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4L

Structure-function studies of egg proteins of invasive molluscs, plagues and vectors of parasites


Director

Heras, Horacio

Researchers
Dreon, Marcos Sebastián
Ituarte, Santiago
Pasquevich, María Yanina

Fellows - Thesis Candidates
Brola, Tabata Romina
Cadierno, María Pilar
Giglio, Matiás Leonel


foto 4 chica Team of work

Collaborators from other institutions
International
Jian-Wen Qiu
Hong Kong Baptist University, Republica Popular de China.

National
Ceolín, Marcelo
INIFTA -Research Institute of Theoretical and Applied Physical Chemistry (CONICET - CCT- La Plata - UNLP). La Plata. Province of Buenos Aires. Argentina
Fernández, Patricia
Cathedra of Pathology, Faculty of Veterinary Science. UNLP. La Plata. Province of Buenos Aires. Argentina

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Objetives

Our group is focused on classifying, identifying and studying the structure and functions of unknown egg proteins of invasive snails, agricultural plague and parasite vectors. We endeavour to understand the snail reproductive strategies from a biochemical perspective and, at the same time, to select some of these new proteins with potentials as bioactive compounds to exploit its application in Biomedicine.

Importance of the study of snail proteins

The knowledge of the structure and protein function is of key relevance for the understanding of all the biological processes. Gastropod molluscs are a source of bioactive molecules commonly used in Pharmacology and Medicine, since they have an outstanding diversity of compounds as reflection of their long evolutionary history. However, proteins of mollusc eggs are very little investigated. Our research group is studying them taking as a model those of Pomacea canaliculata (Lamarck, 1822), also called ampularia, an aquatic snail, worldwide invader which has turned out to be a plague for rice fields. Its uncontrolled expansion entailed the advance of a vector nematode parasite, responsible for human eosinophilic meningoencephalitis in Asia and Latin America. Its eggs strongly coloured by pigmented proteins almost have no predators.

Questions expected to answer

What kind of proteins do the eggs of this snail have?
Which are the properties of these proteins and what role do they play in the defence mechanism of the embryos?
Doe they have any application as tools in Biomedicine?

Employed methodology

Our group uses a multidisciplinary approach with collaborations of other research groups of our country and abroad in order to answer these questions, usually employing a combination of methodologies such as biochemical, molecular biology, biophysics, cellular biology, histopathological and bioassays with laboratory animals. Some of the techniques used are described as follows:

  • Determination of amino acid sequence.
  • Study of biochemical activity of inhibitors, lectins, enzymes isolated from the eggs.
  • Study of structure/function of proteins through dispersion of angle x-rays and biophysical techniques.
  • Microbiological studies of inhibition of bacterial growth.
  • In vitro studies of toxicity and interaction with cultured tumour cells.
  • Basic immunological studies.
  • Toxicity bioassays in murine model.
  • Use of optical and electronic microscopy for histopathological studies, effects on the nervous and digestive systems of potential predators

Main results

We studied the proteome of egg fluid and demonstrated that P. canaliculata has achieved a fascinating antipredatory defence mechanism where perivitellins play a central role, providing the embryos with one of the best defences in animals at biochemical level. We characterize major perivitellins (PcOvo y PcPV2) that participate in the defence as follows:

PcOvo caroprotein provides photoprotective and antioxidant compounds which at the same time give an intense colour. Its striking colouring would warn potential predators of the presence of various protein toxins of perivitellin fluid surrounding the embryo. PcOvo also has associated an activity as protease inhibitor of gastrointestinal tract of the predator, limiting the digestion of egg nutrients (antidigestive defence). Orally administered, it diminishes the growth rate in rats and impedes nutrient acquisition.

PcPV2 is lethal for rodents and is the first protein toxin described within an egg. Its structure differs from all other animal toxins since it combines a lectin subunit that recognizes specific sequences of membrane sugars, and a subunit that forms pores in the membrane. Despite the fact that there is no equal in animals, it is similar to some toxins type AB described in seeds and to the botulinum toxin. Whether this novel defence system is present in the eggs of other species remains unknown. We continue studying its properties and mechanism of action. However, we study the major egg protein of Pomacea scalaris so-called scalarin (PsSC) that provides the eggs with a striking rose-salmon colour that could function as a warning signal. We demonstrated that it is a powerful lectin that shows, nevertheless, many structural and functional similarities with PcOvo, that it is not.

We are studying its defence function as toxin lectin on the digestive system and its potential as biomarker of tumour cells. Other gastropod lectins have impact on medical practice and are commercialized to improve the pharmacokinetics of glycoprotein drugs or in the diagnosis of diseases such as tumour markers.

The structure and function of these three proteins are very resistant to high temperatures, to a very range of pH and to gastrointestinal digestion, what allows reaching biologically active intestines, highlighting even more its biotechnological application.

Evolutional and ecological implications

The selection pressure exerted by predators and hostile environment on the eggs would lead to the acquisition of new properties to endure the air development of an aquatic animal. At biochemical level it seems that these adaptations would involve a new type of perivitellins that besides being a reserve protein they are multifunctional compounds provided with very efficient defences against predation together with protective properties of the embryo facing hostile environment conditions.
The presence of eggs protected with proteins of these characteristics has not been found in other members of the animal kingdom, but it is similar to the plant defences against herbivory.


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Publications

  • Novel animal defenses against predation: A snail egg neurotoxin combining lectin and pore-forming chains that resembles plant defense and bacteria attack toxins.
    Dreon, M. S; Frassa, M. V; Ceolín, M; Ituarte, S; Qiu, J. W; Sun, J; Fernández, P. E;  Heras, H.
    2013.
    Plos one. San Francisco: Public Library Science, vol. 8, n° e637, p. 1-11. ISSN 1932-6203

  • First proteome of the egg perivitelline fluid of a freshwater gastropod with aerial oviposition.
    Sun, J; Zhang, H; Wang, H; Heras, H; Dreon, M. S; Ituarte, S; Ravasi, T; Qian, P; Qiu, J. W.
    2012. Journal of Proteome Research , Washington: Amer Chemical Soc, vol. 11, p.
    2440-2448

  • Agglutinating activity and structural characterization of scalarin  the major egg protein of the snail Pomacea scalaris.
    Ituarte, S; Dreon, M. S; Ceolin, M; Heras, H. (d’Orbigny, 1832).
    2012. Plos One 7(11): e50115. Doi:10.1371/journal.pone.0050115.


  • Antioxidant response and oxidative stress levels in Macrobrachium borellii (Crustacea: Palaemonidae)exposed to the water-soluble fraction of petroleum.
    Lavarías, S; Heras, H;  Pedrini, N; Turnier, H; Ansaldo, M.
    2011. Comp. Biochem. Physiol. 153 C: 415-421. doi:10.1016/j.cbpc.2011.02.002
  • Triacylglycerol catabolism in the prawn Macrobrachium borellii (Crustacea: Palaemoniade).
    Pasquevich, M. Y; Dreon, M. S; Lavarías, S; Heras, H.
    2011. Comp. Biochem. Physiol. 160 B: 201-207. doi:10.1016/j.cbpb.2011.08.006
  • Structure and stability of the neurotoxin PV2 from the eggs of the apple snail Pomacea canaliculata.
    Frassa, V; Ceolin, M; Dreon, M. S; Heras, H.
    2010. Biochimica and biophysica acta. Elsevier. vol. 1804, p. 1492-1499
  • The Role of the Proteinase Inhibitor Ovorubin in Apple Snail Eggs Resembles Plant Embryo Defenses Against Predation.
    Dreón, M. S; Ituarte, S; Heras, H.
    2010. Plos one. Public Library Science. vol. 5, n 12, p. 15059-15068
  • Carbohydrates and glycoforms of the major egg perivitellins from Pomacea apple snails (Architaenioglossa: Ampullariidae).
    Ituarte, S; Dreón, M. S; Pasquevich, M. Y; Fernández, P. E; Heras, H.
    2010. Comparative biochemistry and physiology. part b, biochemistry & molecular biology. Elsevier Science Inc. vol. 157, p. 66-72
  • Isolation and characterization of a novel perivitellin from the eggs of Pomacea scalaris (Mollusca, Ampullariidae).
    Ituarte, S; Dreon, M. S; Ceolin, M; Heras, H.
    2008. Molecular reproduction and development.Wiley-Liss, vol. 75, p. 1441-1448
  • First egg protein with a neurotoxic effect on mice.
    Heras, H; Frassa, V; Fernández, P. E; Galosi, C. M; Gimeno, E; Dreon, M. S.
    2008. Toxicon. Elsevier, vol. 52, p. 481-488
  • Global shape and pH stability of ovorubin, an oligomeric protein from the eggs of Pomacea canaliculata.
    Dreón, M. S; Ituarte, S; Ceolín, M; Heras, H.
    2008. Febs journal. Blackwell Pub. on behalf of the Federation of European Biochemical Societies, vol. 275, n° 18, p. 4522-4530. ISSN 1742-464X
  • Astaxanthin binding and structural stability of apple snail carotenoprotein ovorubin.
    Dreon, M. S; Ceolin, M; Heras, H. 
    2007. Archives of biochemistry and biophysics. vol. 30, p. 359-365
  • Egg carotenoproteins of neotropical ampullariidae (gastropoda: arquitaenioglossa).
    Heras, H; Dreon, M. S; Ituarte, S; Pollero, R. J.
    2007. Comparative biochemistry and physiology. toxicology & pharmacology. vol. 146, p. 158-167

Book - Book Chapters

  • Changes in carbohydrate expression in the cervical spinal cord of mice intoxicated with perivitellin PV2 from Pomacea canaliculata. In: Poisoning by plants, mycotoxins and related toxins.
    Fernández, P. E; Frassa, V; Gimeno, E. J; Dreon, M; Heras, H.
    2011. Edited by Riet-Correa, F., Pfister J., Schild A.L. and Panter K. CAB International, Oxfordshire, UK  482-498. ISBN 13 978 1 84593 833 8. Chapter 83

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International

  • The major egg protein of pomaceanscalaris is a lipo-glyco-carotenoprotein with citotoxic effect on CACO2 cells.
    Ituarte, S; Heras, H; Dreon, M. S.
    2013. La Plata. Resumen. Congreso. 8th International Conference on Lipid Binding Proteins. INIBIOLP-CCT La Plata.
  • Defensive proteins in Pomacea: Hemagglutinating activity of PsSC, the major egg perivitellin of P. scalaris.
    Ituarte, S; Dreon, M. S; Heras, H.
    2012.
    Santiago de Compostela. España. Congreso. Physiomar 12. Universidad de
    Santiago de Compostela.

  • PcPV2 neurotoxin from the Apple snail Pomacea canaliculata eggs: A perivitellin combining perforin and lectin-like chains.
    Dreon, M. S; Ituarte, S; Frassa, V; Fernández, P. E; Heras, H. 
    2012.
    Santiago de Compostela. España. Congreso. Physiomar 12. Universidad de Santiago de Compostela.

  • Transferencia de lípidos in vitro entre lipoproteínas y divertículos intestinales en la araña Polybetes pythagoricus (Araneae, Sparassidae).
    García, F; Laino, A; Heras, H; Herrera Hidalgo, J; Cunningham, M.
    2011. Montenegro. Colombia. III Congreso Latinoamericano de Aracnología.
  • Colorfull and nutritious but lethal and antidigestive. The multiple roles of egg proteins in the successful reproductive strategy of Pomacea canaliculata (Mollusca: Gastropoda)
    2011. Hong Kong Baptist University, China. Invitado por el Dr. J-W Qiu, Department of Biology.
  • Antinutritive perivitellins in Pomacea Perry, 1810: The case of P.scalaris (d Orbigny 1835) (Architaenioglossa, Ampullariidae).
    Ituarte, S; Dreon, M. S; Heras, H.
    2010. Quebec: Quebec.  Resumen. Congreso. Physiomar 10.
  • Biochemical defenses against predation of apple snail eggs resemble plant strategies against herbivory.
    Dreon, M. S; Ituarte, S; Frassa, V; Heras, H.
    2010. Quebec: Quebec. Resumen. Congreso. Physiomar 10.


  • Changes in carbohydrate expression in the cervical spinal cord of mice intoxicated with perivitellin PV2 from Pomacea canaliculata.
    Fernández P. E; Frassa, V; Gimeno, E; Dreon, M. S; Heras, H.
    2009. João Pessoa, Brasil: João Pessoa, Brasil. Artículo Breve. Congreso.
    8th International Symposium on Poisonous Plants (ISOPP 8). Federal University of Campina Grande.
  • Structure-function relationships of ovorubin, a protease inhibitor from the eggs of Pomacea canaliculata.
    Dreon, M. S; Ituarte, S; Ceolin, M; Heras, H.
    2009. Buzios, Brasil. Resumen. Congreso. VII Iberoamerican Congress of Biological Physics. Sociedad Argentina de Biofísica.
  • Changes in carbohydrate expression in the cervical spinal cord of mice intoxicated with perivitellin PV2 from Pomacea canaliculata.
    Fernández P. E; Frassa, V; Gimeno, E; Dreon, M. S; Heras, H.
    2009. João Pessoa, Brasil: João Pessoa, Brasil. Artículo Breve. Congreso.
    8th International Symposium on Poisonous Plants (ISOPP 8). Federal University of Campina Grande.
  • Structure-function relationships of ovorubin, a protease inhibitor from the eggs of Pomacea canaliculata.
    Dreon, M. S; Ituarte, S; Ceolin, M; Heras, H.
    2009. Buzios, Brasil. Resumen. Congreso. VII Iberoamerican Congress of Biological Physics. Sociedad Argentina de Biofísica.
  • Perivitelinas resistentes a la digestión en Pomacea Perry,1810 (Architaenioglossa, Ampullariidae). ¿Un mecanismo antinutritivo de defensa?
    Ituarte, S; Dreon, M. S; Ceolin, M; Heras, H.
    2008. Valdivia, Chile: Valdivia, Chila. 2008. Resumen. Congreso. VII Congreso Latinoamericano de Malacología (CLAMA). Universidad Austral de Chile.


  • Low Resolution Structure and pH stability of Ovorubin, a lipoglycocarotenoprotein from the eggs of Pomacea canaliculata.
    Ituarte, S; Dreon, M. S; Ceolin, M; Heras, H.
    2007. Montevideo, Uruguay: ICBP. Resumen. Conferencia. International Conference of Biophysics of Proteins (ICBP). ICBP.
  • PH stability and Low Resolution Structure of Ovorubin, a protease inhibitor from the eggs of Pomacea canaliculata.
    Ituarte, S; Dreon, M. S; Ceolin, M; Heras, H.
    2007. Acapulco, Mexico: Latin American Protein Society Meeting. Resumen. Congreso. 2nd Latin American Protein Society Meeting. 2nd Latin American Protein Society Meeting.
  • Structural characterization of thermal and urea denaturation of the Pv2 apple snail perivitellin.
    Frassa, V; Dreon, M. S; Ceolin, M; Heras, H.
    2007. Montevideo, Uruguay: ICBP. Resumen. Conferencia.
    International Conference of Biophysics of Proteins (ICBP).


National

  • Aislamiento y caracterización parcial de un inhibidor de proteasas de los huevos de Pomacea Canaliculata (Gastropoda: Ampullariidae).
    Brola, T; Dreon, M. S; Heras, H.
    2013. La Plata. Resumen.  I Congreso Argentino de Malacología (I CAM). FCNyM, UNLP.
  • Efecto de los huevos de Pomacea canaliculata sobre la morfofisiología intestinal de rata. Una nueva defensa antidepredador.
    Heras, H; Dreon, M. S; Fernández, P. E; Gimeno, E.
    2013. Resumen. Taller. Pomacea 2013 V Taller ¨Biología de Ampullariidae¨. Monte Hermoso. UNS.
  • Estudio comparado de la carotenoproteína PMPV1 con otras carotenoproteínas presentes en huevos de Pomacea (Gastropoda: Ampullariidae).
    Pasquevich, M. Y; Dreon, M. S; Ituarte, S; Heras, H.
    2013. La Plata. Resumen. Congreso. I Congreso Argentino de Malacologia (I CAM). FCNyM-UNLP.
  • Estudio de la carotenoproteína mayoritaria de los huevos de Pomacea maculata y su relación con las carotenoproteínas de Pomacea canaliculata y Pomacea scalaris.
    Pasquevich, M. Y; Dreon, M. S; Heras, H.
    2013. Monte Hermoso. UNS. Resumen. Taller. V Taller ¨Biología de Ampullariidae¨.
  • Presencia de un órgano tóxico en el sistema reproductor de Pomacea canaliculata (Lamarck, 1822).
    Cadierno, M. P; Dreon, M. S; Heras, H.
    2013. La Plata. Resumen. Congreso. I Congreso Argentino de Malacología (I CAM). FCNyM, UNLP.
  • Estudio de la composición de los huevos de Pomacea maculata Perry, 1810 y caracterización preliminar de la proteína responsable de sus efectos neurotóxicos.
    Giglio, M; Heras, H; Ituarte, S.
    2013. Monte Hermoso. UNS. Resumen. Taller Pomacea, V Taller ¨Biología de Ampullariidae¨.
  • Inversión maternal en proteínas defensivas en Pomacea: PsSc, lectina citotóxica de Pomacea scalaris.
    Ituarte, S; Brola, T; Dreon, M. S; Heras, H.
    2013. Monte Hermoso. UNS. Resumen. Taller. V Taller Biología de Ampullariidae.
  • La glándula del albumen de Pomacea canaliculata (Lamark,1822) como potencial alimento. Evaluación bioquímica y análisis estructural y funcional de la neurotoxina PcPv2.
    Cadierno, M. P; Dreon, M. S; Heras, H.
    2013. Monte Hermoso. UNS. Resumen. Taller Pomacea 2013, V Taller ¨Biología de Ampullariidae¨
  • Métodos bioquímicos para diferenciar los huevos de dos especies simpátricas: Pomacea maculata y Pomacea canaliculata (Gastropoda: Ampullariidae).
    Pasquevich, M. Y; Dreon, M. S; Heras, H.
    2013. La Plata. Resumen. Congreso. I Congreso Argentino de Malacologia (I CAM). FCNyM-UNLP.
  • First insight into the major egg protein from the apple snail Pomacea insularum perivitellin fluid.
    Pasquevich, M. Y; Dreon, M. S; Heras, H.
    2012. Mendoza. Congreso.
    XLVIII Reunión SAIB y BM.

  • Coloridas y nutritivas pero letales y antidigestivas. El rol múltiple de las proteínas de los huevos en la exitosa estrategia reproductiva de Pomacea.
    Heras, H; Ituarte, S; Frassa, M. V; Dreon, M. S.
    2011. Pto Madryn, Argentina. VIII Congreso Latinoamericano de Malacología (CLAMA).
  • Effect of organophosphate fenitrothion on biochemical parameters of the crustacean Macrobrachium borellii.
    Lavarías, M.S;  García; Crespo, R; Pedrini, N; Heras, H.
    2011. Potrero de los Funes, Argentina. 30 octubre - 2 Noviembre. XLVII Reunión SAIByBM.
  • Hemagglutinating activity and structural stability of a snail egg-protein.
    Ituarte, S; Dreon, M. S;Heras, H.
    2011. Potrero de los Funes, Argentina. 30 octubre - 2 Noviembre. XLVII Reunión SAIByBM.
  • Effect of water-soluble crude oil hydrocarbon exposure on protein expression of a freshwater crustacean.
    Pasquevich, M. Y; Dreon, M. S; Heras, H.
    2011. Potrero de los Funes, Argentina. 30 octubre - 2 Noviembre. XLVII Reunión SAIByBM
  • Effects of urban discharge on lipid and fatty acid composition of Diplodon chilensis.
    Yussoppone, M. S; Pasquevisch, Y; Heras, H; Luquet, C. M; Ríos de Molina, M. C; Rocchetta, I.
    2011. Potrero de los Funes, Argentina. 30 octubre - 2 Noviembre. XLVII Reunión SAIByBM
  • Rol de la neurotoxina PV2 en la defensa de los embriones de Pomacea canaliculata.
    2011. La Plata, Argentina. Invitado por Dr. Catanesi, Instituto de Biología Celular IMBICE.
  • Multifunctional perivitellins in Pomacea: Characteristic of aerial eggs.
    Ituarte, S; Dreon, M. S; Heras, H.
    2010. Tucumán: Tucumán. Resumen. Workshop. IV International workshop on the biology of Ampullariidae.
  • Pomacea canaliculata perivitellins. Storage proteins or defensive molecules.
    Heras, H; Ituarte, S; Frassa, V; Dreon, M. S.
    2010. Tucumán: Tucumán. Resumen. Workshop. IV International workshop on the biology of Ampullariidae. INIBIOLP.
  • PV2: a novel toxin isolated from the eggs of P.
    Frassa, V; Dreon, M. S; Heras, H.
    2010. Canaliculata. S.M. Tucumán: Argentina. 2010. Resumen. Workshop.
    IV International workshop on the biology of Ampullariidae. INIBIOLP.
  • The role of the proteinase inhibitor ovorubin in the apple snail eggs defenses against predation.
    Dreon, M. S; Ituarte, S; Frassa, V; Heras, H.
    2010. Tucumán: Tucumán. Resumen. Workshop. IV International Workshop on the Biology of Ampullariidae.

  • Structure and stability of the snail neurotoxin PV2.
    Frassa, V; Dreon, M. S; Ceolin, M; Heras, H. 
    2009. Tucumán. Resumen. Congreso.
    XLV Reunión SAIB y BM. XLV Reunión SAIB y BM.

  • First egg protein with a neurotoxic effect on mice.
    Frassa, V; Fernández, P. E; Galosi, C. M; Gimeno, E; Dreon, M. S; Heras, H.
    2008. Va. Carlos Paz, Argentina: Biocell. 2008. Resumen. Congreso.
    XLIV Reunión SAIB y BM.
  • Ovorubin as a protease inhibitor in Pomacea canaliculata eggs. Implications in the biochemical defenses of snail embryos.
    Dreon, M. S; Ituarte, S; Ceolin, M; Heras, H.
    2008. La Plata. Resumen. Congreso.
    XXXVII Congreso Soc. Argentina de Biofísica. SAB.
  • Inducción de la síntesis de vitelogeninas por efecto del plaguicida organofosforado fenitrothion sobre el camarón Macrobrachium borelii.
    García, F; Cunningham, M; Pifano, M; Heras, H.
    2008. Mar del Plata, Buenos Aires: Sociedad de Toxicología y Química Ambiental. Simposio SETAC Argentina. Resumen. Congreso.
    Sociedad de Toxicología y Química Ambiental. Simposio SETAC Argentina.


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linea Awards

  • “IFS Jubilee Award 2006”
    Premio otorgado a los investigadores sobresalientes de proyectos subsidiados por la Internacional Foundation for Science  al Dr. Heras Horacio. Suecia – 2006.


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INIBIOLP - Instituto de Investigaciones Bioquímicas de La Plata - Consejo Nacional de Investigaciones Científicas y Técnicas - Facultad de Ciencias Médicas - U.N.L.P. Avenida 60 y 120 - C.P. 1900 La Plata -
Provincia de Buenos Aires - Argentina. Tel. 54-221-4824894 - Fax. 54-221-4258988 - email: info@inibiolp.org.ar