RAB-6.1 and RAB-6.2 Promote Retrograde Transport in C. elegans

Retrograde transport is a critical mechanism for recycling certain membrane cargo. Following endocytosis from the plasma membrane, retrograde cargo is moved from early endosomes to Golgi followed by transport (recycling) back to the plasma membrane. The complete molecular and cellular mechanisms of retrograde transport remain unclear. The small GTPase RAB-6.2 mediates the retrograde recycling of the AMPA-type glutamate receptor (AMPAR) subunit GLR-1 in C. elegans neurons. Here we show that RAB-6.2 and a close paralog, RAB-6.1, together regulate retrograde transport in both neurons and non-neuronal tissue. Mutants for rab-6.1 or rab-6.2 fail to recycle GLR-1 receptors, resulting in GLR-1 turnover and behavioral defects indicative of diminished GLR-1 function. Loss of both rab-6.1 and rab-6.2 results in an additive effect on GLR-1 retrograde recycling, indicating that these two C. elegans Rab6 isoforms have overlapping functions. MIG-14 (Wntless) protein, which undergoes retrograde recycling, undergoes a similar degradation in intestinal epithelia in both rab-6.1 and rab-6.2 mutants, suggesting a broader role for these proteins in retrograde transport. Surprisingly, MIG-14 is localized to separate, spatially segregated endosomal compartments in rab-6.1 mutants compared to rab-6.2 mutants. Our results indicate that RAB-6.1 and RAB-6.2 have partially redundant functions in overall retrograde transport, but also have their own unique cellular- and subcellular functions.     

Development of sporeless/low sporing strains of Pleurotus through mutation

Summary The sporophores of Pleurotus are gymnocarpous and continuously release spores in the atmosphere causing respiratory allergies like hay fever and farmer’s lung disease among workers. The allergy is caused by the antigens present on the walls of the spores. Apart from this, during commercial production, these spores settle on the fruit bodies, germinate and form a velvety film which gives an unpleasant appearance to the mushrooms. The spores emitted may include new genotypes likely to attack wood or trees. Spore allergy is one of the most important limiting factors for the large scale cultivation of this species. Different approaches are being adopted at IIHR for the production of commercial sporeless/low-sporing strains of Pleurotus to alleviate the spore allergy problem. Attempts were made during the present investigation to produce sporeless or low-sporing mutants through u.v. mutation. Mutation of the mycelium did not yield the desired results. Mutation of the spores of Pleurotus sajor-caju yielded an extremely low-sporing mutant after 75 min exposure. The character has been found to be stable for more than 10 generations of subculturing.     

Effect of ganglionic stimulating and blocking agents on the fast components of colonic myoelectrical activity in the rat

The presence of two types of fast myoelectrical activities, medium fast activity and fast activity, has been demonstrated previously in the electromyogram of colon in normal children and in the rat by the authors. An absence of medium fast activity in Hirschsprung's disease and in experimental aganglionosis of colon in the rat has also been described. In the present study the fast components of colonic myoelectrical activity were analysed during the procedures affecting ganglionic transmission. It was observed that ganglionic stimulants, such as balloon inflation, and intra-arterial injections of acetylcholine and small amounts of nicotine, increased the spike activity and the frequency of medium fast activity without affecting fast activity. The intra-arterial injections of ganglionic blocking agents, such as nicotine in large amounts and pentolinium tartrate, completely abolished the medium fast activity. These observations suggest that the ganglionic activity is responsible for the genesis of medium fast activity and that the absence of cholinergic ganglionic transmission is the most important single factor for the reported altered electromyogram pattern in aganglionosis.     

TGF-Beta inhibits the in vitro induction of lymphokine-activated killing activity

Summary Employing serum-free media, human peripheral blood mononuclear cells, and purified recombinant interleukin-2 (IL-2), conditions were observed in which the development of IL-2-driven cytotoxic activity was suppressed. The cytotoxic activity of such IL-2-generated lymphokine activated killing (LAK) was tested against natural killer-resistant cultured tumor cells (Daudi, Raji, and a glioma). LAK generation was inhibited by addition of some normal sera, normal platelets, or some tumor cells. Because recent reports have indicated that transforming growth factor-beta (TGF-beta)-like factors are often secreted by tumors and the acidic alpha granules of platelets and can be present in sera, we tested the effect of purified human TGF-beta on the activation of LAK. Our results indicated that TGF-beta is very suppressive for LAK induction, and can completely prevent both the IL-2-driven proliferation and cytotoxicity at concentrations as low as 5 ng/ml. Titrations of IL-2 and of TGF-beta indicated that the suppression is dose-dependent and can be avoided by employing higher levels of IL-2. It was also found that the suppressive effect of TGF-beta can be overcome by washing suppressed cell populations and further culture in low levels of IL-2. Collectively, these data indicate that TGF-beta can be a potent inhibitor of LAK generation under standard activation conditions, but that this effect is regulated by the relative level of IL-2 and may be overcome and/or reversed in vitro.     

Experimental study of physiological pulsatile flow past valve prostheses in a model of human aorta--I. Caged ball valves

Pulsatile flow development past a caged ball valve in a model human aorta was studied using laser Doppler anemometry. Velocity profiles measured in the ascending aorta and in the mid-arch region were strongly influenced by the geometry of the valve at the root of the aorta. Velocity profiles distal to the valve were asymmetric with jet-like flow in the peripheral region having larger velocity magnitudes towards the left lateral wall. In early diastole, a streamwise vortex motion was observed throughout the model aorta with fluid moving towards the downstream direction along the left lateral wall and reversed flow along the right lateral wall. With the caged ball valve at the root of the aorta, no reversed flow was observed along the inner wall of curvature in the mid-arch region.     

Purification and regulation of glutamine synthetase in a collagenolytic Vibrio alginolyticus strain

Glutamine synthetase (EC 6.3.1.2) has been purified from a collagenolytic Vibrio alginolyticus strain. The apparent molecular weight of the glutamine synthetase subunit was approximately 62,000. This indicates a particle weight for the undissociated enzyme of 744,000, assuming the enzyme is the typical dodecamer. The glutamine synthetase enzyme had a sedimentation coefficient of 25.9 S and seems to be regulated by a denylylation and deadenylylation. The pH profiles assayed by the -glutamyltransferase method were similar for NH₄-shocked and unshocked cell extracts and isoactivity point was not obtained from these eurves. The optimum pH for purified and crude cell extracts was 7.9. Cell-free glutamine synthetase was inhibited by some amino acids and AMP. The transferase activity of glutamine synthetase from mid-exponential phase cells varied greatly depending on the sources of nitrogen or carbon in the growth medium. Glutamine synthetase level was regulated by nitrogen catabolite repression by (NH₄)₂SO₄ and glutamine, but cells grown, in the presence of proline, leucine, isoleucine, tryptophan, histidine, glutamic acid, glycine and arginine had enhanced levels of transferase activity. Glutamine synthetase was not subject to glucose, sucrose, fructose, glycerol or maltose catabolite repression and these sugars had the opposite effect and markedly enhanced glutamine synthetase activity.Abbreviations GS glutamine synthetase - SMM succinate minimal medium - ASMM ammonium/succinate minimal medium - GT -glutamyl transferase - SVP snake venom phosphodiesterase     

Steady flow development past valve prostheses in a model human aorta—I. Centrally occluding valves

In this paper, laser-Doppler anemometry measurement of steady flow development in a model human aorta has been reported. Studies were made with uniform entry flow at the root of the aorta and our measurements showed the establishment of a pair of Dean vortices in the mid-arch region. Subsequently, the nature of flow development past centrally occluding caged ball valves in the model aorta was investigated. Our studies showed that in the ascending aorta, an asymmetric velocity profile is obtained with larger velocity gradients towards the inner wall of tertiary curvature (anatomically the left lateral wall) with centrally occluding valves. The peripheral flow past these valves prevented the development of Dean vortices in the mid-arch region. The caged ball valves at the root of the aorta had no discernible effect on the velocity profiles in the brachio-cephalic artery.     

Effect of systolic flow rate on the prediction of effective prosthetic valve orifice area

The Gorlin equation for the hemodynamic assessment of valve area is commonly used in cardiac catheterization laboratories. A study was performed to test the prediction capabilities of the Gorlin formula as well as the Aaslid and Gabbay formula for the effective orifice area of prosthetic heart valves. Pressure gradient, flow, and valve opening area measurements were performed on four 27 mm valve prostheses (two mechanical bileaflet designs, St. Jude and Edwards-Duromedics, an Edwards pericardial tissue valve, and a trileaflet polyurethane valve) each mounted in the aortic position of an in vitro pulse duplicator. With the known valve orifice area, a different discharge coefficient was computed for each of the four valves and three orifice area formulas. After some theoretical considerations, it was proposed that the discharge coefficient would be a function of the flow rate through the valve. All discharge coefficients were observed to increase with increasing systolic flow rate. An empirical relationship of discharge coefficient as a linear function of systolic flow rate was determined through a regression analysis, with a different relationship for each valve and each orifice area formula. Using this relationship in the orifice area formulas improved the accuracy of the prediction of the effective orifice area with all three formulas performing equally well.     

Structure of the mouth tube of a parasitic copepod,Isobranchia appendiculata Heegaard, 1947 (Siphonostomatoida:Lernaeopodidae)

The structure and musculature of the mouth tube of the lernaeopodidIsobranchia appendiculata Heegaard is described in detail. Situated on the median eminence, the mouth tube appears to be lodged inside a median longitudinal groove on the ventral wall of the cephalothorax. The components of the mouth tube, the labrum and the labium are loosely held together along their lateral edges. The thin, transparent walls of both the labrum and the labium are reinforced by chitinous rods. The musculature of the mouth tube consists of two pairs ofretractores oris, four pairs ofcompressores labri and two pairs oflevatores labii. The mandibles are operated by a pair of antagonistic muscles, theroto adductor mandibulae androto abductor mandibulae. Like other lernaeopodids,I. appendiculata resorts to a browsing type of feeding.