Stereologic analysis was made of cell surface membrane (PM) and two interrelated cytoplasmic membrane systems, the vacuole membranes (VM) and small vesicle membranes (SVM) . Volumes and surface areas of the three membrane compartments were measured during steady-state pinocytosis, when membrane recycling is rapid, and during phagocytosis, when a shift to a lower rate of membrane uptake by endocytosis occurs (B . Bowers, 1977, Exp. Cell Res . 110:409) . Total membrane area in the three compartments was 3.2 pmt/gm3 of protoplasmic volume and was constant throughout the experiments. In pinocytosing cells, 32% of the membrane was in the PM, 25% in the VM, and 43% in the SVM . The vacuole compartment occupies -20% of the total cell volume, and the small vesicle, -3%. As the endocytic uptake of membrane from the surface decreased, there was an increase in PM area and a marked decrease in SVM area . The VM area remained constant even though "empty" vacuoles were almost completely replaced by newly formed phagosomes within 45 min . This demonstrates directly a rapid flux of membrane through this compartment. A model, taking into consideration these and other data on Acanthamoeba, is proposed to account for the observed membrane shifts . The data suggest that the vacuolar (digestive) system of Acanthamoeba is central to cellular control of endocytosis and membrane recycling.Evidence from several different kinds of studies suggests that the bulk of cell surface membrane in free-living endocytic cells is in a reversible equilibrium with cytoplasmic vacuolar system (4,6,12,(15)(16)(17)(18). The rate of cell surface turnover in these cells is very high, on the order of minutes (4,17,18) . The dynamics of the membrane flow and the cytoplasmic membrane relationships are complex so that a complete definition of the pathway of membrane flow in a single system is not yet available. Even fewer clues exist as to how the cell modulates the process . We are using the small amoeba, Acanthamoeba castellanii as a model system for exploring these questions . Acanthamoeba is especially useful because it is easy to culture, it pinocytoses continuously and at a high rate, and the membrane composition appears to be simpler than that of mammalian cells (8).The present study seeks to establish basic information about the volume and membrane area in three interrelated membrane compartments that appear to comprise the bulk of the recirculating membrane in Acanthamoeba . The compartments examined were : plasma membrane, vacuolar membrane (the amoeba digestive system), and an associated membrane system THE JOURNAL OF CELL BIOLOGY " VOLUME 88 MARCH 1981 509-515 of small vesicles. We have made a morphometric analysis of electron micrographs of pinocytosing cells to establish a base line. We then determined shifts in the distribution of membrane in the three compartments when phagocytosis at a maximum rate was superimposed on pinocytosis . Phagocytosis pre-empts the pinocytic mechanism so that the rate of fluid uptake decreases (1). The to...

Infection of exponential-phase suspension cultures of mouse fibroblast cells (L-M) with equine abortion virus (EAV) resulted in inhibition of cell growth and marked alterations in host metabolic processes. The synthesis of deoxyribonucleic acid (DNA) and ribonucleic acid was inhibited within 4 hr after infection and was suppressed by more than 90% by the time of maximal virus replication (14 to 18 hr). The overall rate of protein synthesis, however, was similar in uninfected and virus-producing cells as determined by measurements of net protein and isotope incorporation. The time course of viral DNA and protein synthesis and assembly into mature virus was determined with the inhibitors 5-fluorodeoxyuridine (FUdR) and cycloheximide, respectively. Thus, viral DNA synthesis was essentially completed at 14 hr, and viral protein and infectious virus synthesis was completed at 18 hr. Although the number of plaque-forming units (PFU) produced by FUdR-treated cells (10 3 to 10 4 PFU/ml) was at least 3 logs less than that produced by untreated cells, the yield of physical particles (as determined by electron microscopy) was approximately the same at 30 hr after infection. Besides being relatively non-infective, the particles produced in FUdR-treated cells appeared morphologically incomplete as they contained little or no nucleoid material.

RANDAXLL, CHARILES C. (University of MIississippi School of Mledicine, Jackson), LANELLE (X. GAFFORD, ROBERT W. I)ARLINGTON, AND) J.xAIES M. HYDE. Composition of fowlpox virus and inelusion matrix. J. Bacteriol. 87:939-944. 1964.-Inclusion bodies of fowlpox virus infection are especially favorable starting material for the isolation of virus and inclusion matrix. Electron

Aeciospores of the long-cycle heteroecious rust fungus, Cronartium fusiforme, were found to have an extremely thick cell wall with striking spicules protruding from it. The wall was readily degraded by commercial chitinase, but spicules were unaffected. Quiescent spores contained two nuclei with distinct nuclear membranes possessing many pores. Numerous membrane-bounded lipid bodies were found both in wild-type orange and in white mutant aeciospores. An abundance of irregularly ovoid mitochondria was present in quiescent spores. After glutaraldehydeosmium fixation, the surface of the mitochondria appeared to be covered with ribosomes or microtubules in a paracrystalline array, whereas after permanganate fixation only smooth outer mitochondrial membranes were noted. The latter fixative revealed abundant vesicular endoplasmic reticulum in the spore. Spores incubated at 20 C on agar produced one to five distinct germ tubes within 65 to 180 min. These thin-walled tubes exhibited varying degrees of branching, and reached a total hyphal length of 300 to 500 A prior to rupturing. Emergence of germ tubes took place through a pore in the spore wall and appeared to be mainly a physical flowing of cytoplasm from the spore into the germ tube without division of nuc!ei or other cell organelles. On completion of germination, the protoplasm of the germ tube contained both nuclei and nearly all of the other spore contents. Mitochondria had smooth outer membranes, were greatly elongated, and possessed distinct longitudinal cristae. A limited amount of rough endoplasmic reticulum was arranged parallel to the germ tube wall. Other organelles seen in germ tubes were lipid bodies, concentric membrane figures, and numerous ribosomes. Lipid bodies appeared smaller and fewer in number than in quiescent spores.

Ungerminated and germinated basidiospores and 2-day-old mycelial cultures of Lenzites saepiaria were similar in their fine structure. Fixation with glutaraldehyde, followed by osmium tetroxide, was far superior to permanganate. Cell organelles were seen in cytoplasm of spores and hyphae, and clamp connections were abundant in hyphal elements. Numerous lomasomes, vesicular bodies, and complex concentric membranes occurred in the cytoplasm and were often associated with the cell membrane or the dolipore membrane (parenthesome) of the septum. Endoplasmic reticulum was not found, but numerous ribosomes were seen; polyribosome groupings were frequently noted. The nucleus was bounded by a double membrane which contained few pores. Germinating spores exhibited one or more large osmiophilic bodies in association with a vacuole and membranous elements. Other than possessing a thin wall, the emerging germ tube was similar in structure to the parent spore.