Abstract: The extracellular matrix (ECM) has a molecular machinery composed of diverse proteins and proteoforms that combine properties of tensile strength with extensibility exhibiting growth-regulatory functions and self- and non-self-recognition. The identification of ECM proteoforms is the prerequisite towards a comprehensive understanding of biological functions accomplished by the outermost layer of the cell. Regulatory mechanisms of protein functions rely on post-translational modifications, phosphorylation in particular, affecting enzymatic activity, interaction, localization and stability. To investigate the ECM proteoforms, we have isolated the cell wall proteome and phosphoproteome of a tuberous crop, potato (Solanum tuberosum). LC-MS/MS analysis led to the identification of 38 proteins and 35 phosphoproteins of known and unknown functions. The findings may provide a better understanding of biochemical machinery and the integrated protein and phosphoprotein network of ECM for future functional studies of different developmental pathways and guidance cues in mechanosensing and integrity signaling.
Abstract: Brachypodiumdistachyon is a suitable plant model for studying temperate cereal crops, such as wheat, barley or rice, and helpful in the study of the grain cell wall. Indeed, the most abundant hemicelluloses that are in the B. distachyon cell wall of grain are (1-3)(1-4)-β-glucans and arabinoxylans, in a ratio similar to those of cereals such as barley or oat. Conversely, these cell walls contain few pectins and xyloglucans. Cell walls play an important role in grain physiology. The modifications of cell wall polysaccharides that occur during grain development and filling are key in the determination of the size and weight of the cereal grains. The mechanisms required for cell wall assembly and remodelling are poorly understood, especially in cereals. To provide a better understanding of these processes, we purified the cell wall at three developmental stages of the B. distachyon grain. The proteins were then extracted, and a quantitative and comparative LC-MS/MS analysis was performed to investigate the protein profile changes during grain development. Over 466 cell wall proteins (CWPs) were identified and classified according to their predicted functions. This work highlights the different proteome profiles that we could relate to the main phases of grain development and to the reorganization of cell wall polysaccharides that occurs during these different developmental stages. These results provide a good springboard to pursue functional validation to better understand the role of CWPs in the assembly and remodelling of the grain cell wall of cereals.
Abstract: Seed germination is considered to be one of the most critical phases in the plant life cycle, establishing the next generation of a plant species. It is an energy-demanding process that requires functioning mitochondria. One of the earliest events of seed germination is progressive development of structurally simple and metabolically quiescent promitochondria into fully active and cristae-containing mitochondria, known as mitochondrial biogenesis. This is a complex and tightly regulated process, which is accompanied by sequential and dynamic gene expression, protein synthesis, and post-translational modifications. The aim of this review is to give a comprehensive summary of seed mitochondrial proteome studies during germination of various plant model organisms. We describe different gel-based and gel-free proteomic approaches used to characterize mitochondrial proteomes of germinating seeds as well as challenges and limitations of these proteomic studies. Furthermore, the dynamic changes in the abundance of the mitochondrial proteomes of germinating seeds are illustrated, highlighting numerous mitochondrial proteins involved in respiration, tricarboxycylic acid (TCA) cycle, metabolism, import, and stress response as potentially important for seed germination. We then review seed mitochondrial protein carbonylation, phosphorylation, and S-nitrosylation as well as discuss the possible link between these post-translational modifications (PTMs) and the regulation of seed germination.
Abstract: Muscle dysfunction, characterized by a reductive remodeling of muscle fibers, is a common systemic manifestation in highly prevalent conditions such as chronic heart failure (CHF), chronic obstructive pulmonary disease (COPD), cancer cachexia, and critically ill patients. Skeletal muscle dysfunction and impaired muscle mass may predict morbidity and mortality in patients with chronic diseases, regardless of the underlying condition. High levels of oxidants may alter function and structure of key cellular molecules such as proteins, DNA, and lipids, leading to cellular injury and death. Protein oxidation including protein carbonylation was demonstrated to modify enzyme activity and DNA binding of transcription factors, while also rendering proteins more prone to proteolytic degradation. Given the relevance of protein oxidation in the pathophysiology of many chronic conditions and their comorbidities, the current review focuses on the analysis of different studies in which the biological and clinical significance of the modifications induced by reactive carbonyls on proteins have been explored so far in skeletal muscles of patients and animal models of chronic conditions such as COPD, disuse muscle atrophy, cancer cachexia, sepsis, and physiological aging. Future research will elucidate the specific impact and sites of reactive carbonyls on muscle protein content and function in human conditions.
Abstract: Hsp27—encoded by HspB1—is a member of the small heat shock proteins (sHsp, 12–43 kDa (kilodalton)) family. This protein is constitutively present in a wide variety of tissues and in many cell lines. The abundance of Hsp27 is highest in skeletal muscle, indicating a crucial role for muscle physiology. The protein identified as a beef tenderness biomarker was found at a crucial hub in a functional network involved in beef tenderness. The aim of this study was to analyze the proteins impacted by the targeted invalidation of HspB1 in the Tibialis anterior muscle of the mouse. Comparative proteomics using two-dimensional gel electrophoresis revealed 22 spots that were differentially abundant between HspB1-null mice and their controls that could be identified by mass spectrometry. Eighteen spots were more abundant in the muscle of the mutant mice, and four were less abundant. The proteins impacted by the absence of Hsp27 belonged mainly to calcium homeostasis (Srl and Calsq1), contraction (TnnT3), energy metabolism (Tpi1, Mdh1, PdhB, Ckm, Pygm, ApoA1) and the Hsp proteins family (HspA9). These data suggest a crucial role for these proteins in meat tenderization. The information gained by this study could also be helpful to predict the side effects of Hsp27 depletion in muscle development and pathologies linked to small Hsps.
Abstract: One way cancer cells can escape from targeted agents is through their ability to evade drug effects by rapidly rewiring signaling networks. Many protein classes, such as kinases and metabolic enzymes, are regulated by ATP binding and hydrolysis. We hypothesized that a system-level profiling of drug-induced alterations in ATP-binding proteomes could offer novel insights into adaptive responses. Here, we mapped global ATP-binding proteomes perturbed by two clinical MEK inhibitors, AZD6244 and MEK162, in KRAS mutant lung cancer cells as a model system harnessing a desthiobiotin-ATP probe coupled with LC-MS/MS. We observed strikingly unique ATP-binding proteome responses to MEK inhibition, which revealed heterogeneous drug-induced pathway signatures in each cell line. We also identified diverse kinome responses, indicating each cell adapts to MEK inhibition in unique ways. Despite the heterogeneity of kinome responses, decreased probe labeling of mitotic kinases and an increase of kinases linked to autophagy were identified to be common responses. Taken together, our study revealed a diversity of adaptive ATP-binding proteome and kinome responses to MEK inhibition in KRAS mutant lung cancer cells, and our study further demonstrated the utility of our approach to identify potential candidates of targetable ATP-binding enzymes involved in adaptive resistance and to develop rational drug combinations.