Differential distribution and metabolism of arachidonic acid and docosahexaenoic acid by human placental choriocarcinoma (BeWo) cells

The time course of incorporation of [¹⁴C]arachidonic acid and [³H]docosahexaenoic acid into various lipid fractions in placental choriocarcinoma (BeWo) cells was investigated. BeWo cells were found to rapidly incorporate exogenous [¹⁴C]arachidonic acid and [³H] docosahexaenoic acid into the total cellular lipid pool. The extent of docosahexaenoic acid esterification was more rapid than for arachidonic acid, although this difference abated with time to leave only a small percentage of the fatty acids in their unesterified form. Furthermore, uptake was found to be saturable. In the cellular lipids these fatty acids were mainly esterified into the phospholipid (PL) and the triacyglycerol (TAG) fractions. Smaller amounts were also detected in the diacylglycerol and cholesterol ester fractions. Almost 60% of the total amount of [3H]Docosahexaenoic acid taken up by the cells was esterified into TAG whereas 37% was in PL fractions. For arachidonic acid the reverse was true, 60% of the total uptake was incorporated into PL fractions whereas less than 35% was in TAG. Marked differences were also found in the distribution of the fatty acids into individual phospholipid classes. The higher incorporation of docosahexaenoic acid and arachidonic acid was found in PC and PE, respectively. The greater cellular uptake of docosahexaenoic acid and its preferential incorporation in TAG suggests that both uptake and transport modes of this fatty acid by the placenta to fetus is different from that of arachidonic acid.     

The activities of pyruvate carboxylase, phosphoenolpyruvate carboxylase and fructose diphosphatase in muscles from vertebrates and invertebrates

1. The activities of pyruvate carboxylase, phosphoenolpyruvate carboxylase and fructose diphosphatase in crude homogenates of vertebrate and invertebrate muscles are reported. 2. Pyruvate carboxylase activity was present in all insect flight muscles that were investigated: in homogenates of bumble-bee flight muscle the activity was inhibited by ADP and activated by acetyl-CoA, and it was distributed mainly in the mitochondrial fraction. This is the first demonstration of pyruvate carboxylase activity in muscle. However, the activity appears to be restricted to insect flight muscle, since it was not found in other invertebrate or vertebrate muscles. 3. Since the three enzymes were never found together in the same muscle, it is concluded that these enzymes cannot provide a pathway for the synthesis of glycogen from lactate or pyruvate in muscle. Other roles for these enzymes in muscle are suggested. In particular, pyruvate carboxylase may be present in insect flight muscle for the provision of oxaloacetate to support the large increase in activity of the tricarboxylic acid cycle which occurs when an insect takes flight.     

Insights into Coupled Folding and Binding Mechanisms from Kinetic Studies

Intrinsically disordered proteins (IDPs) are characterized by a lack of persistent structure. Since their identification more than a decade ago, many questions regarding their functional relevance and interaction mechanisms remain unanswered. Although most experiments have taken equilibrium and structural perspectives, fewer studies have investigated the kinetics of their interactions. Here we review and highlight the type of information that can be gained from kinetic studies. In particular, we show how kinetic studies of coupled folding and binding reactions, an important class of signaling event, are needed to determine mechanisms.     

Deep placement of Mn fertiliser on a sandy soil increased grain yield and reduced split seed in Lupinus angustifolius

Experiments were conducted over two years with Lupinus angustifolius L. on a site with acid sandy soil near Esperance, Western Australia to determine if deep placed manganese fertilizer increases lupin grain yield. Manganese at 4 and 8 kg ha⁻¹ was placed below the surface immediately before sowing at 4, 20 and 30 cm and 4, 8, 12, 16 and 20 cm in 1987 and 1988 respectively. Foliar Mn applied at 1 kg ha⁻¹ when the first order laterals were in mid-flowering stage, was also compared. Increasing the depth of Mn placement increased grain yield in both years. The deepest placed Mn increased grain yields by 255 kg ha⁻¹ (10%) and 430 kg ha⁻¹ (106%) in year 1 and year 2 over the shallow (4 cm) placed Mn. The higher responses to deep placed Mn occurred in year 2, the year with the driest spring and most intense aphid infestations. Foliar applied Mn was as effective as most deep placed Mn treatments, except for the highest rate (8 kg ha⁻¹) at the greatest depth (20 cm) in year 2. The higher rate of applied Mn gave the best grain yields. This revised version was published online in June 2006 with corrections to the Cover Date.     

Recruitment of TBK1 to cytosol‐invading Salmonella induces WIPI2‐dependent antibacterial autophagy

Mammalian cells deploy autophagy to defend their cytosol against bacterial invaders. Anti‐bacterial autophagy relies on the core autophagy machinery, cargo receptors, and “eat‐me” signals such as galectin‐8 and ubiquitin that label bacteria as autophagy cargo. Anti‐bacterial autophagy also requires the kinase TBK1, whose role in autophagy has remained enigmatic. Here we show that recruitment of WIPI2, itself essential for anti‐bacterial autophagy, is dependent on the localization of catalytically active TBK1 to the vicinity of cytosolic bacteria. Experimental manipulation of TBK1 recruitment revealed that engagement of TBK1 with any of a variety of Salmonella‐associated “eat‐me” signals, including host‐derived glycans and K48‐ and K63‐linked ubiquitin chains, suffices to restrict bacterial proliferation. Promiscuity in recruiting TBK1 via independent signals may buffer TBK1 functionality from potential bacterial antagonism and thus be of evolutionary advantage to the host.