The field of functional interactomics aims to map system-wide molecular interactions and analyze their function. This is of pivotal importance, as biological structures and functions are generally accomplished by networks of short- and long-term molecular interactions.
As a technology-driven team, our research focuses on the continuous development and optimization of interactomics tools for plant research, more specifically for isolation and analysis of protein complexes and mapping of protein interaction networks. At the core of our technology lies affinity purification coupled to mass spectrometry (AP-MS). The advantage of this approach is that interactions are mapped in situ, enabling proteome-wide investigation of interactions that occur inside plant cells.
We implement these interactomics tools to gain insight on the nutrient signaling network of carbon and nitrogen and how it relates to plant growth. Many important signaling pathways impinge on the growth and proliferation of cells in plants. However, signaling pathways involving key growth regulators such as nutrients remain to a large extent elusive in plants. To broaden our knowledge in these nutrient signaling pathways, we are studying the protein interaction network comprising the antagonistic target of rapamycin (TOR) and SNF1-related (SnRK1) kinases, two master regulators that integrate nutrient signaling with plant growth.
Nutrient signaling and Plant Growth
The TOR kinase is a central regulatory hub that translates environmental and nutritional information into permissive or restrictive growth decisions.
Although the TOR pathway is conserved across eukaryotes, plants developed unique adaptations to this pathway to cope with their autotrophic and sessile nature. Overall, compared to other eukaryotic model species, the current knowledge of TOR signaling in plants is still scarce. Only few TOR pathway components are known and no phosphoproteome or interactome screens targeted to the TOR kinase have been performed.
To fill this gap, we combined a systematic phosphoproteome screen with an extensive state-of-the-art protein complex analysis, generating for the first time a comprehensive TOR signaling network in plants. Integration of both networks significantly increased our understanding of plant TOR signaling, elucidating both evolutionarily conserved as well as novel plant-specific links, covering a broad range of biological processes such as protein and nucleotide biosynthesis, autophagy, auxin signaling, chloroplast development, lipid metabolism, and senescence.
In parallel, we are currently mapping the SnRK1 signaling network to investigate how plants cope with nutrient or energy stress.
Plant Interactomics Facility
We offer the plant research community and companies access to our protein complex purification platform.
Purifications under research service agreement are performed based on well-established protocols for complex purification from either the Arabidopsis cell suspension cultures PSB-D/L, seedlings or isolated plant tissues. We have recently been able to show that our complex purification technology performs equally well in crop species such as corn and rice. We have extensive experience with tandem affinity purification (TAP) for the isolation of stable protein complexes at high purity, whereas for more short-term interactions, we apply pull-downs using GFP-containing tags, or a more efficient protocol based on the Protein A/G moiety present in our TAP tags. More recently, we have developed Turbo-ID protocols and an approach to enrich for unstable protein interactions by means of protein cross-linking.