RESUMO
Protein prenylation is one example of a broad class of post-translational modifications where proteins are covalently linked to various hydrophobic moieties. To globally identify and monitor levels of all prenylated proteins in a cell simultaneously, our laboratory and others have developed chemical proteomic approaches that rely on the metabolic incorporation of isoprenoid analogues bearing bio-orthogonal functionality followed by enrichment and subsequent quantitative proteomic analysis. Here, several improvements in the synthesis of the alkyne-containing isoprenoid analogue C15AlkOPP are reported to improve synthetic efficiency. Next, metabolic labeling with C15AlkOPP was optimized to obtain useful levels of metabolic incorporation of the probe in several types of primary cells. Those conditions were then used to study the prenylomes of motor neurons (ES-MNs), astrocytes (ES-As), and their embryonic stem cell progenitors (ESCs), which allowed for the identification of 54 prenylated proteins from ESCs, 50 from ES-MNs, and 84 from ES-As, representing all types of prenylation. Bioinformatic analysis revealed specific enriched pathways, including nervous system development, chemokine signaling, Rho GTPase signaling, and adhesion. Hierarchical clustering showed that most enriched pathways in all three cell types are related to GTPase activity and vesicular transport. In contrast, STRING analysis showed significant interactions in two populations that appear to be cell type dependent. The data provided herein demonstrates that robust incorporation of C15AlkOPP can be obtained in ES-MNs and related primary cells purified via magnetic-activated cell sorting allowing the identification and quantification of numerous prenylated proteins. These results suggest that metabolic labeling with C15AlkOPP should be an effective approach for investigating the role of prenylated proteins in primary cells in both normal cells and disease pathologies, including ALS.
RESUMO
Protein prenylation is one example of a broad class of post-translational modifications where proteins are covalently linked to various hydrophobic moieties. To globally identify and monitor levels of all prenylated proteins in a cell simultaneously, our laboratory and others have developed chemical proteomic approaches that rely on the metabolic incorporation of isoprenoid analogues bearing bio-orthogonal functionality followed by enrichment and subsequent quantitative proteomic analysis. Here, several improvements in the synthesis of the alkyne-containing isoprenoid analogue C15AlkOPP are reported to improve synthetic efficiency. Next, metabolic labeling with C15AlkOPP was optimized to obtain useful levels of metabolic incorporation of the probe in several types of primary cells. Those conditions were then used to study the prenylomes of motor neurons (ES-MNs), astrocytes (ES-As), and their embryonic stem cell progenitors (ESCs), which allowed for the identification of 54 prenylated proteins from ESCs, 50 from ES-MNs and 84 from ES-As, representing all types of prenylation. Bioinformatic analysis revealed specific enriched pathways, including nervous system development, chemokine signaling, Rho GTPase signaling, and adhesion. Hierarchical clustering showed that most enriched pathways in all three cell types are related to GTPase activity and vesicular transport. In contrast, STRING analysis showed significant interactions in two populations that appear to be cell type dependent. The data provided herein demonstrates that robust incorporation of C15AlkOPP can be obtained in ES-MNs and related primary cells purified via magnetic-activated cell sorting allowing the identification and quantification of numerous prenylated proteins. These results suggest that metabolic labeling with C15AlkOPP should be an effective approach for investigating the role of prenylated proteins in primary cells in both normal cells and disease pathologies, including ALS.
RESUMO
POEMS syndrome-characterized by polyneuropathy, organomegaly, endocrinopathy, M-protein, and skin changes-is an uncommon and complex paraneoplastic disorder encompassing a diverse array of symptoms. Here we report the challenging case of a 34-year-old female who sought medical attention at the emergency department due to distal lower limb weakness. She was breastfeeding her first child at that time. Her condition rapidly deteriorated, making it difficult for her to perform simple tasks independently. Initially, she struggled with activities like jumping or climbing stairs. Eventually, her ability to walk was also compromised. These symptoms underscored the swift evolution of her polyneuropathy. Nerve conduction studies and electromyography confirmed a diagnosis of mixed demyelinating and axonal polyneuropathy. Subsequent investigations, including bone marrow biopsy and immunochemistry testing, revealed a plasma cell disorder characterized by lambda monoclonal gammopathy, along with elevated levels of vascular endothelial growth factor (VEGF > 8000 pg/mL). This pivotal finding led to the diagnosis of POEMS syndrome, prompting the initiation of antineoplastic therapy (daratumumab-lenalidomide-dexamethasone) to manage this condition. An autologous cell transplantation was planned. The rarity of POEMS syndrome and its diverse clinical manifestations often lead to an incorrect or delayed diagnosis. Our case underscores the importance of considering this syndrome in patients presenting with acute or subacute polyneuropathy, even if the patients are young. In conclusion, this case elucidates the diagnostic complexities of POEMS syndrome, emphasizing the integral role of comprehensive multidisciplinary evaluations and the potential influence of increased VEGF as a diagnostic key element and possible therapeutic target.
