From proteome to phenotype: Role of post-translational modifications
FROM PROTEOME TO PHENOTYPE: ROLE OF POST-TRANSLATIONAL MODIFICATIONS
By Lucas Frungillo, Postdoctoral Researcher
Post-translational protein modifications (PTMs) dramatically increase the complexity of cellular proteomics. It is now well established that PTMs are critical players in processes ranging from whole cellular metabolic reprogramming to fine-tuning responses of organisms to ever changing environmental conditions. However, many details on the molecular mechanisms underpinning PTM-mediated cellular signalling are yet to emerge.On the 11th to 13rd of December 2017, over 150 keen researchers from very diverse backgrounds gathered together at the University of Edinburgh, United Kingdom for a SEB Symposium, organised in collaboration with GARNet with the suggestive title of Protein Power. Their aim was to share and discuss trending perspectives on the roles of PTMs in the establishment of plant traits.
After translation of the genetic code, newly synthesized proteins are often subjected to modifications that have profound impacts on their biological role, a process referred to as post-translational protein modification (PTM). The pervasiveness and dynamicity of PTMs in all Kingdoms of life is suggestive of how organisms with small genomes end up with vast protein variants and complex metabolisms.
There are numerous examples of how PTMs impact protein function and stability in cells, dramatically affecting metabolism and whole organism performance. To add yet another layer of complexity to proteomics, different PTMs can cross-talk in a single protein platform, using reversibility and stability as powerful pleas1: this is a challenging research topic that fuelled discussions in the meeting. Despite the relevance of the PTMs across disciplines, fundamental questions such as ‘Why are some proteins targeted by specific PTMs whilst others harbouring similar target sites are not?’ and ‘How are non-enzymatic PTMs timely and spatially controlled in cells?’ are still to be fully understood. It was with those questions in mind, and certainly many more convolutions, that scientists faced a snow blanket - and some disruption caused by Storm Caroline - to meet last December in the beautiful and compelling capital of Scotland.
The symposium was warmly opened by the scientific co-organisers Dr Steven Spoel (University of Edinburgh) and Dr Cyril Zipfel (The Sainsbury Laboratory, Norwich, UK), who gave the audience a flavour of what was to come in the next three days. Their welcome talk was followed by an inspiring plenary talk by Dr Jesper V. Olsen (University of Copenhagen) that guided the audience through the signalling waves and networks of peptide phosphorylation and how analytical expertise and fundamental research go side-by-side in science2.
PROTEIN STABILISATION AND DESTABILISATION
Protein stability and turnover are largely controlled by interplay between different PTMs. The outcomes from a molecular battle between two PTMs, ubiquitination and SUMOylation, can dictate if a given protein is targeted for degradation or stabilised in cells. Diving into the obscurity of specificity determinants in SUMOylation, Dr Ron Hay (University of Dundee) detailed his group’s approach to identifying peptide motifs targeted by SUMOylation on a proteome-wide scale. Further in the presentation, he highlighted the importance of SUMOylation in regulating mechanisms of cell division by aiding in dynamics of protein complexes3.Another interesting aspect in the control of protein stability was brought up by Dr Steven Spoel (University of Edinburgh), who talked about his group’s research on the control of gene expression reprograming by protein ubiquitination. Steven’s group’s cross-cutting research sheds light on the long-standing conundrum of activation of gene expression by targeting transcription factors for degradation4. Further presentation by Dr Daniel J. Gibbs (University of Birmingham) highlighted the control of the beautiful process of blossoming in the spring. His findings link perception of reactive oxygen and nitrogen species availability through non-enzymatic PTMs to protein stability5 and serve as an illustrative example of how an intricate network of PTMs controls cellular signalling.
REGULATION OF PROTEIN ACTIVITY
From the numerous chemical reactions that can occur in a cell in each moment, just a few of these will actually happen. It is the selective regulation of enzymatic catalysis that governs cell behaviour and this is largely achieved by timely PTMs. In his talk, Dr Cyril Zipfel (The Sainsbury Laboratory) addressed exciting, novel features of the well-established PTM, phosphorylation. The investigation of a central node in the regulatory network that shapes growth, development and immunity in plants led Zipfel’s group to uncover an intricate phosphocode targeting a single protein, thus providing novel insights on downstream signalling events and metabolic trade-offs6.With another process, plant immunity, in the spotlight, Dr Gary Loake (University of Edinburgh) highlighted the establishment and recent advances of protein S-nitrosylation as a redox-based cornerstone of cell signalling. S-nitrosylation has been shown to impact hormonal signalling and reactive oxygen species production in cells and only now mechanisms of reversibility and specificity are beginning to emerge7. Gary went on to exemplify the role of S-nitrosylation in plant immunity by sharing his group’s novel findings that S-nitrosylation of a specific peptide suppresses expression of a repressor of immunity in a twisting scientific exercise. The diversity of PTMs was further showcased by Dr Piers Hemsley (University of Dundee), who highlighted exclusive features and mechanism of protein S-acylation in plants. Details about this PTM are just now emerging, although evidence already indicates a critical role of lipid based S-acylation in regulating protein function in cell membranes8. Although the list of proteins targeted to S-acylation is not yet extensive, Piers passionately argued that it will undoubtedly bulk up in the near future.
WORKSHOP ON PROTEOMIC ANALYSIS
To top it all off, on the final day of the meeting, students and early career scientists packed a classroom out to be guided through a set of different tools for proteomic analysis by world-class, passionate experts in the field. Technical and computational advances have aided scientists to routinely identify and quantify peptides targeted to PTMs even in complex and scarce biological samples. However, comprehensive analysis and making sense of increasingly common large amounts of data can be a daunting task. Mastering these techniques is an important step towards breakthroughs in the field, and therefore is of great interest to future scientists. Led by Dr. Alex Jones (University of Warwick), who also chaired a session on state of the art technological advances in PTM detection9, attendees had the chance to explore, view and analyse a sample dataset in a hands-on workshop. Experiences and challenges in the workday of proteomic research were then exchanged in an informal, but still very thoughtful, environment to equip researchers with valuable skills.Overall, the posters and talks presented in the Protein Power meeting illustrated the pervasiveness of PTMs in plant science. From fundamental cross-cutting research in model organisms to the development of crop technologies based on current knowledge, a wide range of subjects was covered by enthusiasts and talented attendees. Lively and fruitful discussions took place in joyful coffee breaks and social events. Established and early career researchers and students benefited from the knowledge exchange and interactions that occurred during the Protein Power meeting, and hopefully we will soon see the germination of new collaborations that were sown there!
1. Skelly MJ, Frungillo L &Spoel SH. (2016) Transcriptional regulation by complex interplay between post-translational modifications. Current Opinion in Plant Biology, 33, 126-132doi: 10.1016/j.pbi.2016.07.004
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6. Smakowska-Luzan E, Mott GA, Parys K, Stegmann M, Howton TC, Layeghifard M, Neuhold J, Lehner A, Kong J, Grünwald K, Weinberger N, Satbhai SB, Mayer D, Busch W, Madalinski M, Stolt-Bergner P,Provart NJ, Mukhtar MS, Zipfel C, Desveaux D, Guttman DS &Belkhadir Y. (2018) An extracellular network of Arabidopsis leucine-rich repeat receptor kinases. Nature, 553, 342-346 doi:10.1038/nature25184
7. Kneeshaw S, Gelineau SS, Tada Y, Loake GJ &Spoel SH. (2014) Selective protein denitrosylation activity of thioredoxin-h5 modulates plant immunity. Molecular Cell, 56, 153-162 doi: 10.1016/j.molcel.2014.08.003
8. Kumar M, Wightman R, Atanassov I, Gupta A, Hurst CH, Hemsley PA & Turner S. (2016) S-Acylation of the cellulose synthase complex is essential for its plasma membrane localization. Science, 353(6295), 166-169 doi: 10.1126/science.aaf4009
9. Christie-Oleza JA, Sousoni D, Armengaud J, Wellington EMH & Jones AM.(2015). Defining a pipeline for metaproteomic analyses. In book: Springer Protocols Handbooksdoi: 10.1007/8623_2015_130