Nuclear protein transitions in cuttle-fish spermiogenesis: Immunocytochemical localization of a protein specific for the spermatid stage

The changes in basic nuclear proteins throughout cuttle-fish spermiogenesis were investigated both by immunocytochemical procedures and by isolation of late spermatid nuclei (by virtue of their resistance to sonication). Antibodies were raised in rabbits to a protein, named protein T, isolated from testis chromatin. The anti-protein T immune serum was found to recognize protein T and not histones from the testis. Immunoperoxidase staining of sections or of smears of testis with anti-protein T antibodies showed that protein T appears in the nuclei of round spermatids, is abundant in elongating spermatid nuclei, but cannot be detected in elongated spermatids. Nuclei from these elongated spermatids were isolated by sonication treatment of testis cells. A protein, named protein Sp, with the characteristic mobility of a protamine, was isolated from elongated spermatid nuclei. This protein has the same mobility as the protamine present in mature spermatozoa. Taken together, the results indicate that in cuttle-fish, nuclear protein transitions involve the replacement of histones by a spermatid-specific protein (protein T), which is replaced at the end of elongation of the nucleus by a protamine (protein Sp). Thus, spermiogenesis of the cuttle-fish (and perhaps of other cephalopods), shows two basic nuclear protein transitions, which are similar to the transitions observed in higher vertebrates such as mammals.     

Immobilization of microsomes into alginate beads is a convenient method for producing glucuronides from drugs

Summary The production of glucuronides from drugs by immobilized microsomal uridine diphosphate (UDP)-glucuronosyltransferase has been investigated. Of all the immobilization methods used (covalent binding, adsorption by ionic or hydrophobic interactions), only entrapment of microsomes into alginate beads in the presence of polyethyleneimine was effective in producing high glucuronidation rates, thus leading to the formation of large amounts of metabolites. The performance of the bioreactor was optimized with the drug 3-azido-3-deoxythymidine (AZT), active against the human immunodeficiency virus, as a model substrate of UDP-glucuronosyltransferase. Calcium (12 mm) could optimally improve the stability of microsomes entrapped in alginate beads. Upon immobilization, enzyme activation occurred, leading to a fivefold increase in specific activity. The determination of apparent K _m and V _max revealed that AZT was a better substrate for the immobilized enzyme than free microsomes. The AZT-glucuronide production obtained after 6 h was threefold higher than that observed with free microsomes. This bioreactor was also efficient in production of glucuronides from structurally different compounds such as bilirubin, 4-nitrophenol, clofibric acid, pirprofen, dextrorphan or morphine, the corresponding glucuronide of which possesses pharmacological or toxicological interest. Offprint requests to: J. Magdalou     

Lack of effect of metoclopramide on plasma aldosterone concentrations in young calves

In five 10-day-old Holstein X Friesian male calves, the intravenous injection of the dopamine blocker metoclopramide (1 mg/kg bwt) had no significant effect on plasma aldosterone concentration. Plasma sodium, potassium, cortisol, corticosterone concentrations and plasma renin activity measured in these animals during 120 min following metoclopramide injection were never significantly different from those simultaneously measured in 5 control calves.     

A thiol protease and an anionic peroxidase are induced by lowering cytokinins during callus growth in Petunia.

We previously identified a group of proteins that increase early in Petunia hybrida calli subcultured on a low-cytokinin medium, unlike the calli subcultured on a high-cytokinin medium. The calli on the low-cytokinin medium do not regenerate (J.-P. Renaudin, C. Tournaire, B, Teyssendier de la Serve [1991] Physiol Plant 82: 48-56). Two of these proteins, P21 and P17, have been identified by peptide sequencing and cloned. P21 is highly homologous to a group of thiol proteases, including barely aleurain, rice oryzain gamma, Arabidopsis SAG2, and mammalian cathepsin H. P17 is highly homologous to a group of anionic peroxidases from potato and tomato. A study of their expression in two P. hybrida lines, PC6 and St40 which differ in their ability to regenerate, showed that the genes for P21 and P17 are differentially expressed depending on the type and the age of the organ, with the highest expression in senescing leaves and in aged calli. The data are in favor of these genes being associated with an early step of senescence, which may be due, in part, to a reduction in total cytokinin. The two Petunia lines are, thus, functionally different concerning the action of cytokinin in two developmental phenomena: in vitro organogenesis and senescence.     

Local adaptation to temperature in populations and clonal lineages of the Irish potato famine pathogen Phytophthora infestans

Environmental factors such as temperature strongly impact microbial communities. In the current context of global warming, it is therefore crucial to understand the effects of these factors on human, animal, or plant pathogens. Here, we used a common‐garden experiment to analyze the thermal responses of three life‐history traits (latent period, lesion growth, spore number) in isolates of the potato late blight pathogen Phytophthora infestans from different climatic zones. We also used a fitness index (FI) aggregating these traits into a single parameter. The experiments revealed patterns of local adaptation to temperature for several traits and for the FI, both between populations and within clonal lineages. Local adaptation to temperature could result from selection for increased survival between epidemics, when isolates are exposed to more extreme climatic conditions than during epidemics. We also showed different thermal responses among two clonal lineages sympatric in western Europe, with lower performances of lineage 13_A2 compared to 6_A1, especially at low temperatures. These data therefore stress the importance of thermal adaptation in a widespread, invasive pathogen, where adaptation is usually considered almost exclusively with respect to host plants. This must now be taken into account to explain, and possibly predict, the global distribution of specific lineages and their epidemic potential.     

Unlocking biodiversity and conservation studies in high‐diversity environments using environmental DNA (eDNA): A test with Guianese freshwater fishes

Determining the species compositions of local assemblages is a prerequisite to understanding how anthropogenic disturbances affect biodiversity. However, biodiversity measurements often remain incomplete due to the limited efficiency of sampling methods. This is particularly true in freshwater tropical environments that host rich fish assemblages, for which assessments are uncertain and often rely on destructive methods. Developing an efficient and nondestructive method to assess biodiversity in tropical freshwaters is highly important. In this study, we tested the efficiency of environmental DNA (eDNA) metabarcoding to assess the fish diversity of 39 Guianese sites. We compared the diversity and composition of assemblages obtained using traditional and metabarcoding methods. More than 7,000 individual fish belonging to 203 Guianese fish species were collected by traditional sampling methods, and ~17 million reads were produced by metabarcoding, among which ~8 million reads were assigned to 148 fish taxonomic units, including 132 fish species. The two methods detected a similar number of species at each site, but the species identities partially matched. The assemblage compositions from the different drainage basins were better discriminated using metabarcoding, revealing that while traditional methods provide a more complete but spatially limited inventory of fish assemblages, metabarcoding provides a more partial but spatially extensive inventory. eDNA metabarcoding can therefore be used for rapid and large‐scale biodiversity assessments, while at a local scale, the two approaches are complementary and enable an understanding of realistic fish biodiversity.     

The endogenous oxindoles 5-hydroxyoxindole and isatin are antiproliferative and proapoptotic

Oxindole-core synthetic molecules are currently being developed as anticancer drugs that target protein tyrosine kinases associated with growth factor receptors. Oxindole, 5-Hydroxyoxindole, and 2, 3-dioxindole [isatin] are natural molecules found in mammalian body fluids and tissues and we addressed the question of similar properties of endogenous oxindoles. 5-Hydroxyoxindole and isatin, but not oxindole, inhibited N1E-115, BALB/c3T3, BBC, PC12, and HL60 proliferation at submicromolar concentrations. Acute treatment with 5-hydroxyoxindole and isatin reduced the activity of extracellular signal regulated protein kinases (ERKs) by 35% at 100 microM and ERK1 activity was strongly inhibited by 5-Hydroxyoxindole at 10 microM. Survival of PMA-differentiated HL60 and FGF(2)-differentiated PC12 cells was not affected by 5-Hydroxyoxindole and isatin treatment, suggesting that endogenous oxindoles interact with growth factors signaling. The physiological implications of these data and the potential utility of 5-Hydroxyoxindole and isatin as antitumor agents are discussed.     

Reduced secretion and expression of gelatinase profile in Toxoplasma gondii-infected human monocytic cells

The apicomplexan Toxoplasma gondii, an obligate intracellular parasite, can infect humans and a wide range of vertebrates. Following oral infection, the parasite invades tissues by crossing non-permissive biological barriers such as the placenta or the blood-brain barrier. But the molecular mechanisms underlying migration of T. gondii remain poorly characterized. The crossing of various basal membranes and infiltration into the extracellular matrix by T. gondii could involve matrix metalloproteinases (MMPs). We demonstrated a decrease in proMMP-2 and proMMP-9 secretion by THP-1 cells at 24 and 48h post invasion with regulation at the mRNA level throughout infection. This down regulation was associated with a decrease in TIMP-2 secretion and an inhibition of its expression. Moreover, results showed an activation of MT1-MMP; its expression was regulated after 6, 24, and 48h.