Abstract
Key message A better understanding of androgenesis with a focus on the changes in plasma membrane fluidity and endogenous ABA content affecting embryogenesis induction in microspore suspension of B. napus.Changes in plasma membrane fluidity (MF) and ABA content associated with androgenesis induction were under the study. Both parameters were monitored in microspores of two Brassica napus L. genotypes differing in their response to androgenic induction under heat (1 day at 32 °C). MF was assessed by DPH method. ABA content was evaluated by ELISA. Heat caused microspores’ plasma membrane to become more rigid. Lower MF in microspores of ‘DH 4079’ (of high androgenic potential) seems to maintain proper cell protection and leads to efficient embryogenesis induction. Plasma membrane remodelling coincided with changes of ABA content in microspores and in the culture medium in both genotypes. ABA concentration (μM) and ABA content (fmol per 104 microspores or pmol g−1 FW) were for the first time measured in microspores. ABA concentration (μM) in microspores and in the culture medium (nM) differed significantly for the genotype and the treatment. The interaction between both variables was also significant. In general, ABA content ranged from <3.5 to 87.1 fmol per 104 microspores. The highest content of ABA was detected in ‘DH 4079’ microspores at 32 °C. Assuming a mean microspores’ radius of 10 μm, it corresponds to ABA concentration of 2.1 μM. Heat shock resulted in quantum of medium pH reduction (0.1–0.2) and increased levels of ABA in microspores and in the medium of both tested genotypes. However, heat induced increase of ABA content in microspores of non-responsive ‘Campino’ had no clear-cut impact, on androgenesis induction efficiency, which suggests a more complex mechanism of process initiation.
Highlights
Rapeseed (Brassica napus L.) microspore suspensions can be used as a convenient model system for the examination of the physiological response to various stress stimuli at the cellular level (Ferrie and Caswell 2011)
Androgenesis induction was obtained only in microspore suspension isolated from low temperature pre-treated (10 °C) donor plants which were subsequently treated with heat shock (1 day at 32 °C)
The experiments reported here have produced interesting observations on the role of abscisic acid (ABA) in cold and heat stress responses in flower buds and in isolated single cells. Both ABA concentration and plasma membrane fluidity were tested in the context of the response to heat stress, which was used as the main trigger in androgenesis induction
Summary
Rapeseed (Brassica napus L.) microspore suspensions can be used as a convenient model system for the examination of the physiological response to various stress stimuli at the cellular level (Ferrie and Caswell 2011). A poor embryogenic response limits the utility of desirable cultivars in breeding programs, where doubled haploid (DH) technology using isolated microspore culture is the fastest route to complete plant homozygosity in a single step (reviewed in Wedzony et al 2009). Heat stress usually increases membrane fluidity, some data showed increased plasma membrane rigidity under high temperature (Wahid et al 2007). Even slight changes in membrane fluidity significantly affect numerous cell functions including growth, solute transport, signal transduction, and membrane-associated enzyme activities. It has been proven that the activity of membrane-bound proteins, such as the translocators of small molecules, ion channels, receptor-associated protein kinases (Hohmann 2003), and sensor proteins (Marczak et al 2009; reviewed in Los and Murata 2004) is directly affected by heat
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