Hydrazino-aza and N-azapeptoids with therapeutic potential as anticancer agents

The ubiquitin-proteasome-mediated degradation pathway plays an important role in regulating protein turnover in eucaryotic cells and, consequently, regulates both cell proliferation and cell death. The proteasome influences many cellular regulatory signals and is thus a potential target for pharmacological agents. The study of proteasome function has led to the identification of several natural and synthetic compounds that can act as tumor cell growth inhibitors. In this study, we have developed a series of hydrazino-aza and N-azapeptoids, analogues of Ac-Leucyl-Leucyl-Norleucinal (ALLN) a non-specific peptidyl aldehyde inhibitor of the proteasome. These peptide analogues share a common backbone and bear different C- and N-terminal functions. Their antiproliferative activity on murine leukemia L1210 cells is reported here.     

The Plasmodium sporozoite journey: a rite of passage

Sporozoites are the most versatile of the invasive stages of the Plasmodium life cycle. During their passage within the mosquito vector and the vertebrate host, sporozoites display diverse behaviors, including gliding locomotion and invasion of, migration through and egress from target cells. At the end of the journey, sporozoites invade hepatocytes and transform into exoerythrocytic stages, marking the transition from the pre-erythrocytic to the erythrocytic part of the life cycle. This article discusses recent work, mostly done with rodent malaria parasites, that has contributed to a better understanding of the sporozoites' complex biology and which has opened up new avenues for future sporozoite research.     

Pharmaceutical biotechnology, Chemical biotechnology: Web alert

A selection of World Wide Web sites relevant to papers published in this issue of Current Opinion in Biotechnology.     

The schizophrenia susceptibility factor dysbindin and its associated complex sort cargoes from cell bodies to the synapse

Dysbindin assembles into the biogenesis of lysosome-related organelles complex 1 (BLOC-1), which interacts with the adaptor protein complex 3 (AP-3), mediating a common endosome-trafficking route. Deficiencies in AP-3 and BLOC-1 affect synaptic vesicle composition. However, whether AP-3-BLOC-1-dependent sorting events that control synapse membrane protein content take place in cell bodies upstream of nerve terminals remains unknown. We tested this hypothesis by analyzing the targeting of phosphatidylinositol-4-kinase type II α (PI4KIIα), a membrane protein present in presynaptic and postsynaptic compartments. PI4KIIα copurified with BLOC-1 and AP-3 in neuronal cells. These interactions translated into a decreased PI4KIIα content in the dentate gyrus of dysbindin-null BLOC-1 deficiency and AP-3-null mice. Reduction of PI4KIIα in the dentate reflects a failure to traffic from the cell body. PI4KIIα was targeted to processes in wild-type primary cultured cortical neurons and PC12 cells but failed to reach neurites in cells lacking either AP-3 or BLOC-1. Similarly, disruption of an AP-3-sorting motif in PI4KIIα impaired its sorting into processes of PC12 and primary cultured cortical neuronal cells. Our findings indicate a novel vesicle transport mechanism requiring BLOC-1 and AP-3 complexes for cargo sorting from neuronal cell bodies to neurites and nerve terminals.     

Unusual anchor of a motor complex (MyoD-MLC2) to the plasma membrane of Toxoplasma gondii

Toxoplasma gondii possesses 11 rather atypical myosin heavy chains. The only myosin light chain described to date is MLC1, associated with myosin A, and contributing to gliding motility. In this study, we examined the repertoire of calmodulin-like proteins in Apicomplexans, identified six putative myosin light chains and determined their subcellular localization in T. gondii and Plasmodium falciparum. MLC2, only found in coccidians, is associated with myosin D via its calmodulin (CaM)-like domain and anchored to the plasma membrane of T. gondii via its N-terminal extension. Molecular modeling suggests that the MyoD-MLC2 complex is more compact than the reported structure of Plasmodium MyoA-myosin A tail-interacting protein (MTIP) complex. Anchorage of this MLC2 to the plasma membrane is likely governed by palmitoylation.     

Structural Chromosome Abnormalities Associated with Obesity: Report of Four New subjects and Review of Literature

Obesity in humans is a complex polygenic trait with high inter-individual heritability estimated at 40-70%. Candidate gene, DNA linkage and genome-wide association studies (GWAS) have allowed for the identification of a large set of genes and genomic regions associated with obesity. Structural chromosome abnormalities usually result in congenital anomalies, growth retardation and developmental delay. Occasionally, they are associated with hyperphagia and obesity rather than growth delay. We report four new individuals with structural chromosome abnormalities involving 10q22.3-23.2, 16p11.2 and Xq27.1-q28 chromosomal regions with early childhood obesity and developmental delay. We also searched and summarized the literature for structural chromosome abnormalities reported in association with childhood obesity.     

From analgesia to myopathy: When local anesthetics impair the mitochondrion

The expanding utilization of local anesthesia and analgesia revealed the occurrence of myopathies induced by local anesthetics. Such iatrogenic effect encouraged anesthesiologists to study the toxicity of local anesthetics and to reevaluate their protocols in order to reduce muscle pain and dysfunction. Studies performed in rats and human cells showed that bupivacaine induces muscle histological damages with sarcomers disruption along with structural alteration of mitochondria, the powerplant of the cell. Bupivacaine-induced myopathies (BIM) are underestimated as patients are not examined by the anesthesiologist after the surgery. Biochemical analyses indicate that BIM could be explained both by the alteration of mitochondrial energetics with consecutive oxidative stress and mitophagy, and the modification of sarcoplasmic reticulum activity with perturbations of calcium homeostasis. BIM is time-dependent, local anesthetic concentration-dependent, enhanced by preexisting metabolism alteration or young age, and could be prevented in part by antioxidant agents and rhEPO. These observations suggest that adapted changes in postoperative analgesia protocols, including the adjustment of LA concentration and volume, a more precise delivery of the drug and an adapted duration of analgesia, may prevent myopathies consecutive to local anesthesia.     

The apoptotic regulator Nrz controls cytoskeletal dynamics via the regulation of Ca2+ trafficking in the zebrafish blastula

Bcl-2 family members are key regulators of apoptosis. Their involvement in other cellular processes has been so far overlooked. We have studied the role of the Bcl-2 homolog Nrz in the developing zebrafish. Nrz was found to be localized to the yolk syncytial layer, a region containing numerous mitochondria and ER membranes. Nrz knockdown resulted in developmental arrest before gastrulation, due to free Ca(2+) increase in the yolk cell, activating myosin light chain kinase, which led to premature contraction of actin-myosin cables in the margin and separation of the blastomeres from the yolk cell. In the yolk syncytial layer, Nrz appears to prevent the release of Ca(2+) from the endoplasmic reticulum by directly interacting with the IP3R1 Ca(2+) channel. Thus, the Bcl-2 family may participate in early development, not only by controlling apoptosis but also by acting on cytoskeletal dynamics and cell movements via Ca(2+) fluxes inside the embryo.     

On the characterization and selection of diverse conformational ensembles with applications to flexible docking

To address challenging flexible docking problems, a number of docking algorithms pregenerate large collections of candidate conformers. To remove the redundancy from such ensembles, a central problem in this context is to report a selection of conformers maximizing some geometric diversity criterion. We make three contributions to this problem. First, we resort to geometric optimization so as to report selections maximizing the molecular volume or molecular surface area (MSA) of the selection. Greedy strategies are developed, together with approximation bounds. Second, to assess the efficacy of our algorithms, we investigate two conformer ensembles corresponding to a flexible loop of four protein complexes. By focusing on the MSA of the selection, we show that our strategy matches the MSA of standard selection methods, but resorting to a number of conformers between one and two orders of magnitude smaller. This observation is qualitatively explained using the Betti numbers of the union of balls of the selection. Finally, we replace the conformer selection problem in the context of multiple-copy flexible docking. On the aforementioned systems, we show that using the loops selected by our strategy can improve the result of the docking process.     

Identification of novel secreted proteases during extracellular proteolysis by dermatophytes at acidic pH

The dermatophytes are a group of closely related fungi which are responsible for the great majority of superficial mycoses in humans and animals. Among various potential virulence factors, their secreted proteolytic activity attracts a lot of attention. Most dermatophyte-secreted proteases which have so far been isolated in vitro are neutral or alkaline enzymes. However, inspection of the recently decoded dermatophyte genomes revealed many other hypothetical secreted proteases, in particular acidic proteases similar to those characterized in Aspergillus spp. The validation of such genome predictions instigated the present study on two dermatophyte species, Microsporum canis and Arthroderma benhamiae. Both fungi were found to grow well in a protein medium at acidic pH, accompanied by extracellular proteolysis. Shotgun MS analysis of secreted protein revealed fundamentally different protease profiles during fungal growth in acidic versus neutral pH conditions. Most notably, novel dermatophyte-secreted proteases were identified at acidic pH such as pepsins, sedolisins and acidic carboxypeptidases. Therefore, our results not only support genome predictions, but demonstrate for the first time the secretion of acidic proteases by dermatophytes. Our findings also suggest the existence of different pathways of protein degradation into amino acids and short peptides in these highly specialized pathogenic fungi.