Protein-based seed germination: Conversion of proteins into carbohydrates, nitrogen compounds and energy during germination and seedling establishment of Lupinus albus

The need for resource-efficient crops is increasing due to climate change, resource scarcity and the growing demand for plant-based protein sources. In this context, white lupin (Lupinus albus) is a particularly promising crop. Due to its high protein content and ability to thrive on nutrient-poor soils thanks to its specialised root structures and symbiosis with nitrogen-fixing bacteria, this legume represents a sustainable production system with great developmental potential. However, realising this potential fully requires a detailed understanding of the molecular mechanisms that govern early seed development and nutrient utilisation. During germination, plant seeds are heterotrophic, relying mostly on their stored nutrients. L. albus is a protein crop, containing up to 40 % protein in its seeds, which act as the primary storage compound. The total seed proteome is made up of 85 % classical storage proteins, which act as reserves of carbon, nitrogen and sulfur. Our results indicated that these storage properties were maintained during germination, with the storage proteins remaining stable. Instead, lipids were used to fuel the germination. The protein fractions that were most mobilised during germination were those related to desiccation tolerance, such as LEA proteins, and storage lipids, such as oleosins. These were also highly abundant in seeds. Degradation of these proteins released large quantities of the amino acids alanine, glycine, and threonine. The catabolism of these amino acids provided the seed with pyruvate and glyoxylate. When combined with acetyl-CoA, a product of lipid β-oxidation, these intermediates were believed to fulfil a glyoxylate cycle bypass, providing the seed with the carbohydrates necessary for energy generation and biomass construction (see Chapter 2: Manuscript 1). We found that the mitochondria in the seeds of L. albus were dynamic, pre-assembled systems that contained all of the necessary respiratory complexes. During imbibition, the activation of cofactor biosynthesis, transport, and redox homeostasis enabled the immediate provision of energy. We identified NAD-dependent formate dehydrogenase as the third most abundant seed mitochondrial protein. The high abundance of this protein, in combination with those of other dehydrogenases involved in mitochondrial amino acid catabolism, suggests that seeds have specialised mitochondrial metabolism, providing energy and carbohydrate precursors while carrying out detoxification. Furthermore, our study revealed that a significant proportion of the mitochondrial proteins found in L. albus seeds were uncharacterised, with no close homologues identified in Arabidopsis. This suggests the existence of legume-specific mitochondrial adaptations (see Chapter 4: Manuscript 3). During the epigeal growth phase after germination, the cotyledons of L. albus were exposed to light and turned green. At this time, the degradation of storage proteins was intensified, leading in an increased concentration of free amino acids, particularly asparagine (see Chapter 2: Manuscript 1). The synthesis of the proteins necessary for photosynthesis was also induced, resulting in these proteins becoming the most abundant in the cotyledon proteome. Photosynthesis of the cotyledon enabled the production of light-based energy and carbohydrates, but it was less efficient than that of true leaves. Therefore, before the first true leaves have fully developed, the seedling relies on stored reserves, especially in nutrient-limiting conditions. Premature cotyledon loss triggered drastic nitrogen-saving strategies, including a reduction in protein levels, particularly those related to photosynthesis and metabolism, and a significant decrease in biomass. Once there were enough true leaves for autotrophy, the cotyledons began to senesce and the metabolism shifted to recycling and export processes (see Chapter 3: Manuscript 2). This thesis provides valuable insights into the molecular biology of protein-rich seeds. It demonstrates that the germination and seedling establishment of L. albus are characterised by the precise interplay of energy metabolism, reserve mobilisation, organ transformation, and organelle reactivation. These processes rely on the seeds stored resources.