Light-induced ultrastructure changes of amyloplasts and effect of nitrogen fertilization on greening in potato tubers (Solanum tuberosum L.)

Highlights

• Chloroplast auto-fluorescence was evident in greening tuber under UV light.

• Formation of grana was observed in amyloplasts (9–30 μm diameter) by TEM.

• Nitrogen application increased the percentage of small granule starch (<30 μm).

• Light-induced tuber greening increased with increasing nitrogen level.

Abstract

Tuber greening causes potato quality decline and economic loss. We investigated physiological mechanisms of greening by fluorescence and transmission electron microscopy. We then explored the effect of nitrogen level (0, 210, and 315 kg N ha⁻¹) on starch granule size and tuber greening of three commercial cultivars. Results showed that chlorophyll content of tuber skin increased (5–8 times) in a quadratic manner with increasing duration of light exposure (0–7 d) in all cultivars. Light-induced greening occurred in cortical parenchyma 0–1.5 mm below the periderm, where chloroplast auto-fluorescence was evident under ultraviolet light. The greening process involved membrane lose, starch granule dissolution, and grana formation in amyloplasts, along with chloroplast development. Formation of grana lamellae was observed around amyloplasts with a diameter of 9–30 μm. Nitrogen application increased the percentage of small granule starch (<30 μm) in tuber skin and thereby promoted tuber greening under light in all cultivars. This study provides new evidence for the mechanisms of tuber greening and nitrogen management in potato.

Introduction

Potato (Solanum tuberosum L.) is an important non-cereal food crop worldwide, with annual consumption of 230 million tons, second only to rice and wheat (FAO, 2014). During post-harvest logistics and marketing processes, fresh potatoes are unavoidably exposed to light, thereby causing greening of the tuber skin (Akeley et al., 1962; Olsen et al., 2018). Potato tuber greening is a process involving light-induced transformation of amyloplasts into chloroplasts and chlorophyll accumulation in cortical parenchyma cells in the ectoderm of tubers (Anstis and Northcote, 1973; Petermann and Morris, 1985), along with a concomitant the cumulation of glycoalkaloids (Friedman and Mcdonald, 1997). Chlorophyll is not harmful in itself, but the glycoalkaloids produced via the greening process are toxic (Edwards and Cobb, 1997; Grunenfelder et al., 2006). Tuber greening negatively impacts consumer interest in purchasing potato (Mekapogu et al., 2016), and as a result is a major cause of potato quality decline (Grunenfelder, 2005) and economic loss (Dourado et al., 2019).

Many studies investigated the mechanisms of tuber greening and the associated influencing factors (Tanios et al., 2018). Microscopy was adopted to observe amyloplast–chloroplast transformation in tuber tissues of potato, including the formation of a membrane capsule between the plastid envelope and the starch granules (Zhu et al., 1984; Muraja-Fras et al., 1994; Ljubicic et al., 1998; Grunenfelder, 2005). There exist differentiation of thylakoid and accumulation of chlorophyll on the newly synthesized membrane. These studies have provided evidence for uncovering the physiological mechanisms of potato greening. Microscopic techniques can now acquire more accurate images and capture more detailed information. In particular, the use of fluorescence microscopy, transmission electron microscopy (TEM), scanning electron microscopy (SEM) can fully realize the visualization of subcellular ultrastructures such as amyloplasts and chloroplasts during greening of potato tubers.

Amyloplasts are leucoplasts that synthesize and store starch. Potato amyloplasts have different volume sizes, approximately 5–80 μm in diameter (Muraja-Fras et al., 1994; Miranda and Aguilera, 2006), whereas chloroplasts are only 5–10 μm in diameter (Trunova et al., 2003; Sun et al., 2011). Zhu et a1. (1984) reported that only small amyloplasts were transformed into chloroplasts in greening potato tubers. Muraja-Fras et al. (1994) failed to corroborate the conclusion of Zhu et a1. (1984), but allowed the possibility of faster greening in smaller amyloplasts. Therefore, the relationship between the size of amyloplasts and their transformation into chloroplasts during tuber greening needs to be further clarified.

Greening of potato tubers is influenced by many factors, including internal factors such as genotype, and external factors such as nutrient management and environmental conditions (Tanios et al., 2018). Nitrogen application promotes plant canopy development and improve tuber yield and quality in potato (Tiwari et al., 2018). However, increasing the level of nitrogen application can negatively affect some quality parameters of potato tubers (Muttucumaru et al., 2013). Braun et al. (2010) reported that the rate of tuber greening increased with increasing nitrogen application level, yet the underpinning mechanisms were unclear. Zhu et al. (2017) and Zhang et al. (2019) reported that nitrogen application altered starch granule size distribution in rice (Oryza sativa L.) and buckwheat (Fagopyrum esculentum Moench.), respectively. Since potato is a starchy crop similar to rice and buckwheat, we hypothesized that nitrogen application level could promote tuber greening through its effect on starch granule size distribution in potato.

The objective of this study was to clarify the relationship between the size of amyloplasts and their transformation into chloroplasts in greening potato tubers by fluorescence microscopy and TEM. Three nitrogen levels were applied in field conditions using three cultivars to investigate tuber greening and its effect on starch granule size.