haemolysin of Escherichia coli: Comparison of pore-forming properties between chromosome and plasmid-encoded haemolysins

Abstract Lipid bilayer experiments were performed with chromosome-encoded haemolysin of Escherichia coli . The addition of the toxin to the aqueous phase bathing lipid bilayer membranes of asolectin resulted in the formation of transient ion-permeable channels with two states at small transmembrane voltages. One is prestate (single-channel conductance 40 pS in 0.15 M KCl) of the open state, which had a single-channel conductance of 420 pS in 0.15 M KCl and a mean lifetime of 30 s. Membranes formed of pure lipids were rather inactive targets for this haemolysin. Experiments with different salts suggested that the haemolysin channel was highly cation-selective at neutral pH. The mobility sequence of the cations in the channel was similar if not identical to their mobility sequence in the aqueous phase. The single-channel data were consistent with a wide, water-filled channel with an estimated minimal diameter of about 1 nm. The pore-forming properties of chromosome-encoded haemolysin were compared with those of plasmid-encoded haemolysin. Both toxins share common features, oligomerize probably to form pores in lipid bilayer membranes. Both types of haemolysin channels have similar properties but different lifetimes.     

Experimentally induced true hermaphroditism in genetically female fowl

Summary Two types of hermaphroditism were experimentally induced in genetically female fowls by grafting of embryonic testes in embryos. Of the 27 hermaphrodites observed during the 8 months after hatching, 20 possessed a right testis and a left ovary and 7 a right testis and a left ovotestis. The testes and ovotestes contained seminiferous tubules with a more or less developed germ cell complement, attaining in many cases the early spermatid stage. The interstitial tissue was poorly functional, as shown by the absence of male secondary sex characters. The ovary or ovarian part of the ovotestes possessed numerous small ovarian follicles. The female arrangement of the plumage and the absence of spurs demonstrated the secretion of oestrogens. A mechanism is proposed for explaining this partial masculinization of genetically female gonads, a phenomenon which occurs during the period of embryonic sex differentiation, and is responsible for this experimental true hermaphroditism.     

Spermiogenesis in the rainbow trout (Salmo gairdneri)

In an ultrastructural study on the spermiogenesis of the rainbow trout (Salmo gairdneri R.) four spermatogenetic stages were identified. In young round spermatids, the nuclear chromatin was first heterogeneous (euchromatin and heterochromatin). Subsequently, it became more homogeneous and started to condense in the form of coarse granules and fibers and then into fibrils associated in ribbon-like elements which eventually partly fused together. During early spermiogenesis, a juxtanuclear vacuole appeared in the area where the nuclear envelope was specialized due to condensation of material between the two envelopes and a slight accumulation of nuclear material. This area was finally located in the anterior part of spermatids and spermatozoa; it probably plays a role during fertilization. A flagellar rootlet appeared early in spermiogenesis; it may play a role in the attachment of the flagellum to the nucleus since it persisted until the centriolar complex was definitively fixed in the implantation fossa. The flagellum did not display a plasma membrane and was first located in the cytoplasm, but when it was later extruded from the cell, it acquired a membrane. The cytoplasm was rich in ribosomes (free or in small groups) but poor in membranous organelles. The few mitochondria polarized around the centriolar complex were finally organized into an annular mid-piece. The spermatids remained connected by intercellular bridges until the end of spermiogenesis. The complexity of trout spermiogenesis is intermediate between that in poecilids and that in carp and pike, which have very simple spermatozoa. The role of the material from the nucleus and the cytoplasm reaching the Sertoli cell in the control of spermatogenesis has been discussed.     

Concerted generation of Ig isotype diversity in human fetal bone marrow

The human fetal bone marrow B cell compartment of 14- to 21-wk gestational age was examined phenotypically and with respect to Ig H chain commitment and diversity. A dramatic expansion of fetal marrow B cell pools at 16- to 18-wk gestational age characterizes a rapid and concerted chain of differentiation events. Transiently up to 1/4 of nucleated marrow cells are CD20+/CD21+ cells which begin to express surface Ig other than IgM. Limiting dilution analysis of EBV-infected marrow cells delineated a virtually exclusive commitment to IgM production until 15 wk and the absolute and relative number of these cells were small (approximately 5% of comparable adult values). In parallel to the rapid increase in total B cell pools size, cells committed and able to secrete any of the five Ig isotypes are generated by 16-wk gestational age and by 18 wk the frequencies of these cells rapidly reach levels typical for adult peripheral tissue such as blood or lymph node. Fetal L chain diversity always anticipated that observed in adult serum. In addition to rising pool sizes and diverse IgH expression, EBV transformability is a major variable during this period of B cell development with up to 2/3 of B lineage cells transformable, about half of which are pre-B cells. By 21-wk gestational age transformable pre-B cells have disappeared and (as in adult tissue) approximately 10 to 20% of CD20+ cells are transformable. The rapid, concerted expression of full H chain diversity during a narrow period in fetal development is unique to marrow and implies a lymphopoietic process in a privileged site rather than an immunologic differentiation event. During this event, the relative proportions between the different IgH classes expressed, resembled that found in adult tissue, perhaps suggesting that B cell inherent programming rather than only antigenic forces determine heavy chain choice. The staggered expression, early in postnatal life, of IgH regions 3' of the C mu locus may reflect regulatory functions rather than inherent immaturity of the B lineage.     

Linkage of PGK1 to X-linked severe combined immunodeficiency (IMD4) allows predictive testing in families with no surviving male

Summary We present a linkage map of DNA probes around the X-linked severe combined immunodeficiency (IMD4) locus at Xq11-13. DXS159 and PGK1 show no cross-overs with the disease locus (Lod 3.01 at = 0.00). The order of loci is DXS1-DXS106-(DXS159-PGK1-IMD4)-DXS72-DXYS1. Members of families whose carrier status has been established by X-inactivation patterns were included in the analysis. As the probe (pSPT/PGK), which is used for investigation of X-inactivation patterns, has been shown to be linked to the disease itself, it is possible to assign phase in mothers of sporadic cases who have been shown to be carriers, even when they have no surviving male offspring.     

A T7 promoter-specific, inducible protein expression system forBacillus subtilis

The adaptation and application of theEscherichia coli T7 RNA polymerase system for regulated and promoter-specific gene expression inBacillus subtilis is reported. The expression cassette used inBacillus subtilis was tightly regulated and T7 RNA polymerase (T7 RNAP) appeared 30 min after induction. The efficiency of T7 promoter-specific gene expression inB. subtilis was studied using one secretory and two cytosolic proteins of heterologous origin. The accumulation ofE. coli -galactosidase, as well as a 1,4--glucosidase fromThermoanaerobacter brockii inB. subtilis after T7 RNAP induction was strongly enhanced by rifampicin inhibition of host RNAP activity. The-amylase ofThermoactinomyces vulgaris, a secretory protein, was found to accumulate in the culture supernatant up to levels of about 70 mg/l 10–20 h after T7 RNAP induction, but was also deposited in cellular fractions. The addition of rifampicin inhibited-amylase secretion, but unexpectedly, after a short period, also prevented its further (intra)cellular accumulation     

Cholesteric-like crystal analogs in glucuronoxylan-rich cell wall composites: Experimental approach of acellular re-assembly from native cellulosic suspension

Summary Many plant cell walls are constructed according to a helicoidal pattern that is analog to a cholesteric liquid crystal order. This raises the question whether the wall assembly passes through a true but temporary liquid crystal state. The paper focuses on experiments performed from aqueous suspensions of extracted quince slime, i.e., a cellulose/glucuronoxylan wall composite that presents a helicoidal order when observed in situ, within the enlarged periplasm of the seed epidermal cells. Experiments carried out in acellular conditions showed that a spontaneous reassociation into a helicoidal order can be obtained from totally dispersed suspensions. The ultrastructural aspect of the reassembled mucilage suspension was different according to the resin used (LR White or nanoplast, a water-soluble melamin resin). It was always typically polydomain, and when an order was visible it was cholesteric-like and similar to the in situ native organization. Transition states with many imperfections expressed the difficulty of the system to reassemble in the absence of constraining surfaces. The possible intervention of glucuronoxylan (GX) in the ordered assembly of the microfibrils was checked by: (1) progressive extraction of GX by trifluoroacetic acid (TFA). The extraction was associated to a control of the fraction by analysis of uronic acid contents and observation at the electron microscope level. Extraction of GX provoked the formation of a flocculent mass, the flocculation being more intense when the TFA was more concentrated; (2) progressive change of pH in order to analyze the influence of pH on flocculation. Low pH (ca. pH 3) led also to a flocculation of the suspension, but the floc was reversibly lost after dialysis against distilled water. The results indicate the antifloc role of the GX due to the anionic charges carried by the side-chains. However, the function of GX as helper twisting agent in the cholesteric-like reassembly must not be ruled out.     

Chromosomal localization of the human histamine H1-receptor gene

We have assigned the human histamine H₁-receptor gene to chromosome 3 by Southern blot analysis of a chromosome mapping panel constructed from humanhamster somatic cell hybrids. This assignment was confirmed by in situ hybridization on metaphase chromosomes and involved bands 3p14–p21.     

Variation and taxonomy in Hordeum depressum and in the H. brachyantherum complex (Poaceae)

The variation pattern in the perennial Hordeum brachyantherum complex and in the annual H. depressum are described. The diploid form of H. brachyantherum s. lat., endemic to California in USA, previously recognized as a separate species is here treated as a subspecies ( H. brachyantherum ssp. californicum ). Despite its restricted distribution it shows a considerable variation and overlap in morphology with the tetraploid ssp. brachyantherum , and no unambiguous determination based on morphology between the two tax a is possible. The tetraploid cytotype has a large distribution area in western North America and easternmost Asia and a very wide morphological variation. It has also a small disjunct distribution area in easternmost Canada. A single hexaploid population from California is referred to ssp. brachyantherum .