Assaad lab research interests
The lab’s predominant interest is in plant adaptive responses and survival strategies. As plant life is essential not only for food security but also for carbon capture and climate stability, we are currently attempting to transition from organismic, cellular and molecular levels of understanding gained with model organism research (see 1. to 3. below) to broader efforts aimed at modeling plant growth and community interactions (see 4. below). This is an interdisciplinary research effort involving collaboration between molecular cell biologists, ecologists and computational scientists, and in which key experiments are conducted on the campus’ state of the art ecotron.
1. PLANT ADAPTIVE RESPONSES, DECISION MAKING AND FUTURE CLIMATE
My predominant interest is in plant adaptive responses to multiple stress conditions and to erratic or contradictory stimuli. Adaptive responses can be broken down into four steps: sensing, signal integration, decision making processes and the execution of these decisions. While progress has been made in the field of Arabidopsis to understand signal perception and integration, little is known about decision making processes, which we refer to as allocation decisions, and their execution. To address this, we have set up a “conflict of interest” forward genetic screen to identify major players in allocation decisions. Together with reverse genetic approaches and screens for interactors, we have uncovered a three-component module. The first component of the module is a family of shaggy-like kinases (AtSKs), which mediate signal integration. The second component is the conserved TRAPPII multi-subunit tethering complex that mediates decision-making processes at the trans-Golgi-network (TGN). The third component comprises a family of Rab GTPases that are posited to execute decisions downstream of the first two components.
We are currently (1) elucidating the impact of post-translational modifications on the assembly, interactomes and function of TGN-associated TRAPP tethering complexes, (2) characterizing functional interactions between TRAPP complexes and Rab GTPases, and (3) assessing how these instruct sorting and trafficking decisions. We are identifying molecular mechanisms by which signaling at the TGN modulates sorting decisions that contribute to cell division, elongation, growth anisotropy and meristem function. Mechanistic insights gained here are laying down a foundation for understanding plant adaptive growth, allocation decisions, and resilience to future climate.
With changing climate, the cues that guide plant growth decisions have become more erratic; in some cases these cues even appear contradictory, as in the case of mild winters followed by late frosts or of drought followed by flooding. Understanding decision making in plants and how such processes respond to erratic or contradictory cues becomes imperative.
To dissect the molecular mechanisms of signal convergence at the AtSK-TRAPP-RAB module, we are using a broad range of methods.
- proteomics, with mass spectrometry at BayBioMS
- biochemistry including kinase assays
- molecular techniques
- high-resolution confocal microscopy at the CALM facility on campus
- stress physiology
- future climate simulations at TUMmesa
We have established very strong local and international collaborations with the groups of Bernhard Küster (WZW, TUM), Christina Ludwig (BayBioMS, WZW, TUM), Pascal Falter-Braun (Helmholz München), Zhiyong Wang (Carnegie; Stanford), the former Ian Moore (Oxford) and David Ehrhardt (Carnegie; Stanford).
2. ELUCIDATING THE FUNCTION OF TRAPP TETHERING COMPLEXES
My lab has been working on the TRAPPII complex since the 1990s. We have identified a number of Arabidopsis TRAPP components by mutation, based on their cytokinesis-defective phenotypes (Jaber et al., 2010; Thellmann et al., 2010; Rybak et al., 2014; Ravikumar et al., 2018). The TRAPPII tethering complex resides at the TGN, where it mediates endocytosis, exocytosis and protein sorting. In the context of cytokinesis, we developed and gathered evidence for a novel concept: namely, that transitions in membrane identity are mediated by a switch in tethering complexes . Most recently, and in collaboration with the group of the former Ian Moore (Oxford University), we have characterized the subunit composition of TRAPP complexes as well as interactions between TRAPP complexes and Rab GTPases in Arabidopsis. We are now gathering evidence for GEF function (Kalde et al., 2019) and working towards a GEF assay.
3. COORDINATION BETWEEN MEMBRANE AND MICROTUBULE DYNAMICS
We are also working towards understanding the coordination between membrane and microtubule dynamics in the context of cytokinesis. Plant cytokinesis occurs within a transient membrane compartment known as the cell plate, to which vesicles are delivered along microtubules. While membrane proteins required for cytokinesis are known, how these are coordinated with microtubule dynamics and regulated by cell cycle cues remains unclear. We have documented physical and genetic interactions between Transport Protein Particle II (TRAPPII) tethering factors and microtubule-associated proteins of the PLEIADE/AtMAP65 family (Steiner et al., 2016). These interactions do not specifically affect the recruitment of either TRAPPII or MAP65 proteins to the cell plate or midzone. Rather, and based on single versus double mutant phenotypes, it appears that they are required to coordinate cytokinesis with the nuclear division cycle (Steiner et al., 2016). Interestingly, TRAPPII-MAP65 interactions appear to be regulated by the phosphorylation status of TRAPPII components; we are currently investigating the significance of these post-translational modifications.
4. STUDYING CELL DIVISION IN CHLAMYDOMONAS AND ARABIDOPSIS
In a joint project with the Carnegie institution on Stanford Campus, our global Marie Curie fellow Frej Tulin is studying cell division in Clamydomonas and Arabidopsis, using proteomic and genetic methods, in combination with live cell microscopy. Chlammydomonas is a model system for conserved features of plant cell biology, such as the regulation of cell division. Results obtained in Chlamydomonas may inform studies in other plant species of agricultural importance, which are themselves less amenable to experimentation. Furthermore, microalgae such as Chlamydomonas are themselves emerging as potential cost-effective production platforms for value-added compounds. This is because of their simple nutritional requirements, rapid biomass accumulation, scale-up production in bioreactors and their ability to fold complex proteins (e.g. antibodies). Numerous bioactive molecules, most recently the receptor-binding domain of the SARS-CoV-2 virus, have been produced in Chlamydomonas. Moreover, a recent toxicological study concluded that consumption of Chlamydomonas present no health concerns (GRAS designation by the US FDA),making it possible to use its biomass either directly for human nutrition, or for the development of "edible" vaccines and other orally administered therapeutics. The potential for Chlamydomonas as a feedstock in biotechnology or agriculture is thus considerable, and recent years have witnessed rapid developments of diverse synthetic biology tools, enabling targeted genetic engeneering of both the nuclear and chloroplast genomes. However, yield is still limited due, in part, to a lack of understanding of factors that limited cell growth and division under different culturing conditions. Here, we aim to understand the core networks that control cell proliferation in Chlamydomonas and other green algae. The results from the project may help define biological constraints on microalgal biomass production, an important step towards realization of the biotechnical potential of this diverse group of organisms.
This project has received funding from the European Union's Horizon 2020 research and innovation programme under grant agreement No. 798198
5. CLIMATE ACTION
My group chairs a forum in which WZW students and faculty unite with industry, government and non-governmental organizations to debate existing mitigation goals and scenarios. The focus is on the agricultural and "land use change and forestry" (LULUCF) sectors. These sectors are described in seven boxes on the forum website, covering (i) nitrous oxide, (ii) consumer behaviour including meat consumption and waste reduction, (iii) grassland preservation, (iv) optimized peatland water levels and crop selection, (v) the species composition and management of our forests, (vi) optimized and cascading wood use as well as (vii) radical changes to our agricultural systems to avert crop failure, ecosystem collapse and climate breakdown.
Temperate European forests are severely impacted by climate change, and foresters as well as forest owners are at a loss as to what species combination to plant for optimized resilience to future climate. Furthermore, sapling mortality is a key factor impeding restoration efforts. Our research consortium is testing the model that community interactions in general and the understory in particular foster the emergence of late- successional tree species in a stratified approach. We are also developing a state of the art 3D digital simulation model for plant growth, which includes machine learning algorithms. This interdisciplinary effort will generate important insights and calibrations, enabling us to be more comprehensive in terms of developing hybrid models and simulations of resilient ecosystems that leverage plant and microbial community interactions.