Post-translational modification of RAS proteins

Mutations of RAS genes drive cancer more frequently than any other oncogene. RAS proteins integrate signals from a wide array of receptors and initiate downstream signaling through pathways that control cellular growth. RAS proteins are fundamentally binary molecular switches in which the off/on state is determined by the binding of GDP or GTP, respectively. As such, the intrinsic and regulated nucleotide-binding and hydrolytic properties of the RAS GTPase were historically believed to account for the entirety of the regulation of RAS signaling. However, it is increasingly clear that RAS proteins are also regulated by a vast array of post-translational modifications (PTMs). The current challenge is to understand what are the functional consequences of these modifications and which are physiologically relevant. Because PTMs are catalyzed by enzymes that may offer targets for drug discovery, the study of RAS PTMs has been a high priority for RAS biologists.     

High mobility group proteins and their post-translational modifications

The high mobility group (HMG) proteins, including HMGA, HMGB and HMGN, are abundant and ubiquitous nuclear proteins that bind to DNA, nucleosome and other multi-protein complexes in a dynamic and reversible fashion to regulate DNA processing in the context of chromatin. All HMG proteins, like histone proteins, are subjected to extensive post-translational modifications (PTMs), such as lysine acetylation, arginine/lysine methylation and serine/threonine phosphorylation, to modulate their interactions with DNA and other proteins. There is a growing appreciation for the complex relationship between the PTMs of HMG proteins and their diverse biological activities. Here, we reviewed the identified covalent modifications of HMG proteins, and highlighted how these PTMs affect the functions of HMG proteins in a variety of cellular processes.     

Post‐translational modifications as key regulators of apicomplexan biology: insights from proteome‐wide studies

Parasites of the Apicomplexa phylum, such as Plasmodium spp. and Toxoplasma gondii, undergo complex life cycles involving multiple stages with distinct biology and morphologies. Post‐translational modifications (PTMs), such as phosphorylation, acetylation and glycosylation, regulate numerous cellular processes, playing a role in every aspect of cell biology. PTMs can occur on proteins at any time in their lifespan and through alterations of target protein activity, localization, protein–protein interactions, among other functions, dramatically increase proteome diversity and complexity. In addition, PTMs can be induced or removed on changes in cellular environment and state. Thus, PTMs are likely to be key regulators of developmental transitions, biology and pathogenesis of apicomplexan parasites. In this review we examine the roles of PTMs in both parasite‐specific and conserved eukaryotic processes, and the potential crosstalk between PTMs, that together regulate the intricate lives of these protozoa.     

Dual Coordination of Post Translational Modifications in Human Protein Networks

Post-translational modifications (PTMs) regulate protein activity, stability and interaction profiles and are critical for cellular functioning. Further regulation is gained through PTM interplay whereby modifications modulate the occurrence of other PTMs or act in combination. Integration of global acetylation, ubiquitination and tyrosine or serine/threonine phosphorylation datasets with protein interaction data identified hundreds of protein complexes that selectively accumulate each PTM, indicating coordinated targeting of specific molecular functions. A second layer of PTM coordination exists in these complexes, mediated by PTM integration (PTMi) spots. PTMi spots represent very dense modification patterns in disordered protein regions and showed an equally high mutation rate as functional protein domains in cancer, inferring equivocal importance for cellular functioning. Systematic PTMi spot identification highlighted more than 300 candidate proteins for combinatorial PTM regulation. This study reveals two global PTM coordination mechanisms and emphasizes dataset integration as requisite in proteomic PTM studies to better predict modification impact on cellular signaling.     

RNF4—A Paradigm for SUMOylation‐Mediated Ubiquitination

Covalent modifications by Small Ubiquitin‐like MOdifier (SUMO) and ubiquitin conjugation are now recognized as independent posttranslational modifications (PTMs) employed by cells to reversibly regulate cellular signaling. SUMOylation in particular has emerged as a crucial cellular mechanism involved in multiple pathologies, including cancers, cardiovascular diseases, immunological and neurological disorders, as well as aging. Convergence of these two PTMs result in the ubiquitination of SUMOylated proteins, adding complexity in the modulation of protein functions. The SUMO‐Targeted Ubiquitin Ligases (STUbL) mediate this process, and RNF4, the mammalian STUbL, has been at the forefront in the understanding of this phenomenon. It has been shown to play important roles in a variety of cellular events, ranging from the maintenance of genomic integrity and hence, oncogenesis, to a role in development. Recent identification of direct and indirect RNF4 targets has revealed that the SUMOylation machinery is in itself targeted by RNF4, highlighting the complex nature of the signaling circuitry tightly regulating these processes. This review will touch upon both SUMOylation and ubiquitination, and will focus on how RNF4, which is at the heart of both these PTMs, modulates cellular signaling and promotes protein degradation. Moreover, the potential of therapeutically targeting RNF4 to improve cancer treatment is also explored.     

Plant redox proteomics

In common with other aerobic organisms, plants are exposed to reactive oxygen species resulting in formation of post-translational modifications related to protein oxidoreduction (redox PTMs) that may inflict oxidative protein damage. Accumulating evidence also underscores the importance of redox PTMs in regulating enzymatic activities and controlling biological processes in plants. Notably, proteins controlling the cellular redox state, e.g. thioredoxin and glutaredoxin, appear to play dual roles to maintain oxidative stress resistance and regulate signal transduction pathways via redox PTMs. To get a comprehensive overview of these types of redox-regulated pathways there is therefore an emerging interest to monitor changes in redox PTMs on a proteome scale. Compared to some other PTMs, e.g. protein phosphorylation, redox PTMs have received less attention in plant proteome analysis, possibly due to technical challenges such as with maintaining the in vivo redox states of proteins and the lability of certain PTMs, e.g. nitrosylations, during sample preparation and mass spectrometric analysis. The present review article provides an overview of the recent developments in the emerging area of plant redox proteomics.     

Post-translational modifications in MeHg-induced neurotoxicity

Mercury (Hg) exposure remains a major public health concern due to its widespread distribution in the environment. Organic mercurials, such as MeHg, have been extensively investigated especially because of their congenital effects. In this context, studies on the molecular mechanism of MeHg-induced neurotoxicity are pivotal to the understanding of its toxic effects and the development of preventive measures. Post-translational modifications (PTMs) of proteins, such as phosphorylation, ubiquitination, and acetylation are essential for the proper function of proteins and play important roles in the regulation of cellular homeostasis. The rapid and transient nature of many PTMs allows efficient signal transduction in response to stress. This review summarizes the current knowledge of PTMs in MeHg-induced neurotoxicity, including the most commonly PTMs, as well as PTMs induced by oxidative stress and PTMs of antioxidant proteins. Though PTMs represent an important molecular mechanism for maintaining cellular homeostasis and are involved in the neurotoxic effects of MeHg, we are far from understanding the complete picture on their role, and further research is warranted to increase our knowledge of PTMs in MeHg-induced neurotoxicity.     

Current knowledge about the functional roles of phosphorylative changes of membrane proteins in normal and diseased red cells

With the advent of proteomic techniques the number of known post-translational modifications (PTMs) affecting red cell membrane proteins is rapidly growing but the understanding of their role under physiological and pathological conditions is incompletely established. The wide range of hereditary diseases affecting different red cell membrane functions and the membrane modifications induced by malaria parasite intracellular growth represent a unique opportunity to study PTMs in response to variable cellular stresses. In the present review, some of the major areas of interest in red cell membrane research have been considered as modifications of erythrocyte deformability and maintenance of the surface area, membrane transport alterations, and removal of diseased and senescent red cells. In all mentioned research areas the functional roles of PTMs are prevalently restricted to the phosphorylative changes of the more abundant membrane proteins. The insufficient information about the PTMs occurring in a large majority of the red membrane proteins and the general lack of mass spectrometry data evidence the need of new comprehensive, proteomic approaches to improve the understanding of the red cell membrane physiology.     

Lysine methylation: beyond histones

Posttranslational modifications (PTMs) of histone proteins, such as acetylation, methylation, phosphorylation, and ubiquitylation, play essential roles in regulating chromatin dynamics. Combinations of different modifications on the histone proteins, termed 'histone code' in many cases, extend the information potential of the genetic code by regulating DNA at the epigenetic level. Many PTMs occur on non-histone proteins as well as histones, regulating protein-protein interactions, stability, localization, and/or enzymatic activities of proteins involved in diverse cellular processes. Although protein phosphorylation, ubiquitylation, and acetylation have been extensively studied, only a few proteins other than histones have been reported that can be modified by lysine methylation. This review summarizes the current progress on lysine methylation of non-histone proteins, and we propose that lysine methylation, like phosphorylation and acetylation, is a common PTM that regulates proteins in diverse cellular processes.     

Influence of combinatorial histone modifications on antibody and effector protein recognition

Increasing evidence suggests that histone posttranslational modifications (PTMs) function in a combinatorial fashion to regulate the diverse activities associated with chromatin. Yet how these patterns of histone PTMs influence the adapter proteins known to bind them is poorly understood. In addition, how histone-specific antibodies are influenced by these same patterns of PTMs is largely unknown. Here we examine the binding properties of histone-specific antibodies and histone-interacting proteins using peptide arrays containing a library of combinatorially modified histone peptides. We find that modification-specific antibodies are more promiscuous in their PTM recognition than expected and are highly influenced by neighboring PTMs. Furthermore, we find that the binding of histone-interaction domains from BPTF, CHD1, and RAG2 to H3 lysine 4 trimethylation is also influenced by combinatorial PTMs. These results provide further support for the histone code hypothesis and raise specific concerns with the quality of the currently available modification-specific histone antibodies.     

Diurnal changes in concerted plant protein phosphorylation and acetylation in Arabidopsis organs and seedlings

Protein phosphorylation and acetylation are the two most abundant post‐translational modifications (PTMs) that regulate protein functions in eukaryotes. In plants, these PTMs have been investigated individually; however, their co‐occurrence and dynamics on proteins is currently unknown. Using Arabidopsis thaliana, we quantified changes in protein phosphorylation, acetylation and protein abundance in leaf rosettes, roots, flowers, siliques and seedlings at the end of day (ED) and at the end of night (EN). This identified 2549 phosphorylated and 909 acetylated proteins, of which 1724 phosphorylated and 536 acetylated proteins were also quantified for changes in PTM abundance between ED and EN. Using a sequential dual‐PTM workflow, we identified significant PTM changes and intersections in these organs and plant developmental stages. In particular, cellular process‐, pathway‐ and protein‐level analyses reveal that the phosphoproteome and acetylome predominantly intersect at the pathway‐ and cellular process‐level at ED versus EN. We found 134 proteins involved in core plant cell processes, such as light harvesting and photosynthesis, translation, metabolism and cellular transport, that were both phosphorylated and acetylated. Our results establish connections between PTM motifs, PTM catalyzing enzymes and putative substrate networks. We also identified PTM motifs for further characterization of the regulatory mechanisms that control cellular processes during the diurnal cycle in different Arabidopsis organs and seedlings. The sequential dual‐PTM analysis expands our understanding of diurnal plant cell regulation by PTMs and provides a useful resource for future analyses, while emphasizing the importance of analyzing multiple PTMs simultaneously to elucidate when, where and how they are involved in plant cell regulation.     

“Direct” and “Indirect” Effects of Histone Modifications: Modulation of Sterical Bulk as a Novel Source of Functionality

The chromatin-regulatory principles of histone post-translational modifications (PTMs) are discussed with a focus on the potential alterations in chromatin functional state due to steric and mechanical constraints imposed by bulky histone modifications such as ubiquitin and SUMO. In the classical view, PTMs operate as recruitment platforms for histone “readers,” and as determinants of chromatin array compaction. Alterations of histone charges by “small” chemical modifications (e.g., acetylation, phosphorylation) could regulate nucleosome spontaneous dynamics without globally affecting nucleosome structure. These fluctuations in nucleosome wrapping can be exploited by chromatin-processing machinery. In contrast, ubiquitin and SUMO are comparable in size to histones, and it seems logical that these PTMs could conflict with canonical nucleosome organization. An experimentally testable hypothesis that by adding sterical bulk these PTMs can robustly alter nucleosome primary structure is proposed. The model presented here stresses the diversity of mechanisms by which histone PTMs regulate chromatin dynamics, primary structure and, hence, functionality.