A role for symplastic gating in the control of the potato tuber life cycle

Abstract

The control of the potato tuber life cycle has been the subject of significant interest over many years. A number of different approaches have been adopted and data is available regarding hormonal, metabolic and gene expression changes that occur over the tuber life cycle. Despite this intense effort, no unifying model for the control of the potato tuber life cycle has emerged. We have undertaken a detailed analysis of the tuber life cycle utilising physiological, biochemical and cell-biological techniques. It has emerged that a major factor contributing to both tuber induction and dormancy break is symplastic gating which controls the allocation of resources to meristematic or vegetative tissues. Future challenges include the determination of factors regulating symplastic gating at the molecular level and the extrapolation of these findings to other systems.

Introduction

The potato represents the fourth most significant global crop¹ with worldwide production of 320 M tons in 2005 (FAO estimate). Potatoes are consumed both in the developed world, primarily as processed foods and also in the developing world where they are becoming an increasingly important crop with dramatic rises in potato production in China, India and the African continent over the last two decades.² The increasing popularity of the potato in developing countries results from a combination of its high nutritional value, ease of vegetative propagation and high yield. As potato represents the highest yielding crop per hectare it is likely to continue to increase in importance as the global population rises.³

In both the developing world and industrialised countries, there is a growing requirement to control the potato tuber life cycle in order to improve scheduling, improve consistency and uniformity of the crop, improve storage characteristics and maximise yield. For example, it is estimated that approximately 30% of stolons in field grown plants fail to tuberise and up to 30% of the crop may be lost postharvest as a result of quality issues associated with dormancy break.⁴

As a result of the global importance of the potato crop and the demonstrable benefits that would arise from improved control, significant research efforts have focused on the potato tuber life cycle. Investigations into tuber initiation have examined the role of environmental cues and associated signalling events resulting in tuber induction⁵ and further work has been undertaken regarding the changes in morphology⁶ and metabolism¹ that occur on the transition from stolon elongation to tuber formation. Significant advances have also been achieved regarding the role of hormones in the control of dormancy status⁴ which have aided applied work concerned with the search for chemical suppressors of sprouting.⁷ More recently, genomic approaches have been adopted to gain insights into the physiological and metabolic changes associated with tuber induction and dormancy release.^8,9

In our research, we attempted to gain a mechanistic understanding of the events leading to tuber formation and dormancy release by combining detailed biochemical analysis of tuber tissues at different stages of development with an analysis of pathways of assimilate unloading.^10,11 Our observations allowed the development of a novel hypothesis for control of the potato tuber life cycle which is outlined here. It is intended that such a model will provide a framework for future avenues of research that will ultimately allow the development of improved methods for potato cultivation and storage.

Tuber Induction and the Transition from Apoplastic to Symplastic Unloading

The introduction of the fluorescent dye carboxyfluorescein diacetate (CFDA) via leaf stomata allowed tracking of the pathways of assimilate unloading in developing stolons and tubers. CFDA is membrane permeable facilitating uptake into cells where endogenous diesterases cleave the compound to the membrane impermeant carboxyfluorescein (CF) trapping the dye within the symplast and allowing its use as a marker for symplastic unloading in distant sink tissues.¹² Patterns of assimilate unloading were confirmed following ¹⁴CO₂ labelling and autoradiography of sink tissues. In elongating stolons CF was confined to phloem strands and there was no evidence of unloading either along the length of the phloem pathway or in the terminal meristematic region. On the contrary, analysis of assimilate unloading by autoradiography showed even distribution of radioactivity in the stolon suggesting that during the elongation phase assimilates were unloaded via an apoplastic pathway and became evenly diluted into stolon tissues. In contrast, a diffuse pattern of CF labelling was observed in swelling stolons indicative of symplastic unloading and this was associated with more intense labelling in autoradiographs. No CF unloading was observed in the apical region and unloading of ¹⁴C-assimilates in this area was relatively weak (Fig. 1). Combined with quantitative data regarding the abundance of sugars along stolon and tuber axes, incorporation of ¹⁴CO₂ into sugars and the distribution of sucrolytic enzyme activities the data were interpreted as strong evidence for a switch from apoplastic to symplastic phloem unloading coincident with tuber initiation.¹⁰

Figure 1.

Pathways of phloem unloading in tuber tissues. (A) Non-swelling stolons from plants labelled with CFDA via leaf stomata. CF restricted to the phloem (P) in both the stolon axis and hook regions. (B) Non-swelling stolons of the same developmental stage as (A) following ¹⁴CO₂ labelling of leaves. Radioactivity distributed evenly across the stolon axis and hook. Scale bar in (B) = 1 mm for (A–C). Visibly swollen stolons from plants labelled with CFDA. CF showed distinct sub-apical unloading (arrowheads) into parenchyma tissues. In this case also note lack of dye movement into apical tissues (acropetal to dotted line). (D) Autoradiograph of swelling stolon following leaf ¹⁴CO₂ labelling shows similar pattern to (C). The apical region (acropetal to dotted line) shows less labelling than sub-apical tissues and increased labelling (arrowheads) is visible in lateral parenchyma (E) tisues. Scale bar in (D) = 1 mm for (C and D). Apical meristem on dormant tubers shows little or no CF import following tuber labelling with CFDA. In the region immediately subtending the meristem (within dotted line), dye is apparent in the phloem but has not unloaded. In comparison, dye unloading is extensive into parenchyma tissues outside this region (arrowheads). P, phloem. Scale bar = 1 mm. (F) At nine weeks post-harvest, tubers show strong transport of CF in the phloem strands subtending and in the growing buds, and extensive dye unloading into the region of the apical meristem. (arrowheads). P, phloem. Scale bar = 500 µm.

Bud Dormancy and Symplastic Isolation

Following harvest, potato tubers remain dormant for a period of up to 15 weeks even when placed in ideal growth conditions.¹³ To gain insights into the control of tuber dormancy, a comprehensive metabolic analysis of both tuber parenchyma and bud tissues during dormancy transition was undertaken.¹¹ Tuber tissues underwent metabolic shifts consistent with transition from a sink to a source organ within days. Carbohydrate metabolism shifted away from starch biosynthesis and towards sucrose generation and autoradiographs of tuber slices incubated with [¹⁴C]sucrose demonstrated a functional transition in the phloem strands from a sucrose distribution network to a sucrose accumulating network. The metabolic capacity of tuber buds did not alter throughout the transition from dormancy to active growth as determined by partitioning of [¹⁴C]glucose, however significant increases were observed in the pools of both organic acids and non-structural carbohydrates consistent with increased substrate availability. Analysis of phloem unloading using CF showed symplastic isolation of the apical bud during dormancy but significant unloading into the meristematic region in actively growing buds (Fig. 1).

The Role of Symplastic Gating in the Potato Tuber Life Cycle

Our data imply a role for symplastic gating in the control of the potato tuber life cycle. Induction of tuberisation in elongating stolons was associated with (1) a switch from apoplastic to symplastic phloem unloading in the subapical region behind the meristem and (2) a reduction in label deposition in the stolon apical meristem from ¹⁴CO₂ supplied to whole plants.¹⁰ In mature tubers, the bud meristem remained dormant despite (1) a switch from starch to sucrose synthesis in the storage parenchyma upon detachment, (2) the development of an active sucrose uptake system by the tuber phloem and (3) no detectable metabolic incompetencies in the apical bud. In developing and freshly harvested tubers, the apical bud remained symplastically isolated, however symplastic connection was re-established in growing buds.¹¹ Given these observations a scheme is presented suggesting that control of the potato tuber life cycle is in part achieved via symplastic gating (Fig. 2). In this scheme, symplastic connectivity or isolation is a key mechanism controlling meristem activity by regulating the supply of substrates and signalling compounds. The next challenge is to determine what environmental and genetic factors are involved in switching symplastic gating within the potato. Such studies should lead to genetic and agronomic tools allowing improved control of the potato tuber life cycle and ultimately improved tuber yield and reduced postharvest losses.

Figure 2.

Role of symplastic gating in the potato tuber life cycle. (A) In non-swelling stolons, phloem unloading is apoplastic both in the region of the apical meristem and in the subapical region. Assimilate unloading into the subapical parenchyma is limited and the unloading of systemic signaling compounds (e.g., hormones) can be controlled by the requirement for specific phloem located transporters. (B) In swelling stolons, induction of symplastic unloading allows photoassimilates to be unloaded into lateral parenchyma cells, resulting in changes in sugar-responsive gene expression and tuber development. Phloem-transported signaling compounds can now be directly unloaded. The dormant apical meristem remains symplastically isolated, maintaining a tightly-controlled cellular domain around the bud. (C) Within days of detachment from the mother plant, tuber phloem acquires the capacity for apoplastic sucrose accumulation (scavenging) and the ability to actively transport assimilates throughout the phloem network. The apical bud remains symplastically isolated limiting substrate availability. Signalling compounds must enter meristematic cells from the apoplastic domain via specific transporters. (D) Dormancy break is associated with symplastic connection of the bud meristem to the tuber phloem network. Solutes become freely available as symplastic unloading occurs into the bud region and signaling molecules present in the phloem are freely diffusible into the meristematic region.

Addendum to: Viola R, Pelloux J, van der Ploeg A, Gillespie T, Marquis N, Roberts AG, Hancock RD. Symplastic connection is required for bud outgrowth following dormancy in potato (Solanum tuberosum L.) tubers. Plant Cell Environ. 2007;30:973–983.