PEG Induces High Expression of the Cell Cycle Checkpoint Gene WEE1 in Embryogenic Callus of Medicago truncatula: Potential Link between Cell Cycle Checkpoint Regulation and Osmotic Stress.

Adel M Elmaghrabi,Hilary J Rogers,Sergio J Ochatt,Dennis Francis

doi:10.3389/fpls.2017.01479

Adel M Elmaghrabi, Hilary J Rogers + Show 2 more

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https://doi.org/10.3389/fpls.2017.01479

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Abstract

Polyethylene glycol (PEG) can be used to mimic osmotic stress in plant tissue cultures to study mechanisms of tolerance. The aim of this experiment was to investigate the effects of PEG (M.W. 6000) on embryogenic callus of Medicago truncatula. Leaf explants were cultured on MS medium with 2 mg L-1 NAA and 0.5 mg L-1 BAP for 5 months. Then, calli were transferred to the same medium further supplemented with 10% (w/v) 6000 PEG for 6 months in order to study physiological and putative molecular markers of water stress. There were no significant differences in growth rate of callus or mitotic index ± PEG although embryogenic potential of PEG treated callus was morphologically enhanced. Cells were rounder on PEG medium and cell size, nuclear size and endoreduplication increased in response to the PEG treatment. Significant increases in soluble sugar and proline accumulation occurred under PEG treatment compared with the control. Significantly, high MtWEE1 and MtCCS52 expression resulted from 6 months of PEG treatment with no significant differences in MtSERK1 or MtP5CS expression but down regulation of MtSOS expression. The results are consistent in showing elevated expression of a cell cycle checkpoint gene, WEE1. It is likely that the cell cycle checkpoint surveillance machinery, that would include WEE1 expression, is ameliorating the effects of the stress imposed by PEG.

Highlights

Water stress can result in reducing crop yield world-wide (Boyer, 1982; Smirnoff, 1993; Gonzalez et al, 1995) and a recent UN survey has underlined the importance of water deficit in our planet and its effects for the coming generations unless urgent measures are taken
There were no significant differences between M. truncatula leaf callus cultured on MANA medium or MANA medium supplemented with 10% w/v PEG6000 over the 6 months of culture, and the linear growth rates for control and Polyethylene glycol (PEG) treatments were 0.155 and 0.168 g month−1 respectively (Figure 1)
Qualitative observations of callus indicated that those treated with PEG were typically bright green in color and exhibited clear evidence of embryogenesis as did the controls; the controls were brown in color and the somatic embryos regenerated looked blocked at an early developmental stage (Figures 2A,B)

Summary

Introduction

Water stress can result in reducing crop yield world-wide (Boyer, 1982; Smirnoff, 1993; Gonzalez et al, 1995) and a recent UN survey has underlined the importance of water deficit in our planet and its effects for the coming generations unless urgent measures are taken. Studying a legume model species is timely and Medicago truncatula is of particular interest given its rather short life cycle and autogamy. It has a small and almost completely annotated genome (500–550 Mbp) which is publicly available (Goodstein et al, 2012), and it is more drought tolerant than other legume crops such as pea, bean, and soybean (Motan et al, 1994; Gonzalez et al, 1998; Costa França et al, 2000; Galvez et al, 2005). The assessment of physiological responses (Nunes et al, 2008) and their genetic mechanisms (Badri et al, 2011) is more limited

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