Ceramide kinase targeting and activity determined by its N-terminal pleckstrin homology domain

The N-terminus of ceramide kinase (CERK) is thought to be myristoylated and to contain a pleckstrin homology (PH) domain. We found that deletion of this region (DeltaPH-CERK) ablates activity. This is not due to prevention of N-terminal myristoylation since a G2A CERK mutant, which cannot be myristoylated, was active. CERK was able to bind liposomes, as well as the isolated unmyristoylated PH domain; DeltaPH-CERK was not. Upon analysis of EGFP-tagged proteins, CERK was found associated with the Golgi complex. Osmotic cell swelling induced translocation of CERK to the plasma membrane and formation of large vesicles displaying bound CERK. None of these features occurred with DeltaPH-CERK, which remained disseminated throughout the cytoplasm. These findings show that the PH domain of CERK is essential for localization, translocation, and activity of this lipid kinase.     

Coupling between the initiation of DNA replication and cell division in the blue-green alga Anacystis nidulans

The effect of mitomycin C on cell mass increase, cell division, RNA synthesis and DNA synthesis in the blue-green alga Anacystis nidulans has been examined. Data suggests that the initiation of DNA replication, rather than its termination was the necessary event for cell division to occur.     

Rho GTPase–independent regulation of mitotic progression by the RhoGEF Net1

Neuroepithelial transforming gene 1 (Net1) is a RhoA-subfamily–specific guanine nucleotide exchange factor that is overexpressed in multiple human cancers and is required for proliferation. Molecular mechanisms underlying its role in cell proliferation are unknown. Here we show that overexpression or knockdown of Net1 causes mitotic defects. Net1 is required for chromosome congression during metaphase and generation of stable kinetochore microtubule attachments. Accordingly, inhibition of Net1 expression results in spindle assembly checkpoint activation. The ability of Net1 to control mitosis is independent of RhoA or RhoB activation, as knockdown of either GTPase does not phenocopy effects of Net1 knockdown on nuclear morphology, and effects of Net1 knockdown are effectively rescued by expression of catalytically inactive Net1. We also observe that Net1 expression is required for centrosomal activation of p21-activated kinase and its downstream kinase Aurora A, which are critical regulators of centrosome maturation and spindle assembly. These results identify Net1 as a novel regulator of mitosis and indicate that altered expression of Net1, as occurs in human cancers, may adversely affect genomic stability.     

Novel cnidocysts of narcomedusae and a medusivorous ctenophore, and confirmation of kleptocnidism

Cnidocysts have been examined from the tentacles of the ctenophore Haeckelia rubra (Euchlora rubra) and five species of hydrozoan narcomedusae (Solmundella bitentaculata, Aegina citrea, Solmissus marshalli, Solmissus albescens, and Cunina sp.) using TEM, both in sections and by firing whole cnidocysts onto EM grids. The study revealed that these apotrichous isorhiza cnidocysts have a novel morphology in which the intracapsular inverted tubule has five circumferential pleats when viewed in transverse section, rather than the usual three pleats. Accordingly, the definition of helicoptychoneme cnidocysts has been broadened to include both the usual three-pleated cnidocysts and these new five-pleated cnidocysts. In general, apotrichous isorhizas have subspherical capsules with a thick, bilayered wall, whose interior is nearly filled with the regularly coiled, helically folded, five-pleated inverted tubule. Upon discharge, the everted tubule is several mm long and the five circumferential pleats become manifested as five helical rows of spines running up the tubule, which has three morphologically different segments. The very short basal segment is devoid of ornamentation; the remaining proximal portion is characterized by five spirals of uniform, closely packed short spines; the long distal portion is characterized by a single spiral of regularly spaced large spines that derive from all five spirals-the five spirals are otherwise demarcated in the distal portion by 'scales' that are visible only with the electron microscope.     

Comparison of chemical dehydration and critical point drying for the stabilization and visualization of aging biofilm present on interior surfaces of PVC distribution pipe

In this study, fixation of attached glycocalyx on the interior surfaces of polyvinyl chloride distribution pipe remnants was compared with and without ruthenium red/osmium tetroxide and, in the final preparatory phase, with chemical dehydration and critical point drying. SEM examination of interior surface of the polyvinyl chloride pipe showed varying concentrations of adherent bacteria, depending on the preparatory technique used. It was concluded that using a combination of ruthenium red/osmium tetroxide and critical point drying is the optimum method for visually demonstrating aging biofilm on the interior surface of contaminated polyvinyl chloride pipe.     

THE STRUCTURE OF THE COLLODION MEMBRANE AND ITS ELECTRICAL BEHAVIOR : X. AN EXPERIMENTAL TEST OF SOME ASPECTS OF THE TEORELL AND MEYER-SIEVERS THEORIES OF ELECTRICAL MEMBRANE BEHAVIOR

1. The Teorell, Meyer-Sievers theory characterizes the electrochemical behavior of membranes by their selectivity constant "A_p" which is derived conventionally from concentration potential measurements at various concentration levels. The selectivity constant may, however, be derived also from entirely independent, different experimental data, namely base exchange studies. The constants arrived at in this second way are designated as "A_b." The selectivity constants derived by these two methods must be in reasonable, at least semiquantitative agreement if the basic assumptions of the theory are correct. 2. The selectivity constants A_p and A_b were determined for eleven different sets of membranes of different electrochemical activity and of different (8.2 to 80 volume per cent) water content. 3. The potentiometric selectivity constants A_p are in most cases several orders of magnitude greater than the corresponding A_b values. With membranes of great porosity and high electrochemical activity the A_b values approach at least in order of magnitude the A_p values. 4. It is concluded that the unexpectedly large discrepancy between the A_p and A_b values is due to some inherent weakness of the Teorell, Meyer-Sievers theory, most likely to its neglect of any structural factors.