STUDIES ON THE AUTECOLOGY OF THE MARINE DIATOM THALASSIOSIRA NORDENSKIOELDII. II. THE INFLUENCE OF CELL SIZE ON GROWTH RATE,AND CARBON,NITROGEN, CHLOROPHYLL a AND SILICA CONTENT1

The effect of cell size on growth rates and some cellular contents of Thalassiosira nordenskioeldii Cleve has been measured at 0 and 10 C. At 0 C the growth rate did not vary with cell size. The 2 smallest clones at this temperature had reduced growth rates because of the induction of sexuality in that size range. The clones grown at 10 C showed a significant negative relationship between growth rate and valve diameter with the cell surface area/volume ratio positively related to growth rate. At both temperatures the smaller cells had proportionately more carbon and nitrogen/unit cell volume. The amount of chlorophyll a and silica/unit cell surface area increased with increasing cell surface area at both 0 and 10 C. Both the C/N and C/chl a ratios showed no significant change with cell size at either temperature but there was a significant increase in the C/chl a ratio at 0 C. The C/Si ratio decreased with increasing cell size at both 0 and 10 C.     

Reconstitution of band 3, the erythrocyte anion exchange protein

Band 3, the erythrocyte membrane protein thought to be responsible for anion transport, was purified to near homogeneity using a Concanavalin A affinity column. Band 3 was then combined with egg lecithin, erythrocyte lipid, cholesterol, and glycophorin, the major erythrocyte sialoglycoprotein, to form vesicles capable of rapid sulfate transport. The transport activity was sensitive to prior treatment of the erythrocytes with pyridoxal phosphate-NaBH₄, a potent inhibitor of anion transport in these cells.     

Human β-glucuronidase: Assignment of the structural gene to chromosome 7 using somatic cell hybrids

-Glucuronidase (GUS) has become an important enzyme model for the genetic study of molecular disease, enzyme realization, and therapy, and for the biogenesis and function of the lysosome and lysosomal enzymes. The genetics of human -glucuronidase was investigated utilizing 188 primary man-mouse and man-Chinese hamster somatic cell hybrids segregating human chromosomes. Cell hybrids were derived from 16 different fusion experiments involving cells from ten different and unrelated individuals and six different rodent cell lines. The genetic relationship of GUS to 28 enzyme markers representing 19 linkage groups was determined, and chromosome studies on selected cell hybrids were performed. The evidence indicates that the -glucuronidase gene is assigned to chromosome 7 in man. Comparative linkage data in man and mouse indicate that the structural gene GUS is located in a region on chromosome 7 that has remained conserved during evolution. Involvement of other chromosomes whose genes may be important in the final expression of GUS was not observed. A tetrameric structure of human -glucuronidase was demonstrated by the formation of three heteropolymers migrating between the human and mouse molecular forms in chromosome 7 positive cell hybrids. Linkage of GUS to other lysosomal enzyme genes was investigated. -Hexosaminidase HEX _B) was assigned to chromosome 5; acid phosphatase₂ (ACP ₂) and esterase A₄ (ES-A ₄) were assigned to chromosome 11; HEX _A was not linked to GUS; and -galactosidase (-GAL) was localized on the X chromosome. These assignments are consistent with previous reports. Evidence was not obtained for a cluster of lysosomal enzyme structural genes. In demonstrating that GUS was not assigned to chromosome 9 utilizing an X/9 translocation segregating in cell hybrids, the gene coding for human adenylate kinase₁ was confirmed to be located on chromosome 9.Supported by NIH Grants HD 05196, GM 20454, and GM 06321, by NSF Grant BMS 73-07072, and by HEW Maternal and Child Health Service, Project 417.     

COLONY DEVELOPMENT IN EUDORINA ELEGANS (CHLOROPHYTA,VOLVOCALES)1

A detailed ultrastructure study was made of cell division and colony development in Eudorina elegans Ehrenberg. At the onset of cell division and prior to nuclear division the nucleus moved from the cell center to the cell surface. During nuclear division the nuclear membrane remained intact, except for openings occurring at the nuclear poles. The spindle microtubules appeared to arise from a MTOC-like (microtubule organizing centers) structure, while centrioles were absent from the nuclear poles. Following telophase, daughter nuclei formed which were separated by several distinct bands of endoplasmic reticulum. Cytokinesis occurred with formation of a cleavage furrow, associated with a typical phycoplast band of microtubules. However, cytokinesis was incomplete, resulting in formation of cytoplasmic bridges between the plakeal cells. Upon completion of up to five successive cell divisions, the plakea underwent inversion, which appeared to involve the production of colonial envelope material and rearrangement of cytoplasmic bridges. A new hypothesis concerning inversion is postulated based on these observations.     

State of water in extremely halophilic bacteria: Heat of dilution ofHalobacterium halobium

Summary Heat of dilution ofHalobacterium halobium was measured when thick pastes of the bacteria, harvested throughout a complete growth cycle, were lysed by mixing with 40 times their volume of water in a microcalorimeter. A series of comparative measurements was made with pastes of bacteria previously disrupted by freezing and thawing but otherwise identical to the pastes of whole bacteria. The frozen-thawed pastes gave endothermic values some 18% greater than those obtained with intact bacteria; the difference was highly significant. Evidence was obtained that the mechanical component of bursting did not contribute to the difference between whole and lysed bacteria. On the other hand, when a correction was applied for heat of mixing of intracellular salts with extracellular NaCl, such as occurs when the bacteria lyse, the difference between whole and disrupted organisms was largely eliminated from exponential phase halobacteria but not from those harvested in stationary phase. It is concluded that there is no evidence, as reflected in heat of dilution, of abnormal solution properties of the cytosol of young halobacteria, which are rich in potassium. On the other hand, and paradoxically, some doubt remains about stationary phase organisms whose cytosol has a much higher Na⁺ content (and Na/K⁺ ratio) than the cytosol of exponential phase bacteria.     

Growth of endothelial and HeLa cells on a new multipurpose microcarrier that is positive,negative or collagen coated

A new cell culture microcarrier that can be covalently bonded by cell attachment proteins and can be thin-sectioned for electron microscopy was synthesized. It was easily made by sulfonating cross-linked polystyrene beads for a negative surface charge followed by covalent attachment of polyethylenimine for a positive charge. Cell attachment proteins, e.g. collagen, was covalently bonded directly to the microcarrier using a carbodiimide or after activating the microcarrier surface with glutaraldehyde. HeLa-S3 cells attached, spread and grew to confluence more efficiently on the positive microcarriers and those coated with collagen than on the negative ones. Endothelial cells grew best on those with a negative surface charge. The nature of the microcarrier surface was not the only aspect involved in cell adhesion but also the type of serum proteins adsorbed. Qualitatively different proteins coated the microcarriers depending upon whether the carrier was negative, positive or coated with collagen. Comparison of various types of available microcarriers indicated that the modified cross-linked polystyrene beads used here were best for transmission and scanning electron microscopy. Endothelial cells grown on the microcarriers had the same ultrastructure as cells grown in monolayers in culture dishes. Of a variety of microcarriers tested the modified cross-linked polystyrene beads were the only ones that could be used for both ultrastructural and biochemical techniques.     

Lysosomal proliferation in rachitic avian intestinal absorptive cells following 1,25-Dihydroxycholecalciferol

Lysosomes in chick intestinal absorptive cells from rachitic (vitamin D-deficient) and vitamin D-replete animals were studied utilizing transmission electron microscopic histochemistry and ultrastructural morphometry. Absorptive cells from rachitic animals, serum calcium = 7.3±0.3 mg%, contained an average of 4.0±0.3 supranuclear lysosomes. In rachitic chicks sacrificed 9 hr post-injection of 1,25-dihydroxycholecalciferol, the active metabolite of vitamin D, the values for both serum calcium, 9.8 ± 0.2 mg%, and the number of apical absorptive cell lysosomes, 12.9±0.6, were increased over non-injected or vehicle-only injected animals. Lysosomes in vitamin D-replete absorptive cells were characterized by their intense staining with pyroantimonate, indicative of their high calcium content. The same organelles also produced a positive reaction for acid phosphatase. Rachitic lysosomes, also acid phosphatase positive, were only lightly stained with pyroantimonate. The lysosomal proliferation apparently induced by 1,25-dihydroxycholecalciferol may be a further indication that these organelles play a role in intestinal calcium transport and/or intracellular calcium homeostasis within the absorptive cell.     

Effect of potassium on the water potential, the pressure potential, the osmotic potential and cell elongation in leaves of Phaseolus vulgaris

The effect of potassium on the water potential, the osmotic potential and the pressure potential in younger and older leaves of Phaseolus vulgaris grown in hydroponic culture was studied. Inadequate potassium supply resulted in an increase of the osmotic potential. In the older leaves the water potential was raised, in the younger leaves the pressure potential was depressed in the treatment insufficiently supplied with potassium as compared with leaves with an adequate potassium supply. Cell size of the younger leaves was smaller in the treatment with the low K⁺ supply in comparison with the leaves well supplied with K⁺. Potassium had a beneficial effect on plant growth, especially on fresh matter production. The water status of leaves (water content, pressure potential, osmotic potential) responded more sensitively to potassium supply than dry matter production. Besides organic N and organic anions, K⁺ was the most abundant solute found in the press sap of the leaves. From the results it is concluded that K⁺ is indispensible for attaining an optimum potential (turgor) in young leaves which in turn has an impact on plant growth.     

Methylation analysis of extracellular polysaccharides from suspension-cultured cells of Nicotiana tabacum

The soluble extracellular polysaccharides (ECP) of suspension-cultured tobacco cells were fractionated by DEAE-Sephadex CC into seven sub-fractions. Su     

Changes in cell-wall polysaccharide composition of developingNitella internodes

Changes in the uronide, neutral-polysacharide, and cellulose composition of the cell wall ofNitella axillaris Braun were followed throughout development of the internodes and correlated with changes in growth rate. As the cells increased in length from 4 to 80 mm during development, the relative growth rate decreased. Cell wall thickness, as measured by wall density, increased in direct proportion to diameter, indicating that cell-wall stress did not change during elogation. Cell-wall analyses were adapted to allow determination of the composition of the wall of single cells. The total amounts of uronides, neutral sugars and cellulose all increased during development. However, as the growth rate decreased, the relative proportions of uronides and neutral sugars, expressed as percent of the dry weight of the wall, decreased, while the proportion of cellulose increased. The neutral sugars liberated upon hydrolysis ofNitella walls are qualitatively similar to those found in hydrolysates of higher plant cell walls: glucose, xylose, mannose, galactose, arabinose fucose and rhamnose. Only the percentage of galactose was found to increase in walls of mature cells, while the percentage of all other sugars decreased. The rate of apposition (g of wall material deposited per unit wall surface area per hour) of neutral polysaccharides decreased rapidly with decreasing growth rate during the early stages of development. The rate of apposition of uronides decreased more steadily throughout development, while that of cellulose, after an early decline, remained constant until dropping off at the end of the elongation period. These correlations between decreasing growth rate and decreasing rate of apposition of neutral sugars and uronides indicate that synthesis of these cell-wall components could be involved in the regulation of the rate of cell elongation inNitella.