Application of a new method for detecting the phenotype of target binding cells

Summary A new method was developed for detecting the phenotype of target binding cells (TBC) in a single-cell assay system. This methodology was evaluated during a clinical trial of recombinant interferon alfa-2a (rIFN alfa-2a) for the treatment of 10 metastatic renal cell carcinoma patients. Total TBC with K562 targets, HNK-1+ TBC, and HLA-DR+ TBC were quantitated during rIFN alfa-2a therapy. A significantly increased proportion of lymphocytes bound to target cells on day 9 of therapy bore the HNK-1 marker. This proportion subsequently declined to pretreatment levels. Total TBC paralleled the rise and fall in HNK-1+ TBC. HLA-DR+ TBC binding to targets remained constant and low throughout therapy. These findings suggest that rIFN alfa-2a early in therapy (day 9) caused the recruitment of additional HNK-1+ cells into binders. However, with continued therapy, this proportion reverts to pretreatment levels. The results of this clinical trial served to illustrate the ability of the modified single-cell assay system to detect TBC phenotype.Supported in part by Hoffman-La Roche, NIH grant CA 12582, and Jonsson Comprehensive Cancer Center grant CA 15866Dr. Figlin is a recipient of an American Cancer Society Junior Faculty Fellowship-JFCF 762-A     

Assignment of the prealbumin (PALB) gene (familial amyloidotic polyneuropathy) to human chromosome region 18q11.2–q12.1

Summary The assignment of the human prealbumin (PALB) gene to chromosome region 18q11–q12.1 has been achieved using a human genomic probe in the study of human-mouse somatic cell hybrids and by in situ hybridization. Because familial amyloidotic polyneuropathy was reported previously to be due to a mutation in prealbumin, it can be inferred that the gene for this disorder also maps to 18q11.2–q12.1.     

Surface changes in the naturally ankylosed teeth of the frog Rana pipiens during growth and maturation: an SEM study

An SEM study of the surface morphology of the major stages of mature and developing teeth of the leopard frog was made using anorganic preparations of the teeth and jaws. After initial development, the crown area changed little during subsequent tooth eruption, ankylosis and maturation. The thin enamel covering extended further down the shaft than expected. After ankylosis, the surfaces of the tooth continued to mature. The unmineralized gap between the crown and the pedestal, which is prominent in most amphibians, gradually filled in as the ankylosed tooth aged. The upper portion of the pedestal initially formed a dentine surface which was globular in appearance due to partial calcification of the surface collagen fibres but became smooth with uniformly calcified fibres as the ankylosed tooth matured. The lower portion of the pedestal was more variable and there was a gradual transition of dentine into a more cellular, bone-like tissue which contained lacunae and larger fibre bundles. This bone-like tissue was very distinct in surface morphology from the bone of the adjacent jaw, and as the tooth matured it changed from a coarse, woven appearance to one more like lamellar bone. Resorption bays were present in both the dentine and bony areas of teeth which were being shed. During development, the pedestal, which attaches the tooth to the jaw, formed as a separate calcification site and did not form a complete ring until fusion of its buccal surface with that of the overlying crown. A bony buccal lip formed early as part of the pedestal.     

An unusual wheat insertion sequence (WIS1) lies upstream of an alpha-amylase gene in hexaploid wheat, and carries a "minisatellite" array

Summary Comparison of the 5′ flanking regions of three α-amylase genes from chromosome 6B of hexaploid wheat by heteroduplex and sequence analysis revealed the presence of a 1.6 kb stem-loop insertion sequence (WIS1) in one of them. Polymorphism among hexaploid wheat varieties suggests the relatively recent insertion/excision of this sequence from its present position. The complete sequence of the stem-loop insertion shows that it has many of the features found in transposable elements, including target site duplication and terminal inverted repeats. One unusual feature is a tandem array of direct repeats comprising a wheat “minisatellite” sequence. Both the insertion sequence and the minisatellite are found at multiple locations in the wheat genome, but the functional significance of their association in WIS1 is unknown. The minisatellite arrays share a common core structure, and long arrays are polymorphic between different hexaploid varieties.     

Cloning and comparison of Aα mating-type alleles of the basidiomycete Schizophyllum commune

Summary An A mating-type allele (A4) was isolated by walking the chromosome from the closely linked PAB1 gene. A cosmid clone containing the A1 allele isolated from the walk was used as a probe to recover the A1 allele from another cosmid library. Cosmids encoding mating-type activity were identified by transforming Schizophyllum cells and screening for activation of A-regulated development. Putative mating-type transformants were confirmed in mating tests and genetic analyses of progeny. The identity of the specific alleles isolated was demonstrated by showing that their effectiveness in transforming for mating type is limited to recipient strains possessing an A allele different from the one encoded by the cloned sequences. Transforming DNA is active in trans, suggesting that A encodes a diffusible product. Restriction mapping shows that A1 and A4 are coded in the same physical region of the genome, but within a subregion that contains extensive sequence divergence. In addition, Southern analyses show that there is only one copy of A1 or A4 per haploid genome, and that they do not cross-hybridize to one another or to any of the other A alleles. A1 and A4 were subcloned as 2.8 and 1.2 kb fragments, respectively, retaining in transformation all the mating-type activity demonstrated of the original cosmids.     

Water use efficiency and carbon isotope composition of plants in a cold desert environment

Summary The effects of the availabilities of water and nitrogen on water use efficiency (WUE) of plants were investigated in a sagebrush steppe. The four species studied wereArtemisia tridentata (shrub),Ceratoides lanata (suffrutescent shrub),Elymus lanceolatus (rhizomatous grass), andElymus elymoides (tussock grass). Water and nitrogen levels were manipulated in a two-by-two factorial design resulting in four treatments: control (no additions), added water, added nitrogen, and added water and nitrogen. One instantaneous and two long-term indicators of WUE were used to testa priori predictions of the ranking of WUE among treatments. The short-term indicator was the instantaneous ratio of assimilation to transpiration (A/E). The long-term measures were 1) the slope of the relationship between conductance to water vapor and maximum assimilation and 2) the carbon isotope composition (¹³C) of plant material. Additional water decreased WUE, whereas additional nitrogen increased WUE. For both A/E and ¹³C, the mean for added nitrogen alone was significantly greater than the mean for added water alone, and means for the control and added water and nitrogen fell in between. This ranking of WUE supported the hypothesis that both water and nitrogen limit plant gas exchange in this semiarid environment. The short- and long-term indicators were in agreement, providing evidence in support of theoretical models concerning the water cost of carbon assimilation.     

Growth and respiration in two mangrove species at a range of salinities

Growth and dark respiration rates were measured in leaves and roots of seedlings of Avicennia marina (Forsk.) Vierh, (grey mangrove), and Aegiceras corniculatum (L.) Blanco (river mangrove). Plants were grown in a soil mixture at ambient temperatures and watered with 0.25 and 100% sea-water. Oxygen uptake was measured in excised root and leaf samples. In both species growth was maximal in 25% sea-water, and root respiration was lowest in 100% sea-water. Differences were found between the two species in the responses of leaf respiration to salinity. In A. corniculatum leaf respiration was raised in both 25 and 100% sea-water, while in A. marina only leaves in 100% sea-water showed higher rates of respiration. These results are consistent with the view that A. marina is the more salt-tolerant of the two species. In A. corniculatum the respiration rates of the hypocotyl were also measured, and were much higher in 100% sea-water than in the other two treatments. The results suggest that at high salinities there is a high metabolic cost in the shoots of both species, and that at such salinities rates of root respiration may be limited by the supply of substrate from the shoots.     

Effect on in Vitro Pollen Growth of an Isolated Style Glycoprotein Associated with Self-Incompatibility in Nicotiana alata

The effect on in vitro pollen tube growth of an isolated style glycoprotein (S₂-glycoprotein) associated with self-incompatibility in Nicotiana alata was investigated. Tube growth of pollen bearing the S₂-allele was inhibited, but tube growth of pollen bearing other alleles was not affected. Inhibition showed a dose response effect. The percentage of pollen grains that germinated was not significantly affected by the S₂-glycoprotein. Growth of S₂-pollen in the presence of the S₂-glycoprotein resulted in increased binding to the pollen of monoclonal antibody (PCBC3) which has a primary specificity for α-l-arabinofuranosyl residues. Growth of pollen bearing other alleles in the presence of the glycoprotein resulted in no increased binding of the antibody.     

Auxin Physiology of the Tomato Mutant diageotropica

The tomato (Lycopersicon esculentum, Mill.) mutant diageotropica (dgt) exhibits biochemical, physiological, and morphological abnormalities that suggest the mutation may have affected a primary site of auxin perception or action. We have compared two aspects of the auxin physiology of dgt and wild-type (VFN8) seedlings: auxin transport and cellular growth parameters. The rates of basipetal indole-3-acetic acid (IAA) polar transport are identical in hypocotyl sections of the two genotypes, but dgt sections have a slightly greater capacity for IAA transport. 2,3,5-Triiodobenzoic acid and ethylene reduce transport in both mutant and wild-type sections. The kinetics of auxin uptake into VFN8 and dgt sections are nearly identical. These results make it unlikely that an altered IAA efflux carrier or IAA uptake symport are responsible for the pleiotropic effects resulting from the dgt mutation. The lack of auxin-induced cell elongation in dgt plants is not due to insufficient turgor, as the osmotic potential of dgt cell sap is less (more negative) than that of VFN8. An auxin-induced increase in wall extensibility, as measured by the Instron technique, only occurs in the VFN8 plants. These data suggest dgt hypocotyls suffer a defect in the sequence of events culminating in auxin-induced cell wall loosening.     

The cell wall-plasmalemma interface in sieve tubes of barley

Both thick- and thin-walled sieve tubes in leaf-blade veins of Hordeum vulgare L. exhibit a distinct, electron-opaque inner wall layer after fixation in glutaraldehyde-osmium tetroxide and staining with uranyl acetate and lead citrate. This inner wall layer is thickest at the sieve plates and lateral sieve areas where it is permeated by a labyrinth of tubules formed by the plasmalemma. Along the lateral walls between sieve areas the inner wall layer apparently is penetrated by numerous microvilli-like evaginations of the plasmalemma, giving the cell wall-plasmalemma interface the appearance of a brush border. It is suggested that a similar brush-border-like structure may occur at the cell wall-plasmalemma interface of sieve elements in a wide variety of vascular plants.Abbreviation ER endoplasmic reticulum