Genetic and molecular analysis of the synaptotagmin family

Secretion is a fundamental cellular process used by all eukaryotes to insert proteins into the plasma membrane and transport signaling molecules and intravesicular proteins into the extracellular space. Secretion requires the fusion of two phospholipid bilayers within the cell, an energetically unfavorable process. A conserved repertoire of vesicle-trafficking proteins has evolved that function to overcome this energy barrier and temporally and spatially control membrane fusion within the cell. Within neurons, opening of synaptic calcium channels and subsequent calcium entry triggers synchronous synaptic vesicle exocytosis and neurotransmitter release into the synaptic cleft. After fusion, synaptic vesicles undergo endocytosis, are refilled with neurotransmitter, and return to the vesicle pool for further rounds of cycling. It is within this local synaptic trafficking pathway that the synaptotagmin family of calcium-binding synaptic vesicle proteins has been postulated to function. Here we review the current literature on the function of the synaptotagmin family and discuss the implications for synaptic transmission and membrane trafficking. Received 14 August 2000; received after revision 20 September 2000, accepted 14 October 2000     

The diversity of molecular motors: an overview

Rapid progress has recently been made in the identification and characterization of a large number of kinesin and myosin motor proteins. Recent work has uncovered roles for these motors in processes such as vesicle trafficking, cytoskeletal organization, and chromosome movements, to name a few. A series of reviews describing some of the significant advances in our understanding of the structure and function of myosins and kinesins is presented.     

Equilibrium constant for calcium ion and ascorbate ion

The combination of calcium and ascorbic acid in water at 25 degrees C has been examined by measuring the change of free calcium ion concentration as ascorbate was added in small increment to a solution of calcium. The data show clearly that complex formation between calcium ion and ascorbate ion occurred. At ionic strength mu = 0.1-0.2, the equilibrium constant of Ca++ and the singly-charged ascorbate ion has been measured to be 2.1 M-1. The precision of the result is better than 5% and the accuracy is estimated to be better than 20%. The application of the equilibrium constants is discussed.     

Phospholipid composition of Dipylidium caninum

The phospholipid composition of Dipylidium caninum has been studied. Chloroform-methanol-soluble fraction amounted to 2.4% and phospholipids to 0.5% of the wet weight of the parasite. Phosphatidyl choline and phosphatidyl ethanolamine represented the bulk of the phospholipids, whereas phosphatidyl serine, phosphatidyl inositol, lysolecithin and lysophosphatidyl ethanolamine were present in minor amounts. Sulfatides were also identified in this parasite.     

Phospholipid composition ofDipylidium caninum

Summary The phospholipid composition ofDipylidium caninum has been studied. Chloroform-methanol-soluble fraction amounted to 2.4% and phospholipids to 0.5% of the wet weight of the parasite. Phosphatidyl choline and phosphatidyl ethanolamine represented the bulk of the phospholipids, whereas phosphatidyl serine, phosphatidyl inositol, lysolecithin and lysophosphatidyl ethanolamine were present in minor amounts. Sulfatides were also identified in this parasite.     

The mannophosphoinositides of Nocardia asteroides.

The phospholipid of N. asteroides has been investigated. It was found to contain phosphatidyl ethanolamine, cardiolipin, phosphatidyl inositol and a family of mannophosphoinositides. Dimannophosphoinositides with 3 and 4 moles of fatty acid per phosphate residue represented the major glycophospholipids besides small amounts of other more glycosylated mannophosphoinositides.     

Identification and analysis of the gas vesicle gene cluster on an unstable plasmid ofHalobacterium halobium

In our efforts to elucidate the mechanism of high-frequency mutation ofHalobacterium halobium to a gas vesicle deficient state, we discovered insertions, deletions, inversions, and complex DNA rearrangements associated with a large endogenous plasmid, pNRC100. The rearrangements are mostly IS element-mediated, and when they occur in a region of pNCRC100 containing a cluster of thirteen genes, gas vesicle mutants result. We have characterized the structure and expression of this gas vesicle protein (gvp) gene cluster and demonstrated its requirement for gas vesicle synthesis and cell flotation by genetic transformation.     

The mannophosphoinositides ofNocardia asteroides

Summary The phospholipid ofN. asteroides has been investigated. It was found to contain phosphatidyl ethanolamine, cardiolipin, phosphatidyl inositol and a family of mannophosphoinositides. Dimannophosphoinositides with 3 and 4 moles of fatty acid per phosphate residue represented the major glycophospholipids besides small amounts of other more glycosylated mannophosphoinositides.     

In situ localization of ACTH receptor-like mRNA in molluscan and human immunocytes

Adrenocorticotropin hormone (ACTH) receptor-like messenger RNA was localized in molluscan hemocytes and human peripheral blood mononuclear cells by in situ hybridization using a digoxigenin-labelled bovine complementary DNA probe. These findings suggest that the ACTH receptor gene has been highly conserved during evolution. Moreover, these data represent further support for a relationship between the immune and neuroendocrine systems in invertebrates, as documented in our previous studies [1]. Received 25 September 1997; received after revision 12 November 1997; accepted 12 November 1997     

Lipid composition ofMicrosporum gypseum

Summary The lipid composition ofMicrosporum gypseum has been studied. The lipids amounted to 10.1% and phospholipids to 1.1% of the mycelial dry weight. Phosphatidyl choline, phosphatidyl serine and phosphatidyl ethanolamine were the major components, while lysophosphatidyl choline, and phosphatidyl inositol were present in smaller quantities. Neutral lipids consisted of monoglycerides, diglycerides, triglycerides, free and esterified cholesterol.Acknowledgments: Thanks are due to Prof. P. Talwar, Department of Microbiology of this Institute for the supply ofM. gypseum. Technical assistance of Mr Adarsh Kumar is acknowledged.     

Equilibrium constant for calcium ion and ascorbate ion

Summary The combination of calcium and ascorbic acid in water at 25°C has been examined by measuring the change of free calcium ion concentration as ascorbate was added in small increment to a solution of calcium. The data show clearly that complex formation between calcium ion and ascorbate ion occurred. At ionic strength =0.1–0.2, the equilibrium constant of Ca⁺⁺ and the singly-charged ascorbate ion has been measured to be 2.1 M^–1. The precision of the result is better than 5% and the accuracy is estimated to be better than 20%. The application of the equilibrium constants is discussed.     

Über die Basizität der Knochenmineralien

Summary Interpretations regarding the structure of the bone mineral, based on quantitative data as to the basicity of calcium phosphate, have been complemented by X-ray and electron microscopic investigations. The chemical nature of the bone material is thereby thought to be substituted apatite.

---

---

Ergänzende Bemerkungen zur Arbeit vonJ. Raaflaub, Exper.17, 443 (1961).     

Role of phospholipid transfer protein in rabbit lung development

A 36-kDa phospholipid transfer protein (PLT-P_R), which preferentially transfers phosphatidyl choline (PC) compared to phosphatidyl inositol (PI), was purified 827-fold from rabbit lung homogenate. Incorporation of cholesterol in unilamellar vesicles reduced the PC transfer activity of PLTP_R. Dipalmitoyl phosphatidyl choline uptake by alveolar type II cells was increased in the presence of the protein, and further enhanced in the presence of surfactant liposomes. However, a decrease in uptake was noted with cholesterol in host membranes. Incorporation of PI into host membranes had a low stimulatory effect on the process. All these effects were more pronounced in adult type II cells compared to premature, term and 3-day-old pups. Received 12 September 2001; accepted 11 October 2001     

Tailored substrates for studies of attached cell culture

Substrates for studies of the interactions of attached cells with extracellular matrix components are often prepared by allowing a protein to adsorb from solution onto a glass or polystyrene substrate. This method is simple and effective for many studies, but it can fail in cases that require rigorous control over the structure and composition of adsorbed protein. Self-assembled monolayers formed by the spontaneous ordering of terminally functionalized alkanethiols onto a gold substrate are a class of well-ordered substrates and provide a convenient method for tailoring substrates with ligands, proteins and other groups. Methods that can pattern the monolayers provide a general strategy to create substrates that control the size, shape and spacing of attached cells. This review illustrates recent work that has used these methods of surface chemistry to create tailored substrates for studies in cell biology. Received 14 November 1997; received after revision 10 March 1998; accepted 10 March 1998     

Mechanisms of self-incompatibility in flowering plants

Self-incompatibility is a widespread mechanism in flowering plants that prevents inbreeding and promotes outcrossing. The self-incompatibility response is genetically controlled by one or more multi-allelic loci, and relies on a series of complex cellular interactions between the self-incompatible pollen and pistil. Although self-incompatibility functions ultimately to prevent self-fertilization, flowering plants have evolved several unique mechanisms for rejecting the self-incompatible pollen. The self-incompatibility system in the Solanaceae makes use of a multi-allelic RNase in the pistil to block incompatible pollen tube growth. In contrast, the Papaveraceae system appears to have complex cellular responses such as calcium fluxes, actin rearrangements, and programmed cell death occurring in the incompatible pollen tube. Finally, the Brassicaceae system has a receptor kinase signalling pathway activated in the pistil leading to pollen rejection. This review highlights the recent advances made towards understanding the cellular mechanisms involved in these self-incompatibility systems and discusses the striking differences between these systems. Received 10 May 2001; received after revision 20 June 2001; accepted 20 June 2001     

A role for the hinge/ear domain of the β chains in the incorporation of AP complexes into clathrin-coated pits and coated vesicles

The clathrin-associated adaptor protein (AP) complexes drive the polymerization of clathrin in coated pits to form coated vesicles. It has previously been shown that the carboxyl-terminal hinge/ear domain of the β2 chain contains a binding site for clathrin and that removal of this domain from APs or from isolated β2 chains abrogates their ability to form clathrin coats in vitro. We show here that the hinge/ear domain is necessary for efficient incorporation of AP complexes into coated pits and coated vesicles in cells, a result that is consistent with the view that the β chains indeed provide an important interaction between the AP complexes and clathrin. Received 7 April 1997; received after revision 22 May 1997; accepted 28 May 1997     

Changes in phospholipid composition ofNocardia polychromogenes during temperature adaptation

Summary The effect of growth temperature on the phospholipid composition of Nocardia polychromogenes has been examined. When the growth temperature was decreased from 37°C to 27°C, there was a large decrease in phosphatidyl ethanolamine with an increase in cardiolipin and phosphoinositides. These changes are discussed in context with the control of membrane fluidity.Acknowledgment. This investigation was financed in part by a grant from the Indian Council of Medical Research. Technical assistance of Mr Adarsh Kumar is acknowledged.     

Triadin: a multi-protein family for which purpose?

Triadin is a protein first identified as a member of the muscle calcium release complex, involved in calcium release for muscle contraction. However, its precise function in this complex is still undefined. Recently, triadin has been shown to be a multi-protein family, with different distribution of the various splice variants within the sarcoplasmic reticulum, raising the possibility of multiple functions for this family of polypeptides. Such functions may include involvement in excitation-contraction coupling, in triad targeting, in structural function or in muscle differentiation. The putative role(s) of triadin(s) will be discussed here.Received 5 May 2004; received after revision 4 June 2004; accepted 7 June 2004