrix receptors, syndecan, was studied during the hormone-induced change in the phenotype of Shionogi 115 (S115) mouse mammary tumor cells. In the presence of testosterone, when S115 cells express fibroblastic phenotype, they increased their growth rate and became gradually anchorage independent. These cells, however, revealed strong RGDS-dependent binding to fibronectin (FN) but not binding to the heparin-binding domain of FN. Instead, S115 cells growth without testosterone showed epithelial morphology and binding to the heparin-binding domain of FN, suggesting an alteration of syndecan expression in hormone-treated S115 cells. As quantitated by radioimmunoassay and by Western blot, the amounts of both matrix-binding ectodomain of syndecan and syndecan mRNA (2.6 kb) declined in hormone-treated S115 cells. The addition of antiandrogen cyproterone acetate to culture medium opposed the effect of testosterone on syndecan mRNA. We thus propose that the inactivation of syndecan gene and the consequent suppression of syndecan expression is related to the altered adhesion properties, the disappearance of epithelial phenotype, and, on the other hand, to the appearance of transformed-like phenotype in hormone-treated S115 cells.

The Structure and Function of Candida albicans Cell Wall

The wall and outer surface of pathogenic fungi is involved in adhesion and cell shape maintenance, and acts as a barrier to metabolites and drugs. In addition the cell wall contains immunogenic determinants, immunomodulators and secreted enzymes. The cell wall of C. albicans comprises some 30% of the dry weight of the cell and is composed predominantly of mannoproteins and β-glucans with small quantities of chitin, protein and lipid. Transmission electronmicroscopy of the wall reveals a number of different layers; the thickness and number of these layers varies during germ-tube formation and the cell cycle. The outer fuzzy layer of the wall contains receptors that bind plastic, laminin, fibrinogen, and C3 fragments of complement. The β1, 3 and β1, 6 glucans and chitin are the polymers which endow rigidity on the cell wall. These polymers form a microfibrillar scaffolding upon which other macromolecules, particularly mannoproteins are bound.The wall mannoproteins are highly branched polysaccharides primarily composed of mannose units attached to protein through a G 1 cNac dimer bridge and asparagine. There are also smaller numbers of 0-linked mannoproteins where the mannose units are attached to protein through serine and threonine residues. The mannoproteins have a backbone of α-1, 6 linked mannose units to which side chains of mannoses attach via α-1, 2-and occasionally α-1, 3 bonds. The side chains are the major antigenic determinants and the A and B serotypes are distinguished by differences between these side chains. A model for cell wall growth and development will is described.

Organization of microtubules in cochlear hair cells

The organization of microtubules in hair cells of the guinea-pig cochlea has been investigated using transmission electron microscopy and correlated with the location of tubulin-associated immunofluorescence in surface preparations of the organ of Corti. Results from both techniques reveal consistent distributions of microtubules in inner and outer hair cells. In the inner hair cells, microtubules are most concentrated in the apex. Reconstruction from serial sections shows three main groups: firstly, in channels through the cuticular plate and in a discontinuous belt around its upper perimeter; secondly, forming a ring inside a rim extending down from the lower perimeter of the plate; and thirdly, in a meshwork underlying the main body of the plate. In the cell body, microtubules line the inner face of the subsurface cistern and extend longitudinally through a tubulo-vesicular track between the apex and base. In outer hair cells, the pattern of microtubules associated with the cuticular plate is similar, although there are fewer present than in inner hair cells. In outer hair cells from the apex of the cochlea, microtubules occur around an infracuticular protrusion of cuticular plate material. In the cell body, many more microtubules occur in the region below the nucleus compared with inner hair cells. The possible functions of microtubules in hair cells are discussed by comparison with those found in other systems. These include morphogenesis and maintenance of cell shape; intracellular transport, e.g., of neurotransmitter vesicles; providing a possible substrate for motility; mechanical support of structures associated with sensory transduction.

Action of general and α-smooth muscle-specific actin antibody microinjection on stress fibers of cultured smooth muscle cells

Arterial smooth muscle cells express α- and γ-smooth muscle, as well as β- and γ-cytoplasmic actins. Two actin antibodies, one recognizing smooth muscle and cytoplasmic actin isoforms, the other recognizing specifically α-smooth muscle actin, were microinjected into cultured aortic smooth muscle cells. The effect of these antibodies on stress fiber organization was examined by staining with rhodamine-labeled phalloidin and by immunofluorescence with the same antibodies. Microinjection of the general actin antibody abolished most of the stress fiber staining with all reagents, but did not significantly affect the shape of the injected cells. This suggests that stress fiber integrity is not absolutely necessary for the maintenance of cell shape within the time of observation. Microinjection of the specific α-smooth muscle antibody abolished to various extents the staining of stress fibers with this antibody, but left practically intact their staining with rhodamine-labeled phalloidin and with the general actin antibody. This suggests that the incorporation of α-smooth muscle actin is not absolutely necessary for the maintenance of stress fiber integrity in cultured smooth muscle cells.

Involvement of cell shape in the replication and segregation of chromosomes in Escherichia coli

Chromosome replication appears to initiate in E. coli when the dnaA boxes in oriC become filled with DnaA protein, which could simultaneously mediate both the unwinding of the origin for the start of polymerization and the attachment of oriC to the cell envelope (Bramhill and Kornberg, 1988; Løbner-Olesen et al., 1989; Pierucci et al., 1989). The attachment takes place somewhere within the cell half in which the oriC resides. The boundaries of this attachment/replication zone, which cannot include the polar cap, could be demarcated by the polar and centrally located periseptal annuli (Rothfield, this Forum). Since attachment and polymerization are two aspects of the same process, the attachment probably takes place via the polymerizing strand. Once polymerization begins, the oriC with the older template strand moves away from the younger one, by mechanisms unknown, to eventually take up residence in the equivalent domain of the complementary sister cell. Thus, the template strand that stays within its domain corresponds to the strand that was attached during the previous round of replication, and the template that moves away is the one that was not attached. The driving force for this translocation is not specified by our model, but a number of plausible alternatives have been proposed by others (reviewed in Leonard and Helmstetter, 1990). Throughout the ensuing replication and cell division, the chromosomes are located (or can move freely) within the attachment/replication zone of the developing daughter cell (lateral cylinder and septum). At some time during the course of this process, but before the next initiation event, the replication origins must be released from the attachment sites so that the entire process can be repeated. Thus, the probabilistic non-random chromosome segregation is due to the asymmetry of the attachment/replication zone in the cell, whereas the partitioning system itself must possess a mechanism to discriminate between template strands of different ages. This apparent mechanistic relationship between chromosome replication, chromosome partitioning and the maintenance of cell shape may provide an interesting framework for future experiments.

Breakthrough in microtubule research

A breakthrough occurred in microtubule research during 1988 by the demonstration of Koshland and co-workers using a microtubule-chromosome model system. They discovered that chromosome movement towards the cell pole during the anaphase of cell division is both powered and regulated by microtubule depolymerization at the chromosome end of the microtubule [1]. The experiment has bearings on further directions of basic research on cell division [2]. It is thus timely to summarize past and present linkages between microtubules and clinical practice. Microtubules are tube-shaped structures with an outer diameter of approximately 24 nm and a variable length, sometimes reaching 10–25 μm. The main building block is a dumb-bell shaped dimer protein composed of α-tubulin and β-tubulin. Microtubule-associated proteins (MAPs) and guanosine nucleotides, GTP and GDP, are essential for microtubule structure, stability and function [3, 4]. Microtubules are ubiquitous cell structures mainly involved in the formation and maintenance of cell shape and in cell transport phenomena such as chromosome separation in cell division, transport within the axon of the nerve cell, leucocyte chemotaxis and phagocytosis, cilial and flagellar movements [3–6]. The mitotic spindle is mainly composed of microtubules. Low concentrations of microtubule depolymerizers, such as colchicine, block cell division in prometaphase, the classical and conspicuous picture of ‘metaphase arrest’. Interestingly, the microtubule polymerizer taxol, which has an opposite mechanism of action, blocks cell division even more efficiently at the same stage of the cell cycle [7]. It should be emphasized that the effect of microtubule depolymerizers is not restricted to the mitosis stage of the cell cycle. Inhibition was demonstrated both in the G1 stage and G2 stage of the interphase between two cell divisions [3, 8]. Another unexplained effect of microtubule depolymerizers is that DNA synthesis was stimulated in several cell systems. The microtubule polymerizer taxol lacked such effects on interphase cells [3]. Much of the information available on the biological properties of microtubules has been derived from experiments with microtubule depolymerizers (colchicine, Catharantus derivatives, podophyllotoxin, griseofulvin), each with specific binding sites on the tubulin molecule [3]. These substances have played a rôle in current anti-inflammatory, antiviral, antifungal and antineoplastic chemotherapy. In this context, it is pertinent to remember that colchicine is the classical remedy for acute gout [3]. The mechanism of action is likely to be inhibition of neutrophil leucocyte chemotaxis due to suppression of the release of signal substances from phagocytosing leucocytes. The antifungal agent griseofluvin is as efficient as colchicine in the treatment of acute gout and lacks the serious side-effects of colchicine [cf. 9]. A semisynthetic podophyllin preparation (SPG) has been used in Scandinavia as an alternative to other cytotoxic agents in the treatment of severe rheumatoid arthritis. An interphase block in the G2 stage was found in bone marrow cells of patients treated with the preparation [10]. The observation does not exclude metaphase arrest at higher concentrations. Pure podophyllotoxin, which is stable in solution, has now replaced crude podophyllin preparations in the treatment of venereal warts, condyloma accuminatum [11]. An analogous development would seem desirable in anti-rheumatoid therapy [12]. In antineoplastic chemotherapy, a colchicine derivative (demecolchine, Colcemid®) was introduced for the treatment of leukaemia in the 1950s. The launch failed, but the substance was more convenient and stable than the parent compound and became popular in research laboratories and in the laboratories of clinical genetics for the accumulation of mitoses and improvement of chromosome morphology. The latter phenomenon is thought to be due to chromosome stretching by microtubule depolymerization [7], analogous to chromosome transport originally observed by Inové [13], nuclear segmentation of interphase cells [14], and chromosome transport in the new microtubule-chromosome model [1, 2]. The part of the microtubule inhibitors on the antineoplastic market was conquered by the Catharantus derivatives, formerly called Vinca alkaloids, in the 1960s [15]. Since then, modifications of the original molecules have emerged and disappeared. A fascinating metamorphosis is that oestramustine, originally thought to be an alkylating agent directed to oestrogen receptors by the hormone part of the molecule, is now considered to be a microtubule inhibitor with a specific action on microtubule-associated proteins [cf. 6]. The common denominator in microtubule depolymerizer therapy of inflammatory, viral, fungal, and neoplastic diseases is interference with growth and communication of key cells in the pathological process. The new microtubule-chromosome model [1, 2], together with other recent advances in the investigation of microtubule biology [16, 17], is expected to provide a tool for the improvement of such therapy.

Dependence of proliferation of Bacteroides forsythus on exogenous N-acetylmuramic acid.

Bacteroides forsythus is the first wild-type or mutant bacterium found to require exogenous N-acetylmuramic acid for proliferation and maintenance of cell shape. This implies so far unknown pathways for peptidoglycan synthesis and a strict dependence of B. forsythus on other bacteria in its oral habitat. Addition of N-acetylmuramic acid to conventional bacteriological media allows routine cultivation of this fastidious organism.

Chromosomal assignment of two myosin alkali light-chain genes encoding the ventricular/slow skeletal muscle isoform and the atrial/fetal muscle isoform (MYL3, MYL4).

In all eukaryotes, myosin plays a major role in the maintenance of cell shape and in cellular movement; in association with actin and other contractile proteins it is also a major structural component of the muscle sarcomere. Several isoforms of myosin alkali light chain have been identified, associated with different muscle types. We have recently localized the gene encoding the fast skeletal muscle alkali light-chain isoforms MLC1F and MLC3F (HGM symbol, MYL1) to human chromosome 2q32.1-qter (Cohen-Haguenauer 1988). We present here the chromosomal assignment of two loci encoding the ventricular muscle isoform MLC1V (equivalent to the slow skeletal muscle isoform MLC1Sb) and the atrial muscle isoform MLC1A (equivalent to the fetal isoform MLC1emb) using a panel of 25 independent man-rodent somatic cell hybrids. The MLC1V gene (HGM symbol, MYL3) was mapped to human chromosome 3 using a human full-length cDNA probe that hybridizes to a single major human TaqI2.8-kb fragment. The MLC1A probe (HGM symbol, MYL4) was a 360-bp mouse cDNA fragment that gave a distinct signal with human DNA using low stringency conditions of hybridization and washings and after presaturation of the Southern blots with rodent DNA. A single PstI 7.8-kb fragment gives an intense signal, and its presence correlates with the presence of chromosome 17 among the hybrids. These data are in keeping with the localizations of the MLC1V gene to mouse chromosome 9, and of the MLC1A gene to mouse chromosome 11, which share some markers in common with human chromosomes 3 and 17 respectively.

Transient reduction in erythrocyte membrane sialoglycoprotein β associated with the presence of elliptocytes

Erythrocyte membranes from an anaemic patient receiving gold therapy for rheumatoid arthritis had reduced beta-sialoglycoprotein (beta-SGP) content but normal expression of sialoglycoproteins alpha, delta and gamma. Elliptocytes were present in the peripheral blood. The serum of the patient contained anti-beta-SGP which did not appear to bind to her own cells. It reacted with all erythrocytes apart from beta-SGP deficient Leach phenotype cells. The antibody was inhibited by purified beta-SGP from normal red cells, bound to beta-SGP on immunoblots and also reacted with the abnormal beta-related-SGP in erythrocyte membranes of both the Gerbich type and Yus type of Gerbich negative. Two years later the patient was no longer anaemic, no elliptocytes were seen in her peripheral blood film and her erythrocyte membranes had normal beta-SGP content. Antibody was no longer present in her serum and antibody from the earlier sample now reacted with the patient's erythrocytes. Erythrocyte membrane beta-SGP is known to be important in the maintenance of normal cell shape. It is likely that the transient occurrence of elliptocytes in the patient resulted from the concurrent temporary reduction in beta-SGP content of her erythrocyte membranes.

Transient reduction in erythrocyte membrane sialoglycoprotein β associated with the presence of elliptocytes

Erythrocyte membranes from an anaemic patient receiving gold therapy for rheumatoid arthritis had reduced β‐sialoglycoprotein (β‐SGP) content but normal expression of sialoglycoproteins α, δ and γ. Elliptocytes were present in the peripheral blood. The serum of the patient contained anti‐β‐SGP which did not appear to bind to her own cells. It reacted with all erythrocytes apart from β‐SGP deficient Leach phenotype cells. The antibody was inhibited by purified β‐SGP from normal red cells, bound to β‐SGP on immunoblots and also reacted with the abnormal β‐related‐SGP in erythrocyte membranes of both the Gerbich type and Yus type of Gerbich negative. Two years later the patient was no longer anaemic, no elliptocytes were seen in her peripheral blood film and her erythrocyte membranes had normal β‐SGP content. Antibody was no longer present in her serum and antibody from the earlier sample now reacted with the patient's erythrocytes.Erythrocyte membrane β‐SGP is known to be important in the maintenance of normal cell shape. It is likely that the transient occurrence of elliptocytes in the patient resulted from the concurrent temporary reduction in β‐SGP content of her erythrocyte membranes.

Impaired echinocytic transformation of ankyrin- and spectrin-deficient erythrocytes in mice.

The membrane skeleton of the red blood cell plays an important role in the determination of cell deformability and cell shape. Under various in vitro conditions, red blood cells undergo an echinocytic or stomatocytic shape transformation. The mechanism of this fundamental process is not well understood. We have studied the red cell shape transformation in normoblastic anemia mice (nb/nb) and spherocytic anemia mice (sph/sph), which are deficient in ankyrin and spectrin, respectively. We found that both ankyrin-deficient cells (nb/nb) and spectrin-deficient cells (sph/sph) have a reduced capacity to undergo echinocytic transformation with various echinocytogenic treatments, that is, incubation with sodium salicylate (40 and 120 mM), calcium loading (50 microM A23187 + 2.2 mM Ca2+), or metabolic depletion (24 hr at 37 degrees C). These results suggest that the functional integrity of the membrane skeleton is essential for the maintenance and transformation of the red cell shape.

Secretion of Protein-bound Hydroxyproline from Moss Callus Cells

Abstract A glycoprotein rich in hydroxyproline was found in wall preparations of callus cells of the moss Physcomitrium pyriforme Brid. It is apparently attached to the non-cellulosic polysaccharides of the wall, and the majority is extractable by boiling the wall fraction or by using a chaotropic salt at room temperature. A pulse-chase technique was used to study the transport of this protein to the wall. Cytochalasin B seems to inhibit its secretion from the callus cells. Some of this wall-associated protein is probably secreted from the cells into the medium. Electron microscopic evidence shows vesicular activity in the cytoplasm and secretion and incorporation into the wall layers (not shown). The protein probably functions in the maintenance of cell shape in the moss callus tissue, being partly secreted out of the callus cells before the filamentous morphology appears.