Characterisation of a Desmosterol Reductase Involved in Phytosterol Dealkylation in the Silkworm, Bombyx mori

Most species of invertebrate animals cannot synthesise sterols de novo and many that feed on plants dealkylate phytosterols (mostly C29 and C28) yielding cholesterol (C27). The final step of this dealkylation pathway involves desmosterol reductase (DHCR24)-catalysed reduction of desmosterol to cholesterol. We now report the molecular characterisation in the silkworm, Bombyx mori, of such a desmosterol reductase involved in production of cholesterol from phytosterol, rather than in de novo synthesis of cholesterol. Phylogenomic analysis of putative desmosterol reductases revealed the occurrence of various clades that allowed for the identification of a strong reductase candidate gene in Bombyx mori (BGIBMGA 005735). Following PCR-based cloning of the cDNA (1.6 kb) and its heterologous expression in Saccharomyces cerevisae, the recombinant protein catalysed reduction of desmosterol to cholesterol in an NADH- and FAD- dependent reaction

Preferenze alimentari in Bombyx mori: saggi comportamentali

Le larve del baco da seta (Bombyx mori L.) sono strettamente monofaghe, infeudate al gelso (Morus spp.) e legate alla presenza di foglia fresca. I dati presentati riguardano lo studio dell’attrattività da parte di alcuni ceppi del baco da seta (due cinesi, due giapponesi e due poliibridi) nei confronti di diete sostitutive, preparate con diverse percentuali di polvere di foglia di gelso essiccata (da 0% a 40%). Lo scopo è quello di determinare il ceppo/i con le migliori “prestazioni” verso la dieta contenente la minor percentuale di gelso compatibile con lo sviluppo dell’organismo. L’indagine è eseguita su larve in I età in condizioni chimico-fisiche controllate. I valori più alti di affluenza al cibo sono stati raggiunti dai ceppi giapponesi, indipendentemente dalla percentuale di gelso; per gli altri ceppi l’attrattività aumenta proporzionalmente alla quantità di gelso presente. Per meglio comprendere i fattori che possono essere responsabili di questo comportamento, su tre diete (0%, 25% e 40%) è stata eseguita, per ogni ceppo, un’analisi delle traiettorie percorse dalle larve per il raggiungimento della dieta. Lo sviluppo cinematico e geometrico della locomozione dell’organismo sono studiati tramite i seguenti indici: l’indice geometrico, che esprime il rapporto tra la distanza dal punto iniziale al punto finale del percorso (D) e la lunghezza (L) effettiva del tragitto (Ig = D/L), descrivendo la geometria complessiva della traettoria seguita; l’indice cinetico (Ik= L/t), che esprime la velocità media della larva; il tasso di dislocazione (Rd = Ik . Ig = D/t), che esprime il tasso di spostamento teorico dell’organismo indipendentemente dalla geometria della traettoria e dalla velocità effettiva posseduta. I risultati ottenuti indicano che, per quanto concerne l’efficienza geometrica del percorso, non esistono differenze significative tra i vari ceppi. Per quanto riguarda l’indice cinetico sono riscontrate delle differenze significative: in particolare, il ceppo cinese 121 risulta più veloce quanto più è bassa la percentuale di gelso, mentre il ceppo giapponese 71 è tanto più veloce quanto maggiore è la quantità di gelso nella dieta. Gli altri ceppi mostrano un comportamento intermedio. In conclusione, si può affermare che il sistema di analisi applicato in questo studio è in grado di fornire indicazioni utili per la scelta del ceppo e della percentuale di gelso idonei per l’allevamento del baco da seta con dieta sostitutiva

Modification of the nutritional parameters and of midgut biochemical and absorptive functions induced by the IGR fenoxycarb in Bombyx mori larvae

Fifth instar larvae of B. mori were topically or orally treated with increasing amounts of the Insect Growth Regulator (IGR) fenoxycarb in a single application, in order to determine its effects on the nutritional parameters, the midgut functional activities and the growth of the silk glands. The IGR affected in a dose-dependent manner the progress of the life cycle of the insect, causing a delay or inhibition of spinning, alteration of the feeding behaviour, decrease of the nutritional parameters, impairment of the growth of the silk glands, and an increased mortality during larval-pupal transformation. Measurement of leucine uptake into midgut brush border membrane vesicles and midgut histochemistry revealed a reduced absorption of leucine by the midgut and a large alteration of a number of midgut enzyme activities as a result of treatments with a high dose of fenoxycarb (2.5 micro g). Treatments with a dose of 2.5 femto g/larva caused an increase in leucine uptake by the midgut, an increased weight of the cocoon shell, and a modification of some midgut enzyme activities. The lepidopteran midgut appears to be a larval organ that responds promptly to the exposure to fenoxycarb. The epithelial columnar cells modify their absorptive functions, at least with regard to amino acid uptake, as well as their metabolic activity, with a modification of the oxidative status of the cells that is detectable with a single dose of the chemical as low as few fg/larva

Post-translational control of human hemoglobin synthesis The role of the differential affinity between globin chains in the control of mutated globin gene expression

The interactions between β-thalassemia and the humen hemoglobin (Hb) α-chain variants, Hb Hasharon, Hb O Indonesia and Hb J Paris, and between α-thalassemia and the β-chain variants, Hb S, Hb C and Hb G San José, which are characterized by preferential decrease of the abnormal Hb level in peripheral blood, have been studied. Both biosynthesis studies in reticulocytes and determination of the relative affinity of abnormal chains for normal complementary chains by in vivo recombination experiments, involving globin chains previously isolated in their native form, have been carried out in order to provide insights on the molecular events following the synthesis of the mutant chains under conditions of complementary chain deficiency. Furthermore, we have measured the relative affinity for complementary chain of βD Los Angeles- and αJ Rovigo-chains, the level of which does not decay in thalassemic carriers, and of αLegnano- and βOsu Christiansborg-chains, which have not yet been observed in association with thalassemias. Our experiments indicated that the differential affinity for β-chains is not always the major post-translational control mechanism which regulates the level of certain α-chain variants in β-thalassemic heterozygotes, and that preferential removal of abnormal chains by proteolytic enzymes is likely to play an important role in most cases. On the other hand, the low affinity of certain variant β-chains for α-chains may offer an explanation for the low level of certain β-chain variants in peripheral blood of non-thalassemic carriers, as well as to their decrease under conditions of relative α-chain deficiency (α-thalassemias). © 1980

Modulation of leucine absorption in the larval midgut of Bombyx mori (Lepidoptera, Bombycidae)

In the larval midgut of Bombyx mori a K+-dependent transporter for leucine and amino acids with a hydrophobic side chain is responsible for the absorption of most essential amino acids. We investigated if a modulation of its activity occurred as a result of starvation or after hormonal treatments. We measured amino acid uptake in brush border membrane vesicles (BBMV) purified from the anterior-middle (AM) and posterior (P) regions of the midgut in fifth instar larvae. Silkworms were either starved or topically treated with low dosages of fenoxycarb, a molecule often used as a juvenile hormone mimic. The maximal uptake value of K+-driven leucine transport was increased in BBMV of AM- and P-midgut regions of starved larvae. The initial uptake rates of serine and glutamine, two amino acids transported by the same cotransporter as leucine, were also increased. Leucine kinetics proved that Vmax was the kinetic parameter modified by starvation in both midgut regions. Topical applications of fenoxycarb at a dose of 2.5 fg/larva immediately after the fourth ecdysis, induced an increase of leucine initial uptake rates and of intravesicular accumulation of leucine in both AM- and P-BBMV. Kinetic analysis of leucine uptake indicated again that Vmax was increased in BBMV from both midgut regions in treated larvae