The mechanism of cell membrane ruffling relies on a phospholipase D2 (PLD2), Grb2 and Rac2 association

Membrane ruffling is the formation of actin rich membrane protrusions, essential for cell motility. The exact mechanism of ruffling is not fully known. Using YFP and CFP fluorescent chimeras, we show for the first time a co-localization of Phospholipase D2 (PLD2) and Growth factor Receptor Bound protein-2 (Grb2) with actin-rich membrane protrusions of macrophages. Grb2 cooperates with PLD2 in enhancing membrane ruffling, whether in resting cells or in cells stimulated with the growth factor M-CSF, although in the latter an increase in dorsal ruffles was observed, consistent with receptor-ligand internalization. Cells transfected with PLD2 mutated in the PH domain (Y169F) or with Grb2 mutated in the SH2 site (R86K) negate this effect, indicating an association PLD2(Y169)-SH2-Grb2 that was confirmed by immunoprecipitation and Western blotting. The association results in enhanced PLD activity, but the lipase activity can only partially explain the formation of membrane ruffles in vivo. A third component involves the Rho-GTPase Rac2 and it is only when Rac2 is overexpressed along with PLD2 and Rac2 that a full biological effect, including actin polymerization in vivo, is obtained. The mechanism involved is, then, as follows: PLD enzymatic action, after having been increased due to the binding to Grb2-SH2 via Y169, cooperates with Rac2, and the three molecules stimulate actin polymerization and consequently, membrane ruffle formation. Since membrane ruffling precedes cell migration, the results herein provide a novel mechanism for control of membrane dynamics, crucial for the physiology of leukocytes.     

Development of the cypriniform protrusible jaw complex in Danio rerio: Constructional insights for evolution

Studies on the evolution of complex biological systems are difficult because the construction of these traits cannot be observed during the course of evolution. Complex traits are defined as consisting of multiple elements, often of differing embryological origins, with multiple linkages integrated to form a single functional unit. An example of a complex system is the cypriniform oral jaw apparatus. Cypriniform fishes possess an upper jaw characterized by premaxillary protrusion during feeding. Cypriniforms effect protrusion via the kinethmoid, a synapomorphy for the order. The kinethmoid is a sesamoid ossification suspended by ligaments attaching to the premaxillae, maxillae, palatines, and neurocranium. Upon mouth opening, the kinethmoid rotates as the premaxillae move anteriorly. Along with bony and ligamentous elements, there are three divisions of the adductor mandibulae that render this system functional. It is unclear how cypriniform jaws evolved because although the evolution of sesamoid elements is common, the incorporation of the kinethmoid into the protrusible jaw results in a function that is atypical for sesamoids. Developmental studies can show how biological systems are assembled within individuals and offer clues about how traits might have been constructed during evolution. We investigated the development of the protrusible upper jaw in zebrafish to generate hypotheses regarding the evolution of this character. Early in development, the adductor mandibulae arises as a single unit. The muscle divides after ossification of the maxillae, on which the A1 division will ultimately insert. A cartilaginous kinethmoid first develops within the intermaxillary ligament; it later ossifies at points of ligamentous attachment. We combine our structural developmental data with published kinematic data at key developmental stages and discuss potential functional advantages in possessing even the earliest stages of a system for protrusion. J. Morphol. 2010. © 2010 Wiley‐Liss, Inc.     

Morphological and cytochemical analysis of an unusual nucleus-pyrenoid association in a unicellular red alga

Summary Protrusions of the nucleus that extend into the pyrenoid of the unicellular red algaRhodella maculata have been characterised morphologically and cytochemically. Serial reconstructions indicate that cells frequently have two tubular nuclear protrusions that are up to 1.2 m long. Cytochemical analyses were undertaken to investigate the nucleic acid content of the protrusions. DNA was not detected in the nuclear protrusions, though clear labelling was seen in all other DNA-containing zones (viz. the main nuclear compartment, the chloroplast and the mitochondria). High concentrations of RNA were observed in the nuclear protrusion. In situ hybridization experiments indicate that ribosomal RNAs are not a major component of the protrusion RNA. Possible roles of the protrusion and its RNA content are discussed.Abbreviations DNA deoxyribonucleic acid - RNA ribonucleic acid - rRNA ribosomal RNA     

Trophic apparatus in cyprinodontiform fishes: Functional specializations for picking and scraping behaviors

Cyprinodontiforms are a diverse and speciose order that includes topminnows, pupfishes, swordtails, mosquitofishes, guppies, and mollies. Sister group to the Beloniformes and Atheriniformes, Cyprinodontiformes contains approximately twice the number of species of these other two orders combined. Recent studies suggest that this group is well suited to capturing prey by “picking” small items from the water surface, water column, and the substrate. Because picking places unusual performance demands on the feeding apparatus, this mode of prey capture may rely upon novel morphological modifications not found in more widespread ram‐ or suction‐based feeding mechanisms. To assess this evolutionary hypothesis, we describe the trophic anatomy of 16 cyprinodontiform species, selected to broadly represent the order as well as capture intrageneric variation. The group appears to have undergone gradual morphological changes to become increasingly specialized for picking and scraping behaviors. We also identify a suite of functional characters related to the acquisition of a novel and previously undescribed mechanism of premaxillary protrusion and retraction, including: modification of the “premaxillomandibular” ligament (which connects each side of the premaxilla to the ipsilateral mandible, or lower jaw), a novel architecture of the ligaments and bony elements that unite the premaxillae, maxillae and palatine bones, and novel insertions of the adductor muscles onto the jaws. These morphological changes to both the upper and lower jaws suggest an evolutionary trend within this group toward increased reliance on picking individual prey from the water column/substrate or for scraping encrusting material from the substrate. We propose that the suite of morphological characters described here enable a functional innovation, “picking,” which leads to novel trophic habits. J. Morphol., 2009. © 2008 Wiley‐Liss, Inc.     

Distribution of Cadmium,Lead, Nickel,and Strontium in Imbibing Maize Caryopses

Seed germination is tolerant to heavy metals apparently because the seed coat is impermeable to metal ions. However, it is not clear whether the seed coat is a universal barrier for all metals. In addition, depending on their physical and chemical properties, a distribution of various metals may differ within an imbibing caryopsis, and therefore they produce dissimilar effects on seed germination. The toxic effects of Cd(NO₃)₂, Pb(NO₃)₂, Ni(NO₃)₂, and Sr(NO₃)₂ were estimated from the germination rates of maize (Zea mays L.) caryopses following two-day incubation with these salts. The distribution of heavy metals and Sr was studied by histochemical methods based on the formation of colored complexes with dithizone (Cd and Pb), dimethylglyoxyme (Ni), and sodium rhodizonate (Sr). Although the metals under study did not affect maize radicle protrusion, they inhibited seed germination in the following order: Cd > Ni ≈ Pb > Sr. Cd and Pb accumulated mainly in the seed coat cells, but Sr and Ni in the embryo cells and in the cells of endosperm (Sr) and scutellum (Ni). Although Cd was found only in the seed coat, it was the strongest inhibitor of seed germination. Apparently, due to high toxicity, Cd exerted its inhibitory effect at the concentrations too low for histochemical assay. In spite of easy translocation across the seed coat of imbibing caryopses, Sr did not considerably inhibit radicle protrusion and seed germination, apparently because of its low toxicity and predominant localization in the apoplast of embryo and endosperm cells.__________Translated from Fiziologiya Rastenii, Vol. 52, No. 4, 2005, pp. 635–640.Original Russian Text Copyright © 2005 by Seregin, Kozhevnikova.     

From the anther to the proctodeum: Pear (Pyrus communis) pollen digestion in Osmia cornuta larvae

Modifications of the pollen grains of Pyrus communis Linneaus that occur during the digestion by Osmia cornuta (Latreille) larvae were studied histochemically. We compared the features of the pollen grains found in the anthers, in the larval cell provisions and in the alimentary canal of the 5th instar larvae. Modifications were already evident in the provisions and consisted of protoplast protrusions through the apertures and a decrease in the number of starch-containing pollen grains. After pollen grains were ingested by the larvae, the protoplast appeared retracted from the pollen wall. Pollen digestion began in the anterior part of the midgut, where we observed: (1) disorganised intine at the apertures; (2) disappearance of DAPI staining of nuclear pollen DNA; (3) fewer pollen grains containing starch than in the anthers; (4) some empty pollen grains. Pollen grains in the proctodeum appeared extremely compressed and crushed. Some grains appeared to be unaffected by the digestive process. We hypothesise that the protrusion of the intine and of the protoplast from the apertures in bee provisions could be considered a kind of pre-treatment necessary to initiate the digestion process in the larval alimentary canal.     

Anatomy and functional morphology of the feeding apparatus of the lesser electric ray, Narcine brasiliensis (Elasmobranchii: Batoidea)

Protrusion of the jaws during feeding is common in Batoidea (rays, skates, sawfishes, and guitarfishes), members of which possess a highly modified jaw suspension. The lesser electric ray, Narcine brasiliensis, preys primarily on polychaete annelids using a peculiar and highly derived mechanism for jaw protraction. The ray captures its prey by protruding its jaws beneath the substrate and generating subambient buccal pressure to suck worms into its mouth. Initiation of this protrusion is similar to that proposed for other batoids, in that the swing of the distal ends of the hyomandibulae is transmitted to Meckel's cartilage. A "scissor-jack" model of jaw protrusion is proposed for Narcine, in which the coupling of the upper and lower jaws, and extremely flexible symphyses, allow medial compression of the entire jaw complex. This results in a shortening of the distance between the right and left sides of the jaw arch and ventral extension of the jaws. Motion of the skeletal elements involved in this extreme jaw protrusion is convergent with that described for the wobbegong shark, Orectolobus maculatus. Narcine also exhibits asymmetrical protrusion of the jaws from the midline during processing, accomplished by unequal depression of the hyomandibulae. Lower jaw versatility is a functional motif in the batoid feeding mechanism. The pronounced jaw kinesis of N. brasiliensis is partly a function of common batoid characteristics: euhyostylic jaw suspension (decoupling the jaws from the hyoid arch) and complex and subdivided cranial musculature, affording fine motor control. However, this mechanism would not be possible without the loss of the basihyal in narcinid electric rays. The highly protrusible jaw of N. brasiliensis is a versatile and maneuverable feeding apparatus well-suited for the animal's benthic feeding lifestyle.     

Actin and Microtubules in Cell Motility: Which One is in Control?

The cytoskeleton is composed of three distinct elements: actin microfilaments, microtubules and intermediate filaments. The actin cytoskeleton is thought to provide protrusive and contractile forces, and microtubules to form a polarized network allowing organelle and protein movement throughout the cell. Intermediate filaments are generally considered the most rigid component, responsible for the maintenance of the overall cell shape. Cytoskeletal elements must be coordinately regulated for the cell to fulfill complex cellular functions, as diverse as cell migration, cell adhesion and cell division. Coordination between cytoskeletal elements is achieved by signaling pathways, involving common regulators such as the Rho guanosine-5'-triphosphatases (GTPases). Furthermore, evidence is now accumulating that cytoskeletal elements participate in regulating each other. As a consequence, although their functions seem well defined, they are in fact overlapping, with actin playing a role in membrane trafficking and microtubules being involved in the control of protrusive and contractile forces. This cytoskeletal crosstalk is both direct and mediated by signaling molecules. Cell motility is a well-studied example where the interplay between actin and microtubules appears bidirectional. This leads us to wonder which, if any, cytoskeletal element leads the way.