Characterization of Snakin/GASA gene family in Solanum tuberosum and functional analysis of Snakin-1

NAHIRÑAK V; GONZALEZ DE URRETA, MARTIN; ALMASIA N I; HOPP H E; VAZQUEZ ROVERE C

Bruselas

Congreso; 19th Triennial Conference of the European Association for Potato Research; 2014

Characterization of Snakin/GASA gene family in Solanum tuberosum and functional analysis of Snakin-1Vanesa Nahirñak1, 2; Martín Gonzalez de Urreta1, 2; Natalia I. Almasia1; Esteban Hopp1 and Cecilia Vazquez Rovere1, 21Instituto de Biotecnología, CICVyA, CNIA, Instituto Nacional de Tecnología Agropecuaria (INTA), Buenos Aires, Argentina2 Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Buenos Aires, ArgentinaIntroductionSnakin/GASA proteins are widely distributed among plant species. They have been implicated in diverse biological activities: cell elongation/division, root formation, stem growth, flowering time, fruit ripening, and stress tolerance [1-4]. Interestingly, all of them maintain 12 cysteines of the C-terminus in highly conserved positions that are probably responsible for their protein structure and were shown to be essential for their biochemical activity as antioxidants [5, 6].Snakin-1 is a member of the Snakin/GASA family isolated from Solanum tuberosum that was found to be active against pathogens in vitro and we demonstrated that its overexpression in potato enhances resistance to Rhizoctonia solani and Erwinia carotovora [7, 8]. We recently showed that Snakin-1 silencing resulted in an abnormal phenotype affecting cell division, leaf primary metabolism and cell wall composition demonstrating that it has additional roles in growth and development [9]. Materials and MethodsSnakin/GASA genes were in silico screened in the Potato Genome Sequencing Consortium database using the InterPro GASA domain annotation (IPR003854) as query. For functional analysis, Snakin-1 transgenic potato lines and conventional protocols were used [9].ResultsIn this work, we carried out a genome-wide search and 14 novel Snakin/GASA genes were identified in the potato genome in addition to the previously reported genes (Snakin-1, Snakin-2 and Snakin-3). Chromosome localization studies of Snakin/GASA genes indicated that they correspond to different loci and are distributed on nine of the 12 potato chromosomes. In silico analyses revealed that Snakin/GASA gene upstream sequences carry a variety of potential hormone and stress responsive cis-regulatory elements. Expression analyses showed a tissue specific expression pattern for each family member and also some differences from the previously reported data.Further analyses of transgenic potato lines suggested that the mechanism of action possibly involves the participation of Snakin-1 in redox homeostasis.Conclusion and perspectivesSnakin/GASA family in potato consist of 17 members and phylogenetic analysis based on sequence alignments of the full-length proteins resulted in three major groups. Expression studies indicated that the spatial regulation of these novel Snakin/GASA genes is highly specific suggesting distinct functions. And as it was described for others Snakin/GASA genes, Snakin-1 may play its role by modulating reactive oxygen species. References[1] Ben-Nissan G, Lee JY, Borohov A, Weiss D (2004) GIP, a Petunia hybrida GA-induced cysteine-rich protein: a possible role in shoot elongation and transition to flowering. Plant J 37: 229-238[2] Roxrud I, Lid SE, Fletcher JC, Schmidt ED, Opsahl-Sorteberg HG (2007) GASA4, one of the 14-member Arabidopsis GASA family of small polypeptides, regulates flowering and seed development. Plant Cell Physiol 48: 471-483[3] Zhang S, Yang C, Peng J, Sun S, Wang X (2009) GASA5, a regulator of flowering time and stem growth in Arabidopsis thaliana. Plant Mol Biol 69: 745-759[4] Nahirñak V, Almasia N I, Hopp E H y Vazquez Rovere C. Snakin/GASA proteins: Involvement in hormone crosstalk and redox homeostasis. Plant Signaling & Behavior 7:8, 1-5. 2012. [5] Wigoda N, Ben-Nissan G, Granot D, Schwartz A, Weiss D (2006) The gibberellin-induced, cysteine-rich protein GIP2 from Petunia hybrida exhibits in planta antioxidant activity. Plant J 48: 796-805[6] Rubinovich L, Weiss D (2010) The Arabidopsis cysteine-rich protein GASA4 promotes GA responses and exhibits redox activity in bacteria and in planta. Plant J 64: 1018-1027[7] Segura A, Moreno M, Madueno F, Molina A, Garcia-Olmedo F (1999) Snakin-1, a peptide from potato that is active against plant pathogens. Mol Plant Microbe Interact 12: 16-23[8] Almasia NI, Bazzini AA, Hopp HE, Vazquez-Rovere C (2008) Overexpression of snakin-1 gene enhances resistance to Rhizoctonia solani and Erwinia carotovora in transgenic potato plants. Mol Plant Pathol 9: 329-338 [9] Nahirñak V, Almasia N I, Fernandez P V, Hopp E H, Estevez J M, Carrari F y Vazquez Rovere C. Potato snakin-1 gene silencing affects cell division, primary metabolism and cell wall composition. Plant Physiology. 158 (2012) 252-263.