Phosphorylation | EPFL Graph Search

In biochemistry, phosphorylation is the attachment of a phosphate group to a molecule or an ion. This process and its inverse, dephosphorylation, are common in biology. Protein phosphorylation often activates (or deactivates) many enzymes. During respiration and photosynthesis Phosphorylation is essential to the processes of both anaerobic and aerobic respiration, which involve the production of adenosine triphosphate (ATP), the "high-energy" exchange medium in the cell. During aerobic respiration, ATP is synthesized in the mitochondrion by addition of a third phosphate group to adenosine diphosphate (ADP) in a process referred to as oxidative phosphorylation. ATP is also synthesized by substrate-level phosphorylation during glycolysis. ATP is synthesized at the expense of solar energy by photophosphorylation in the chloroplasts of plant cells. Phosphorylation of glucose Glucose metabolism Phosphorylation of sugars is often the first stage in their

Crosstalk between Drp1 phosphorylation sites during mitochondrial dynamics and metabolic adaptation

Miriam Valera Alberni

Mitochondria are highly dynamics organelles that undergo coordinated cycles of fission and fusion, referred to as mitochondrial dynamics. Mitochondrial dynamics regulate mitochondrial bioenergetics and allow cells to adapt to changing cellular stresses and physiological challenges. The balance between different GTPase enzymes dictates the shift from interconnected tubular networks to fragmented individual units. Among them, mitochondrial fission is regulated by the mitochondrial fission orchestrator, Drp1. Drp1 function is modulated by post-translational modifications, of which the phosphorylation at two specific sites have gained most attention. Drp1 phosphorylation at the S579 site results in Drp1 recruitment to mitochondria to promote fission, whereas the research on the role of the Drp1 S600 phosphorylation have yielded contradictory results on whether it promotes fission or fusion. Importantly, the crosstalk and physiological impact of these phosphorylations is poorly understood. In this project, we focused first on elucidating the interplay between Drp1 S600 and Drp1 S579 phosphorylations. We described how Drp1 S600 phosphorylation promotes S579 phosphorylation by protecting against its dephosphorylation. The activation at both S600 and S579 sites is required to, then, promote mitochondrial fragmentation. To explore the physiological relevance of these phosphorylations, we generated a Drp1 S600A knock-in (Drp1KI) mouse model. Drp1KI mice displayed enhanced lipid oxidation rates and respiratory capacity, granting improved glucose tolerance and thermogenic capacity upon high-fat feeding. Housing mice at thermoneutrality blunted these differences, suggesting a role for the brown adipose tissue in the protection of Drp1KI mice against metabolic damage. Therefore, this work unveils for the first time the crosstalk between Drp1 phosphorylation sites and their relationship to impact on mitochondrial architecture. Moreover, we demonstrate that targeting the Drp1 S600 site can grant protection against diet-induced insulin resistance, suggesting that the modulation of these phosphorylations could be used in the treatment and prevention of metabolic diseases.

EPFL2020

A New Chemoenzymatic Semisynthetic Approach Provides Insight into the Role of Phosphorylation beyond Exon1 of Huntingtin and Reveals N-Terminal Fragment Length-Dependent Distinct Mechanisms of Aggregation

Rajasekhar Kolla, Hilal Lashuel, Gopinath Pushparathinam, Andreas Reif, Jonathan Jean-Pierre Ricci, Iman Rostami

Huntington’s disease is a neurodegenerative dis- order caused by the expansion of a polyglutamine repeat (>36Q) in the N-terminal domain of the huntingtin protein (Htt), which renders the protein or fragments thereof more prone to aggregate and form inclusions. Although several Htt N-terminal fragments of different lengths have been identified within Htt inclusions, most studies on the mechanisms, sequence, and structural determinants of Htt aggregation have focused on the Httexon1 (Httex1). Herein, we investigated the aggregation properties of mutant N- terminal Htt fragments of various lengths (Htt171, Htt140, and Htt104) in comparison to mutant Httex1 (mHttex1). We also present a new chemoenzymatic semisynthetic strategy that enables site-specific phosphorylation of Htt beyond Httex1. These advances yielded insights into how post-translational modifications (PTMs) and structured domains beyond Httex1 influence aggregation mechanisms, kinetics, and fibril morphology of longer N-terminal Htt fragments. We demonstrate that phosphorylation at T107 significantly slows the aggregation of mHtt171, whereas phosphorylation at T107 and S116 accelerates the aggregation, underscoring the importance of crosstalk between different PTMs. The mHtt171 proteins aggregate via a different mechanism and form oligomers and fibrillar aggregates with morphological properties that are distinct from that of mHttex1. These observations suggest that different N-terminal fragments could have distinct aggregation mechanisms and that a single polyQ-targeting antiaggregation strategy may not effectively inhibit the aggregation of all N-terminal Htt fragments. Finally, our results underscore the need for further studies to investigate the aggregation mechanisms of Htt fragments and how the various fragments interact with each other and influence Htt toxicity and disease progression.

2021

Sequence and structural determinants of Tau aggregation and seeding capacity: Implications for Neurofibrillary Tangle formation and Tau toxicity in Alzheimer's disease

Nadine Aït Bouziad

Despite decades of research in Alzheimerâs disease (AD), no treatment has been found to efficiently prevent or cure its progression. The microtubule binding protein Tau plays central roles in the pathogenesis of AD and is found, in the form of paired helical filaments (PHFs), as the main component of the neurofibrillary tangles, a major pathological hallmark of AD. Therefore, a better understanding of the molecular and structural determinants of Tau normal and pathological functions is crucial to elucidate the role of Tau in AD. The work presented in this thesis represents our efforts to contribute to addressing this knowledge gap by investigating the sequence, molecular and structural determinants of Tau aggregation, membrane binding, and toxicity and by exploring the interplay between these different properties. In Chapter 1, we assess the effect of manipulating the aggregation conditions on the fibril polymorphism of 4R Tau, the K18 fragment and the four individual repeat peptides within the microtubule binding domain, namely R1, R2, R3 and R4. The fibrilar structures formed by Tau, K18, R2 and R3 are diverse, polymorphic and depend upon the aggregation conditions. This work paves the way to a better understanding of molecular basis underlying Tau fibrils formation and demonstrates the importance of interplay between the different sequence motifs in Tau. In Chapter 2, we describe the discovery of a Tau-derived peptide that forms fibrils with a unique capacity to seed the aggregation of full-length Tau and templates its aggregation into highly ordered twisted fibrils that resemble native PHFs. This has tremendous potential applications in the understanding the biophysical properties of the PHFs, but also provide a powerful platform for designing Tau-specific imaging probes or the screening for small molecule modulators of fibrillization and clearance. In Chapter 3, we investigate the sequence determinants and structural consequences of the interactions of Tau with membranes, and describe for the first time the formation of stable protein/lipid complexes. Using NMR, we determined that the core of these complexes is comprised of short motifs localized in R2 and R3. We designed novel mutants that disrupt Tau interactions with lipids, thus providing powerful tools for investigating the role of membrane interactions in regulating the functions of Tau. Our findings point toward a novel form of Tau complexes that might be part of a membrane-dependent mechanism that regulates Tau oligomerization and toxicity. In Chapter 4, we explore the effect of tyrosine phosphorylation in regulating Tau properties. We report that phosphorylation at Y310, which is located within R3, results in the greatest inhibition of Tau fibril formation. We also identified two novel Tau kinases, Fes and Btk that phosphorylate Tau at positions Y310 and Y394, and developed antibodies specific for pY310 and pY394, thereby providing novel targets and tools that could lead to the discovery of novel mechanisms and pathways involved in regulating Tau functions. In addition to providing several tools to allow further understanding of Tau functions and implications in AD, our work underscores the importance of investigating the individual domains of Tau, especially R2 and R3, as structural determinant of Tau self-assembly and binding to membranes and demonstrates the critical involvement of phosphorylation of the Y310 residue in Tau biophysical and cellular properties.

EPFL2017