Planar polarization of the denticle field in the Drosophila embryo: roles for Myosin II (zipper) and fringe

Epithelial planar cell polarity (PCP) allows epithelial cells to coordinate their development to that of the tissue in which they reside. The mechanisms that impart PCP as well as effectors that execute the polarizing instructions are being sought in many tissues. We report that the epidermal epithelium of Drosophila embryos exhibits PCP. Cells of the prospective denticle field, but not the adjacent smooth field, align precisely. This requires Myosin II (zipper) function, and we find that Myosin II is enriched in a bipolar manner, across the parasegment, on both smooth and denticle field cells during denticle field alignment. This implies that actomyosin contractility, in combination with denticle-field-specific effectors, helps execute the cell rearrangements involved. In addition to this parasegment-wide polarity, prospective denticle field cells express an asymmetry, uniquely recognizing one cell edge over others as these cells uniquely position their actin-based protrusions (ABPs; which comprise each denticle) at their posterior edge. Cells of the prospective smooth field appear to be lacking proper effectors to elicit this unipolar response. Lastly, we identify fringe function as a necessary effector for high fidelity placement of ABPs and show that Myosin II (zipper) activity is necessary for ABP placement and shaping as well.     

How to induce non-polarized cells of hepatic origin to express typical hepatocyte polarity: generation of new highly polarized cell models with developed and functional bile canaliculi

Few in vitro models expressing complex hepatocyte polarity are available. We used the unpolarized rat Fao cell line to isolate the polarized WIF-B line. These complex rat-human hybrid cells form functional simple bile canaliculi. To obtain Fao-derived polarized models with a simpler chromosome content and developed bile canaliculi, we employed two approaches. Partial success was achieved with monochromosomal hybrids. As shown by the immunolocalization of apical, basolateral, and tight-junctional proteins, monochromosomal hybrid 11-3 cells were polarized. They formed simple functional bile canaliculi and transiently expressed the typical polarity of simple epithelial cells. One subclone blocked in this polarity state was isolated. A more robust approach was provided by spheroid culture, a three-dimensional system that strengthens cell-cell contacts. Transient spheroid culture induced irreversible polarization of Fao cells. This induction occurred in most spheroids (approximately 1% of the cells). From populations enriched in stably polarized cells, we generated new polarized cell models, designated Can. Can 3-1 cells formed simple functional bile canaliculi when plated at high density. Regardless of plating density, Can 9 and Can 10 cells formed long tubular branched canaliculi competent for vectorial transport of organic anions and bile acids, and involving several dozen adjacent cells. Thus, we have generated new cell models stably expressing typical hepatocyte polarity. Among these models, Can 9 and Can 10 are the first capable of forming functional, highly developed bile canaliculi similar to those formed in vivo. This work was supported by grants from the Association pour la Recherche sur le Cancer (no.6551), the Institut Curie (PIC Signalisation Cellulaire, no. 914) and the Institut National de la Santé et de la Recherche Médicale (contract PRISME 98-09).     

Interpretation of the dissolution of insoluble peptide sequences based on the acid-base properties of the solvent

The dissolution process of model insoluble peptide sequences was investigated in view of the electron acceptor (AN) and electron donor (DN) solvent properties. The Alzheimer's disease-inducing (1-42) Abeta-amyloid peptide and its (1-21) fragment, the (66-97) transmembrane bradykinin B2 receptor sequence, and the strongly aggregated VVLGAAIV were selected as models of insoluble peptides. Solvents presenting similar AN and DN values failed, despite their polarities, to dissociate peptide chains (free in solution or bound to a polymer). The maximum solubility of these aggregated sequences was attained in solvents presenting the highest possible (AN-DN) values (in positive or negative mode). The AN-DN values ranged from approximately -20 to +80 and, notably, the lowest dissociation power was ascribed to solvents presenting values of approximately +40. The strong hydrogen bond donor water is located in this region, indicating that, for dissociation of specific insoluble segments, the solvent should appropriately combine its acid/base strength with the potential for van der Waals interactions. We also observed a sequence-dependent pH effect on peptide solubility confirmed through circular dichroism spectroscopy. This approach also revealed a complex but, in many cases, consistent influence of peptide conformation on its solubility degree, even when structure-inducing solvents were added. In conclusion, the random method of selecting solvents to dissolve insoluble and intractable peptide sequences, still in use by some, could be partially supplanted by the strategy described herein, which may be also applicable to other solute dissociation processes.     

A simple time delay model for eukaryotic cell cycle

We propose a seven variable model with time delay in one of the variables for the cell cycle in higher eukaryotes. The model consists of four important phosphorylation-dephosphorylation (P-D) cycles that govern the cell cycle, namely Pre-MPF-MPF, Cdc25P-Cdc25, Wee1P-Wee1 and APCP-APC. Other variables are cyclin, free cyclin dependent kinase (Cdk) and mass. The mass acts as a G2/M checkpoint and the checkpoint is represented by a saddle node loop bifurcation. The key feature of the model is that a time lag has been introduced in the activation of anaphase promoting complex (APC) by maturation promoting factor (MPF). This is effected by treating MPF as a time-delayed variable in the activation step of APC. The time lag acts as a spindle checkpoint. Absence of time delay induces a bistability in our model. Time delay also brings about variability in G1 phase timings. The model also reproduces the mutant phenotype experiments on wee1 cells. Stochasticity has been introduced in the model to simulate the dependence of the cycle time on cell birth length. Mutant phenotypes in the stochastic model reproduce the experimental observations better than the deterministic model.     

Inversin, Wnt signaling and primary cilia

Mutations of the ankyrin-repeat protein Inversin, a member of a diverse family of more than 12 proteins, cause nephronophthisis (NPH), an autosomal recessive cystic kidney disease associated with extra-renal manifestations such as retinitis pigmentosa, cerebellar aplasia and situs inversus. Most NPH gene products (NPHPs) localize to the cilium, and appear to control the transport of cargo protein to the cilium by forming functional networks. Inversin interacts with NPHP1 and NPHP3, and shares with NPHP4 the ability to antagonize Dishevelled-stimulated canonical Wnt signaling, potentially through recruitment of the Anaphase Promoting Complex (APC/C). However, Dishevelled antagonism may be confined towards the basal body, thereby polarizing motile cilia on the cells of the ventral node and respiratory tract. Inversin is essential for recruiting Dishevelled to the plasma membrane in response to activated Frizzled, a crucial step in planar cell polarity signaling. During vertebrate pronephros development, the Inversin-mediated translocation of Dishevelled appears to orchestrate the migration of cells and differentiation of segments that correspond to the mammalian loop of Henle. Thus, defective tubule migration and elongation may contribute to concentration defects and cause cyst formation in patients with NPH.     

The eggshell in the C. elegans oocyte-to-embryo transition

In egg-laying animals, embryonic development takes place within the highly specialized environment provided by the eggshell and its underlying extracellular matrix. Far from being simply a passive physical support, the eggshell is a key player in many early developmental events. Herein, we review current understanding of eggshell structure, biosynthesis, and function in zygotic development of the nematode, C. elegans. Beginning at sperm contact or entry, eggshell layers are produced sequentially. The earlier outer layers are required for secretion or organization of inner layers, and layers differ in composition and function. Developmental events that depend on the eggshell include polyspermy barrier generation, high fidelity meiotic chromosome segregation, osmotic barrier synthesis, polar body extrusion, anterior-posterior polarization, and organization of membrane and cortical proteins. The C. elegans eggshell is proving to be an excellent, tractable system to study the molecular cues of the extracellular matrix that instruct cell polarity and early development.     

Co-regulation of polar mRNA transport and lifespan in budding yeast Saccharomyces cerevisiae

Recent studies have uncovered the links between aging, rejuvenation and polar protein transport in the budding yeast Saccharomyces cerevisiae. Here, we examined a still unexplored possibility for co-regulation of polar mRNA transport and lifespan. To monitor the amount and distribution of mRNA-containing granules in mother and daughter cells, we used a fluorescent mRNA-labeling system, with MFA2 as a reporter gene. The results obtained showed that deletion of the selected longevity regulators in budding yeast had a significant impact on the polar mRNA transport. This included changes in the amount of mRNA-containing granules in cytoplasm, their aggregation and distribution between the mother and daughter cells. A significant negative correlation was found between strain-specific longevity, amount of granules and total fluorescent intensity both in mother and daughter cells. As indicated by the coefficient of determination, approximately 50–75% of variation in yeast lifespan could be attributed to the differences in polar mRNA transport.     

N-cadherin: A new player in neuronal polarity

Comment on: Gärtner A, et al. EMBO J 2012; <span class="b">31</span>:1893-903     

Shifts in soil organic carbon for plantation and pasture establishment in native forests and grasslands of South America

The replacement of native vegetation by pastures or tree plantations is increasing worldwide. Contradictory effects of these land use transitions on the direction of changes in soil organic carbon (SOC) stocks, quality, and vertical distribution have been reported, which could be explained by the characteristics of the new or prior vegetation, time since vegetation replacement, and environmental conditions. We used a series of paired‐field experiments and a literature synthesis to evaluate how these factors affect SOC contents in transitions between tree‐ and grass‐dominated (grazed) ecosystems in South America. Both our field and literature approaches showed that SOC changes (0–20 cm of depth) were independent of the initial native vegetation (forest, grassland, or savanna) but strongly dependent on the characteristics of the new vegetation (tree plantations or pastures), its age, and precipitation. Pasture establishment increased SOC contents across all our precipitation gradient and C gains were greater as pastures aged. In contrast, tree plantations increased SOC stocks in arid sites but decreased them in humid ones. However, SOC losses in humid sites were counterbalanced by the effect of plantation age, as plantations increased their SOC stocks as plantations aged. A multiple regression model including age and precipitation explained more than 50% (p < 0.01) of SOC changes observed after sowing pastures or planting trees. The only clear shift observed in the vertical distribution of SOC occurred when pastures replaced native forests, with SOC gains in the surface soil but losses at greater depths. The changes in SOC stocks occurred mainly in the silt+clay soil size fraction (MAOM), while SOC stocks in labile (POM) fraction remained relatively constant. Our results can be considered in designing strategies to increase SOC storage and soil fertility and highlight the importance of precipitation, soil depth, and age in determining SOC changes across a range of environments and land‐use transitions.     

Seasonal variation in nitrogen uptake and turnover in two high-elevation soils: mineralization responses are site-dependent

In arctic and alpine ecosystems, soil nitrogen (N) dynamics can differ markedly between winter and summer months, and nitrogen losses can be measurable during the spring and fall transitions. To explore the effect of seasonality on biogeochemical processes in a temperate alpine environment, we used a combination of field incubations (year-round) and ¹⁵N tracer additions (late fall, early spring, summer) to characterize soil N dynamics in a wet and dry meadow in the Sierra Nevada, California. The snowmelt to early summer season marked a period of high ¹⁵N uptake and turnover in the two soils, coincident with the increase in microbial N pools at the start of snowmelt (wet and dry meadow); an increase in net N mineralization and net nitrification as snowmelt progressed (wet meadow only); and measureable net production of ¹⁵N-NH₄ ⁺ in mid-summer (wet and dry meadow). Whereas fluctuations in microbial biomass were generally synchronous between the wet and dry meadow soils, only wet meadow soils appeared to mineralize N in response to declines in the microbial N pool. Net N mineralization and net nitrification rates in the dry meadow soil were negligible on all but one sampling date, in spite of periodic decreases in biomass of up to 60%. Across both sites, high ¹⁵N recoveries in microbial biomass N, rapid ¹⁵N-NH₄ ⁺ turnover, and low or negative net ¹⁵N-NH₄ ⁺ fluxes suggested tight cycling of N, particularly in the late fall and early spring.