Cell-specific gene and protein expression in a orchid mycorrhizal interaction

• Grant number: 12/06623-7
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate
• Start date: November 01, 2012
• End date: June 30, 2013
• Field of knowledge: Biological Sciences - Genetics - Molecular Genetics and Genetics of Microorganisms
• Principal Investigator: Marcio Rodrigues Lambais
• Grantee: Rafael Borges da Silva Valadares
• Supervisor: Silvia Perotto
• Host Institution: Escola Superior de Agricultura Luiz de Queiroz (ESALQ). Universidade de São Paulo (USP). Piracicaba , SP, Brazil
• Institution abroad: Università degli Studi di Torino (UNITO), Italy
• Associated to the scholarship: 11/01727-6 - Identification and characterization of proteins involved in the regulation and development of orchid mycorrhizas, BP.DR

Abstract

Orchids have minute seeds lacking energetic resources for germination. In nature, orchid seed germination occurs in mutualistic symbiotic association with certain mycorrhizal fungi, which transfer simple sugars to the plant tissues. All orchids are myco-heterotrophic during the first stages of development, evolving to a mixotrophic phase, when the supply of sugars is derived from fungus and photosynthesis, and finally to an autotrophic phase, when adult. The molecular mechanisms controlling the development of orchid mycorrhiza are unknown, but it is likely that proteins accumulate specifically during the symbiotic phase and their expression is regulated by the energetic demands of each developmental stage. The preliminary results obtained in our laboratory indicate that proteins related to the plant defense system, carbon metabolism, molecular signaling, hormonal regulation and redox metabolism are differentially expressed in the different stages of the symbiotic germination of Orchidaceae, but the location of these proteins in different cell types remain unclear. The aim of this project is to verify the differential regulation of genes and proteins within cells colonized and non-colonized by the orchid mycorrhizal fungi. Plantlets of Oeceoclades maculata, symbiotically germinated with Tullanella calospora, will be fixed in paraffin, sectionated and, cells of interest will be captured with a Laser Microdissection system. From these cells, RNA and protein will be isolated and sequenced using the large-scale workflows Illumina-RNAseq and tandem mass spectrometry for proteins. Bioinformatics analysis will be carried aiming to identify, quantify and distinguish plant or fungi-derived genes and proteins. It is expected that genes and proteins specifically related to symbiosis regulation will exhibit increased expression in orchid mycorrhizal cells. (AU)