A Kinome RNAi Screen Identified AMPK as Promoting Poxvirus Entry through the Control of Actin Dynamics

Poxviruses include medically important human pathogens, yet little is known about the specific cellular factors essential for their replication. To identify genes essential for poxvirus infection, we used high-throughput RNA interference to screen the Drosophila kinome for factors required for vaccinia infection. We identified seven genes including the three subunits of AMPK as promoting vaccinia infection. AMPK not only facilitated infection in insect cells, but also in mammalian cells. Moreover, we found that AMPK is required for macropinocytosis, a major endocytic entry pathway for vaccinia. Furthermore, we show that AMPK contributes to other virus-independent actin-dependent processes including lamellipodia formation and wound healing, independent of the known AMPK activators LKB1 and CaMKK. Therefore, AMPK plays a highly conserved role in poxvirus infection and actin dynamics independent of its role as an energy regulator.     

Myxosporida (Protozoa): The determination and maintenance of spore size and shape

Data are presented which suggest that the size of fish myxosporidan spores is primarily determined by the parasite's development in the tissues or organ's cavities and the spore's shape is determined by the presence of physiologically and behaviorally suitable fish.     

Genetic heterogeneity and pathology of congenital nephrotic syndrome and its significance in prenatal diagnosis

Familial occurrence of congenital nephrotic syndrome (CN) has been most extensively studied in Finland where it is known to be inherited as an autosomal recessive disease. In this Finnish type of congenital nephrotic syndrome (CNF) prenatal diagnosis by alphafetoprotein (AFP) determination in the amniotic fluid has been reported to be conclusive. Within the last six years we observed 10 cases of CN occurring in 5 families. In all patients renal histology has been studied. Although each family presented a different type of CN, the appearance within the same sibship was always identical.     

Cell cycle dependent changes of chromosomes in mouse fibroblasts.

Mouse fibroblast interphase nuclei stained with quinacrine dihydrochloride show distinctive differences in their fluorescent characteristics analogous to those which we have already observed in human and Syrian hamster cells. These patterns reflect the position of any given nucleus within the cell cycle. The brightly fluorescent chromocenters in the mouse nuclei were found to be in absolute aggreement with those stained by the C-banding technique, indicating that they represent centromeric heterochromatin. Furthermore, their number and size per nucleus were shown to vary in relation to the progress of the cell cycle.     

Zschokkella (Protozoa: Myxosporida) in macrourid fishes

Six species of Zschokkella, three new, were recovered from eleven genera and forty-three species of macrourid fishes found in both the Atlantic and Pacific Oceans and many adjacent seas. The constancy in spore size and shape, over great distances and among many hosts, was remarkable.     

Molecular mapping of idiotopes of anti-arsonate antibodies

As part of understanding molecular function in structural terms, we have been attempting to map the idiotypic topography of specific anti-arsonate (Ars) antibodies. A panel of anti-Ars hybridomas of which the complete primary sequences are known were used. These molecules show a varied reactivity profile with a panel of monoclonal anti-idiotypic antibodies. By judicious chain recombination experiments and chemical modifications that altered this reactivity profile, we were able to identify particular amino acid residues or discreet regions of anti-Ars antibodies as having crucial roles in the expression of idiotypic determinants. Idiotopes were mapped to the heavy chain second hypervariable region and D segment, and to the light chain first and third hypervariable regions.     

N-Acetylation of glucosamine-6-phosphate in Leuconostoc mesenteroides

A partially purified enzyme (120-fold) from Leuconostoc mesenteroides catalyzed the reversible N-acetylation of d-glucosamine-6-phosphate. Coenzyme A was not required and inhibited the reaction rate. Neither d-glucosamine nor N-acetyl-d-glucosamine served as a substrate for the reversible reaction. The enzyme preparation retained 50% of its original activity after 5 min at 100 C. The K(m) for acetate was 7.7 x 10(-2)m in the presence of 2 x 10(-2)md-glucosamine-6-phosphate. The K(m) for d-glucosamine-6-phosphate was 5.0 x 10(-3)m in the presence of 0.64 m acetate. The product of the reaction was characterized by comparison with N-acetyl-d-glucosamine-6-phosphate prepared by enzymatic phosphorylation of N-acetyl-d-glusamine. The characterization tests were: chromatographic migration, acid hydrolysis, enzymatic dephosphorylation, sodium borohydride reduction, and periodate oxidation. The equilibrium constant for the reaction was about 7.5 m for the expression K = (d-glucosamine-6-phosphate)(acetate)/N-acetyl-d-glucosamine-6-phosphate. The standard free energy of the reaction was approximately 1,200 cal per mole.     

Oldest camarasauromorph sauropod (Dinosauria) discovered in the Middle Jurassic (Bajocian) of the Khadir Island, Kachchh, western India

Fragmentary isolated remains of large (up to 20 m or more) sauropods from the Middle Jurassic (Bajocian) Khadir Formation of Khadir Island (Kachchh, W India) are described and compared in detail. Three of the bone fragments (a metacarpal, a first pedal claw and a fibula) can be assigned with confidence to the Camarasauromorpha and represent the oldest known record of that derived dinosaur group. The new finds from western India further close a temporal and geographical gap in our knowledge of sauropods and contribute to understanding their early phylogeny.      

E-Combretastatin and E-resveratrol structural modifications: Antimicrobial and cancer cell growth inhibitory β-E-nitrostyrenes

As part of a broad-based SAR investigation of E-resveratrol (strong sirtuin activator and antineoplastic) and the anticancer vascular-targeting combretastatin-type stilbenes, a series of twenty-three β-E-nitrostyrenes was synthesized in order to evaluate potential antineoplastic, antitubulin, and antimicrobial activities. The β-E-nitrostyrenes evaluated ranged from monosubstituted phenols to trimethoxy and 3-methoxy-4,5-methylenedioxy derivatives. Two of the β-nitrostyrenes were synthesized as water-soluble sodium phosphate derivatives (4t, 4v). All except four (4r, 4s, 4t, 4u) of the series significantly inhibited a minipanel of human cancer cell lines. All but eight led to an IC₅₀ of <10 μM for inhibition of tubulin polymerization, and all except three (4l, 4t, 4v) displayed antimicrobial activity.