Biology Reference
In-Depth Information
Chapter 29
Suspension-Cultured Plant Cells as a Tool to Analyze
the Extracellular Proteome
Ana B. Sabater-Jara , Lorena Almagro , Sarai Belchí-Navarro ,
María J. Martínez-Esteso , Sabry M. Youssef ,
Juan Casado-Vela , Juan C. Vera-Urbina , Susana Sellés-Marchart ,
Roque Bru-Martínez , and María A. Pedreño
Abstract
Suspension-cultured cells (SCC) are generally considered the most suitable cell systems to carry out scientifi c
studies, including the extracellular proteome (secretome). SCC are initiated by transferring friable callus
fragments into fl asks containing liquid culture medium for cell biomass growth, and they are maintained
in an orbital shaker to supply the suffi cient oxygen that allows cell growth. SCC increase rapidly during the
exponential phase and after 10-20 days (depending on the cell culture nature), the growth rate starts to
decrease due to limitation of nutrients, and to maintain for decades these kinds of cell cultures is needed
to transfer a portion of these SCC into a fresh culture medium. Despite the central role played by extracel-
lular proteins in most processes that control growth and development, the secretome has been less
well characterized than other subcellular compartments, meaning that our understanding of the cell wall
physiology is still very limited. Useful proteomic tools have emerged in recent years to unravel metabolic
network that occurs in cell walls. With the recent progress made in mass spectrometry technology, it has
become feasible to identify proteins from a given organ, tissue, cells, or even a subcellular compartment.
Compared with other methods used to isolate cell wall proteins, the spent medium of SCC provides a
convenient, continuous, and reliable and unique source of extracellular proteins. Therefore, this biological
system could be used as a large-scale cell culture from which these proteins can be secreted, easily separated
from cells without cell disruption, and so, without any cytosolic contamination, easily recovered from the
extracellular medium. This nondestructive cell wall proteome approach discloses a set of proteins that are
specifi cally expressed in the remodelling of the cell wall architecture and stress defense.
Key words
Suspension-cultured cells, Extracellular proteome, Liquid chromatography, Mass
spectrometry-based protein identifi cation, De novo sequencing, Functional annotation
1
Introduction
Plant cells can be cultivated under sterile conditions in liquid
(as suspension-cultured cells, SCC) or solid (as callus) culture
media maintaining an undifferentiated cell state and showing an
