Impact of pr-10a overexpression on the cryopreservation success of Solanum tuberosum suspension cultures

Abstract

Although many genes are supposed to be a part of plant cell tolerance mechanisms against osmotic or salt stress, their influence on tolerance towards stress during cryopreservation procedures has rarely been investigated. For instance, the overexpression of the pathogenesis-related gene 10a (pr-10a) leads to improved osmotic tolerance in a transgenic cell culture of Solanum tuberosum cv. Désirée. In this study, a cryopreservation method, consisting of osmotic pretreatment, cryoprotection with DMSO and controlled-rate freezing, was used to characterize the relation between cryopreservation success and pr-10a expression in suspension cultures of S. tuberosum wild-type cells and cells overexpressing pathogenesis-related protein 10a (Pr-10a). By varying the sorbitol concentration, thus modifying the strength of the osmotic stress during the pretreatment phase, it can be shown that the wild type can successfully be cryopreserved only in a relatively narrow range of sorbitol concentrations, while the pr-10a overexpression leads to an enhanced cryopreservation success over the whole range of applied sorbitol concentrations. Together with transcription data we show that the pr-10a overexpression causes an enhanced osmotic tolerance, which in turn leads to enhanced cryopreservability, but also indicates a role of pr-10a in signal transduction. An increased cryopreservability of the transgenic cell line occurs for pretreatments longer than 24 h. Since both genotypes, characterized by distinct baseline levels of expression, exhibited similar patterns of expression induction, the induction of pr-10a appears to be a key step in the stress signal transduction of plant cells under osmotic stress.

Impacts of pr-10a Overexpression at the Molecular and the Phenotypic Level

“…Whereas, other studies reported enhanced salt and/or osmotic tolerance due to pr-10a overexpression [3,18,19], the results in the context of pathogen attack are not that easy to interpret. In potato plants, overexpressing pr-10a did neither lead to increased resistance against Phytophthora infestans nor against potato virus X [20], whereas in the legume Medicago truncatula , silencing of PR-10 -like proteins increased the tolerance against infection with Aphanomyces euteiches [21].…”

“…Because elevated PR-10a proteins are repeatedly found in salt and osmotically stressed entire plants, as well as cell cultures [3–7], a role in stress perception or signal transduction has been postulated [7]. …”

Proteomic Analysis of Copper-Binding Proteins in Excess Copper-Stressed Roots of Two Rice (Oryza sativa L.) Varieties with Different Cu Tolerances

“…PR proteins participate in a wide range of cell functions, including cell wall rigidity, signal transduction, and antimicrobial activity. Overexpression of PR proteins in plants or cells increased tolerance to salt, osmotic [ 44 , 45 ], cold [ 46 ], heavy metal and pathogen stresses [ 47 ].…”

ATP Content and Cell Viability as Indicators for Cryostress Across the Diversity of Life

“…For cryostress experiments suspended cells were taken from the logarithmic growth phase 3 days after the last transfer. The controlled-rate cooling approach based on the method of Withers and King ( Withers and King, 1980 ) was applied using the minitest system ( Heine-Dobbernack et al, 2008 ) and the protocol described by Vaas et al (2012) . Cell suspensions were pretreated for 48 h with sorbitol (final concentrations, 0, 0.3, 0.6, and 1.2 M) then incubated in 5% (v/v) DMSO for 1.5 h, cooled at rate of -0.25°C min -1 to -40°C, and finally immersed in liquid nitrogen.…”