Host Cell Poly(ADP-Ribose) Glycohydrolase Is Crucial for Trypanosoma cruzi Infection Cycle

Trypanosoma cruzi, etiological agent of Chagas’ disease, has a complex life cycle which involves the invasion of mammalian host cells, differentiation and intracellular replication. Here we report the first insights into the biological role of a poly(ADP-ribose) glycohydrolase in a trypanosomatid (TcPARG). In silico analysis of the TcPARG gene pointed out the conservation of key residues involved in the catalytic process and, by Western blot, we demonstrated that it is expressed in a life stage-dependant manner. Indirect immunofluorescence assays and electron microscopy using an anti-TcPARG antibody showed that this enzyme is localized in the nucleus independently of the presence of DNA damage or cell cycle stage. The addition of poly(ADP-ribose) glycohydrolase inhibitors ADP-HPD (adenosine diphosphate (hydroxymethyl) pyrrolidinediol) or DEA (6,9-diamino-2-ethoxyacridine lactate monohydrate) to the culture media, both at a 1 µM concentration, reduced in vitro epimastigote growth by 35% and 37% respectively, when compared to control cultures. We also showed that ADP-HPD 1 µM can lead to an alteration in the progression of the cell cycle in hydroxyurea synchronized cultures of T. cruzi epimastigotes. Outstandingly, here we demonstrate that the lack of poly(ADP-ribose) glycohydrolase activity in Vero and A549 host cells, achieved by chemical inhibition or iRNA, produces the reduction of the percentage of infected cells as well as the number of amastigotes per cell and trypomastigotes released, leading to a nearly complete abrogation of the infection process. We conclude that both, T. cruzi and the host, poly(ADP-ribose) glycohydrolase activities are important players in the life cycle of Trypanosoma cruzi, emerging as a promising therapeutic target for the treatment of Chagas’ disease.     

Malaria Parasite-Infected Erythrocytes Secrete PfCK1, the Plasmodium Homologue of the Pleiotropic Protein Kinase Casein Kinase 1

Casein kinase 1 (CK1) is a pleiotropic protein kinase implicated in several fundamental processes of eukaryotic cell biology. Plasmodium falciparum encodes a single CK1 isoform, PfCK1, that is expressed at all stages of the parasite’s life cycle. We have previously shown that the pfck1 gene cannot be disrupted, but that the locus can be modified if no loss-of-function is incurred, suggesting an important role for this kinase in intra-erythrocytic asexual proliferation. Here, we report on the use of parasite lines expressing GFP- or His-tagged PfCK1 from the endogenous locus to investigate (i) the dynamics of PfCK1 localisation during the asexual cycle in red blood cells, and (ii) potential interactors of PfCK1, so as to gain insight into the involvement of the enzyme in specific cellular processes. Immunofluorescence analysis reveals a dynamic localisation of PfCK1, with evidence for a pool of the enzyme being directed to the membrane of the host erythrocyte in the early stages of infection, followed by a predominantly intra-parasite localisation in trophozoites and schizonts and association with micronemes in merozoites. Furthermore, we present strong evidence that a pool of enzymatically active PfCK1 is secreted into the culture supernatant, demonstrating that PfCK1 is an ectokinase. Our interactome experiments and ensuing kinase assays using recombinant PfCK1 to phosphorylate putative interactors in vitro suggest an involvement of PfCK1 in many cellular processes such as mRNA splicing, protein trafficking, ribosomal, and host cell invasion.     

How costly is clutch formation in the Audouin's Gull Larus audouinii?

During the Audouin's Gull's breeding season at the Ebro Delta in 1993, 24 fresh eggs from eight three-egg clutches (modal clutch-size) were collected at the peak of the laying period. Eggs were processed to obtain formalin-fixed yolks, which were halved and stained using the potassium dichromate method. Digitized images of the yolks were examined to assess the daily rates of yolk deposition. We used these data in combination with egg compositional analysis to build a model of energy demands during the formation of an average clutch in Audouin's Gull. To show how the different parameters of clutch formation affect the daily energy investment peak, we performed a simulation analysis in which the rapid yolk development (RYD) period, the follicle triggering interval (FTI), the laying interval (LI) and the albumen synthesis period (ASP) were allowed to vary simultaneously. In our sample, the mean RYD period was seven days with a range from six to eight days. There were no significant differences in yolk volume among eggs in a clutch, but albumen volume was significantly smaller in third eggs. According to our model the albumen synthesis of the a-egg coincides with the energy demand peak for clutch formation. This peak represents an increase by ca. 42% in female energy requirements. Values obtained from the simulation analysis showed that only the ASP of the a-egg and the RYD durations of the second and third follicles produced noticeable reductions in peak energy investment. We predict that in gulls, whose laying intervals seem to be kept constant, significant increases of the durations of the RYD periods of second and third eggs, or even significant reductions of yolk size of these eggs, may operate simultaneously to match the energy demands during clutch formation to the prevailing food conditions.     

Turnover of plant lineages shapes herbivore phylogenetic beta diversity along ecological gradients

Understanding drivers of biodiversity patterns is of prime importance in this era of severe environmental crisis. More diverse plant communities have been postulated to represent a larger functional trait‐space, more likely to sustain a diverse assembly of herbivore species. Here, we expand this hypothesis to integrate environmental, functional and phylogenetic variation of plant communities as factors explaining the diversity of lepidopteran assemblages along elevation gradients in the Swiss Western Alps. According to expectations, we found that the association between butterflies and their host plants is highly phylogenetically structured. Multiple regression analyses showed the combined effect of climate, functional traits and phylogenetic diversity in structuring butterfly communities. Furthermore, we provide the first evidence that plant phylogenetic beta diversity is the major driver explaining butterfly phylogenetic beta diversity. Along ecological gradients, the bottom up control of herbivore diversity is thus driven by phylogenetically structured turnover of plant traits as well as environmental variables.     

Alterations of Choroidal Blood Flow Regulation in Young Healthy Subjects with Complement Factor H Polymorphism

A common polymorphism in the complement factor H gene (rs1061170, Y402H) is associated with a high risk of age-related macular degeneration (AMD). In the present study we hypothesized that healthy young subjects homozygous for the high-risk haplotype (CC) show abnormal choroidal blood flow (ChBF) regulation decades before potentially developing the disease. A total of 100 healthy young subjects were included in the present study, of which 4 subjects were excluded due to problems with genotyping or blood flow measurements. ChBF was measured continuously using laser Doppler flowmetry while the subjects performed isometric exercise (squatting) for 6 minutes. The increase in ChBF was less pronounced than the response in ocular perfusion pressure (OPP), indicating for some degree of choroidal blood flow regulation. Eighteen subjects were homozygous for C, 47 subjects were homozygous for T and 31 subjects were heterozygous (CT). The increase in OPP during isometric exercise was not different between groups. By contrast the increase in ChBF was more pronounced in subjects homozygous for the high risk C allele (p = 0.041). This was also evident from the pressure/flow relationship, where the increase in ChBF in homozygous C carriers started at lower OPPs as compared to the other groups. Our data indicate that the regulation of ChBF is abnormal in rs1061170 CC carriers. So far this polymorphism has been linked to age related macular degeneration (AMD) mainly via inflammatory pathways associated with the complement system dysfunction. Our results indicate that it could also be related to vascular factors that have been implicated in AMD pathogenesis.