Newsletter of the Pineapple Working Group, International Society for Horticultural Science Proteases Expressed in Response to in Vitro Culture of Pineapple (Ananas comosus (L.) Merr.) Hernández M , Pérez A , Lorenzo J , Carvajal C , Mora M , Natalucci C , Jorrín J. 1 1 2 1 1 3 4 Laboratorio de Ingeniería Metabólica. Centro de Bioplantas. Universidad de Ciego de Ávila, Ciego de Ávila, CP 69450, Cuba. 1 E-mail: mhernandez@bioplantas.cu . Laboratorio de Mejoramiento Genético. Centro de Bioplantas. Universidad de Ciego de Ávila, Ciego de Ávila, CP 69450, Cuba. 2 LIPROVE, Departamento de Ciencias Biológicas, Facultad de Ciencias Exactas, Universidad Nacional de la Plata. La Plata. 3 Argentina. Departamento de Bioquímica y Biología Molecular. Universidad de Córdoba. Córdoba. Campus de Rabanales, Edificio Severo 4 Ochoa (C6), 14071 Córdoba, España. Biotechnology has become an important tool to produce proteases. Bromeliaceae family plants usually contain high concentration of thiol proteases. Although pineapple plants have been found to produce proteases, most of the biotechnological investigations on this crop have been focused on propagation. Plant tissue culture techniques have provided many solutions to basics questions and practical problems in plant biology. Therefore, considerable attention has been focused on the possibility of applying efficient plant tissue culture methods to physiologically active enzymes isolation. We decided to modify pre-elongation phase during pineapple micropropagation in temporary immersion bioreactors (TIB), looking for an increase of protease excretion. Seven experiments were performed to evaluate the effects of culture duration, levels of gibberellic acid (GA), 6- benzyladenine (BA), different levels of sucrose, inorganic salts, inositol and thiamine. The following indicators were recorded: shoot fresh mass per bioreactor; and protein concentration, proteolytic activity, and specific protease activity in culture media. Specific protease activity was highest at 21 d of culture, with 90g/L of sucrose, 4.2 μmol/L of GA, 100% MS salt strength, 0.1 mg/L of thiamine. Moreover, BA and inositol produced a negative effect. Proteases expression in response to in vitro culture by 2D- electrophoresis was evaluated. We found proteolytic activity in pineapple shoots cultured in vitro . The highest specific protease activity was recorded in shoots cultured in TIB. Multiplication phase in vitro did not cause a remarkable protease production in shoots. Proteome of shoots cultured in different in vitro phase were compared. Molecular mass of some protein spots were between 21 500- 31 000 Da. This parameter was similar to those indicated for cysteine proteases from Bromeliaceae. A protease was detected in TIB culture media. Retention time in RP-HPLC and molecular mass of the major protein detected in TIB culture media showed high similarity to stem bromelain. News From France The Domestication of Pineapple: Context and Hypotheses Geo Coppens d’Eeckenbrugge, CIRAD, UMR 5175 CEFE, 1919 Route de Mende, 34293Montpellier, France geo.coppens@cirad.fr Marie-France Duval, CIRAD, UPR Multiplication Végétative, Avenue Agropolis, TA A-75/03, 34398 Montpellier, Francemarie-france.duval@cirad.fr Introduction The present economic importance of pineapple is easily justified by its unique characteristics as a fruit, which ensured its very rapid diffusion and adoption, first at the continental level in all of tropical and subtropical America, and later at the global level in all tropical areas. However, as with other major crops of Amazonian or peri-Amazonian origin (e.g., cassava), how these unique characteristics were developed in pre-Columbian times, i.e., the process of its evolution under cultivation or domestication, has been poorly investigated. The reason for this must be mostly sought in the relatively limited interest that has long prevailed for the domestication of vegetatively propagated crops, particularly fruit crops. There also has been limited interest or limited study of the prehistory of the humid tropics, because there is poor conservation of archaeological macro-remains in the region. On the other hand, the views of archaeologists on crop domestication in relation to the birth of agriculture and the evolution of human societies have evolved considerably in the last decade. The greater acceptance within the scientific community of an Amazonian cradle of plant domestication and agriculture provides a favourable context for a new evaluation of the pineapple case. Within that context, we review here the results of recent research on pineapple genetic resources and the available archaeological data in an attempt to trace the path of development of this impressive fruit. Crop domestication and its archaeological context Domestication is a form of co-evolution in which humans and their crops and animals become dependent upon each other. The domestication syndrome is a set of morphological, phenological and physiological traits that are modified by conscious or 15