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The posttranslational modification of proteins critically influences many biological processes and is a key mechanism that regulates the function of the RNA-binding protein Hu antigen R (HuR), a hub in liver cancer. Here, we show that HuR is SUMOylated in the tumor sections of patients with hepatocellular carcinoma in contrast to the surrounding tissue, as well as in human cell line and mouse models of the disease. SUMOylation of HuR promotes major cancer hallmarks, namely proliferation and invasion, whereas the absence of HuR SUMOylation results in a senescent phenotype with dysfunctional mitochondria and endoplasmic reticulum. Mechanistically, SUMOylation induces a structural rearrangement of the RNA recognition motifs that modulates HuR binding affinity to its target RNAs, further modifying the transcriptomic profile toward hepatic tumor progression. Overall, SUMOylation constitutes a mechanism of HuR regulation that could be potentially exploited as a therapeutic strategy for liver cancer.

The origin of eukaryotic organisms involved the integration of mitochondria into the ancestor cell, with a massive gene transfer from the original proteobacterium to the host nucleus. Thus, mitochondrial performance relies on a mosaic of nuclear gene products from a variety of genomes. The concerted regulation of their synthesis is necessary for metabolic housekeeping and stress response. This governance involves crosstalk between mitochondrial, cytoplasmic, and nuclear factors. While anterograde and retrograde regulation preserve mitochondrial homeostasis, the mitochondria can modulate a wide set of nuclear genes in response to an extensive variety of conditions, whose response mechanisms often merge. In this review, we summarise how mitochondrial metabolites and proteins-encoded either in the nucleus or in the organelle-target the cell nucleus and exert different actions modulating gene expression and the chromatin state, or even causing DNA fragmentation in response to common stress conditions, such as hypoxia, oxidative stress, unfolded protein stress, and DNA damage.

The regular functioning of the nucleolus and nucleus-mitochondria crosstalk are considered unrelated processes, yet cytochrome c (Cc) migrates to the nucleus and even the nucleolus under stress conditions. Nucleolar liquid-liquid phase separation usually serves the cell as a fast, smart mechanism to control the spatial localization and trafficking of nuclear proteins. Actually, the alternative reading frame (ARF), a tumor suppressor protein sequestered by nucleophosmin (NPM) in the nucleoli, is shifted out from NPM upon DNA damage. DNA damage also triggers early translocation of respiratory Cc to nucleus before cytoplasmic caspase activation. Here, we show that Cc can bind to nucleolar NPM by triggering an extended-to-compact conformational change, driving ARF release. Such a NPM-Cc nucleolar interaction can be extended to a general mechanism for DNA damage in which the lysine-rich regions of Cc-rather than the canonical, arginine-rich stretches of membrane-less organelle components-controls the trafficking and availability of nucleolar proteins.

The origin of eukaryotic organisms involved the integration of mitochondria into the ancestor cell, with a massive gene transfer from the original proteobacterium to the host nucleus. Thus, mitochondrial performance relies on a mosaic of nuclear gene products from a variety of genomes. The concerted regulation of their synthesis is necessary for metabolic housekeeping and stress response. This governance involves crosstalk between mitochondrial, cytoplasmic, and nuclear factors. While anterograde and retrograde regulation preserve mitochondrial homeostasis, the mitochondria can modulate a wide set of nuclear genes in response to an extensive variety of conditions, whose response mechanisms often merge. In this review, we summarise how mitochondrial metabolites and proteins—encoded either in the nucleus or in the organelle—target the cell nucleus and exert different actions modulating gene expression and the chromatin state, or even causing DNA fragmentation in response to common stress conditions, such as hypoxia, oxidative stress, unfolded protein stress, and DNA damage.

Neddylation is a post-translational mechanism that adds a ubiquitin-like protein, namely neural precursor cell expressed developmentally downregulated protein 8 (NEDD8). Here, we show that neddylation in mouse liver is modulated by nutrient availability. Inhibition of neddylation in mouse liver reduces gluconeogenic capacity and the hyperglycemic actions of counter-regulatory hormones. Furthermore, people with type 2 diabetes display elevated hepatic neddylation levels. Mechanistically, fasting or caloric restriction of mice leads to neddylation of phosphoenolpyruvate carboxykinase 1 (PCK1) at three lysine residues—K278, K342, and K387. We find that mutating the three PCK1 lysines that are neddylated reduces their gluconeogenic activity rate. Molecular dynamics simulations show that neddylation of PCK1 could re-position two loops surrounding the catalytic center into an open configuration, rendering the catalytic center more accessible. Our study reveals that neddylation of PCK1 provides a finely tuned mechanism of controlling glucose metabolism by linking whole nutrient availability to metabolic homeostasis.

Obesity is associated with adipose tissue dysfunction through the differentiation and expansion of pre-adipocytes to adipocytes (hyperplasia) and/or increases in size of pre-existing adipocytes (hypertrophy). A cascade of transcriptional events coordinates the differentiation of pre-adipocytes into fully differentiated adipocytes; the process of adipogenesis. Although nicotinamide N-methyltransferase (NNMT) has been associated with obesity, how NNMT is regulated during adipogenesis, and the underlying regulatory mechanisms, remain undefined. In present study we used genetic and pharmacological approaches to elucidate the molecular signals driving NNMT activation and its role during adipogenesis. Firstly, we demonstrated that during the early phase of adipocyte differentiation NNMT is transactivated by CCAAT/Enhancer Binding Protein beta (CEBPB) in response to glucocorticoid (GC) induction. We found that Nnmt knockout, using CRISPR/Cas9 approach, impaired terminal adipogenesis by influencing the timing of cellular commitment and cell cycle exit during mitotic clonal expansion, as demonstrated by cell cycle analysis and RNA sequencing experiments. Biochemical and computational methods showed that a novel small molecule, called CC-410, stably binds to and highly specifically inhibits NNMT. CC-410 was, therefore, used to modulate protein activity during pre-adipocyte differentiation stages, demonstrating that, in line with the genetic approach, chemical inhibition of NNMT at the early stages of adipogenesis impairs terminal differentiation by deregulating the GC network. These congruent results conclusively demonstrate that NNMT is a key component of the GC-CEBP axis during the early stages of adipogenesis and could be a potential therapeutic target for both early-onset obesity and glucocorticoid-induced obesity.

Obesity is associated with adipose tissue dysfunction through the differentiation and expansion of pre-adipocytes to adipocytes (hyperplasia) and/or increases in size of pre-existing adipocytes (hypertrophy). A cascade of transcriptional events coordinates the differentiation of pre-adipocytes into fully differentiated adipocytes; the process of adipogenesis. Although nicotinamide N-methyltransferase (NNMT) has been associated with obesity, how NNMT is regulated during adipogenesis, and the underlying regulatory mechanisms, remain undefined. In present study we used genetic and pharmacological approaches to elucidate the molecular signals driving NNMT activation and its role during adipogenesis. Firstly, we demonstrated that during the early phase of adipocyte differentiation NNMT is transactivated by CCAAT/Enhancer Binding Protein beta (CEBPB) in response to glucocorticoid (GC) induction. We found that Nnmt knockout, using CRISPR/Cas9 approach, impaired terminal adipogenesis by influencing the timing of cellular commitment and cell cycle exit during mitotic clonal expansion, as demonstrated by cell cycle analysis and RNA sequencing experiments. Biochemical and computational methods showed that a novel small molecule, called CC-410, stably binds to and highly specifically inhibits NNMT. CC-410 was, therefore, used to modulate protein activity during pre-adipocyte differentiation stages, demonstrating that, in line with the genetic approach, chemical inhibition of NNMT at the early stages of adipogenesis impairs terminal differentiation by deregulating the GC network. These congruent results conclusively demonstrate that NNMT is a key component of the GC-CEBP axis during the early stages of adipogenesis and could be a potential therapeutic target for both early-onset obesity and glucocorticoid-induced obesity.

Few genome-wide association studies (GWAS) analyzing genetic regulation of morphological traits of white blood cells have been reported. We carried out a GWAS of 12 morphological traits in 869 individuals from the general population of Sardinia, Italy. These traits, included measures of cell volume, conductivity and light scatter in four white-cell populations (eosinophils, lymphocytes, monocytes, neutrophils). This analysis yielded seven statistically significant signals, four of which were novel (four novel, PRG2, P2RX3, two of CDK6). Five signals were replicated in the independent INTERVAL cohort of 11 822 individuals. The most interesting signal with large effect size on eosinophil scatter (P-value = 8.33 x 10-32, beta = -1.651, se = 0.1351) falls within the innate immunity cluster on chromosome 11, and is located in the PRG2 gene. Computational analyses revealed that a rare, Sardinian-specific PRG2:p.Ser148Pro mutation modifies PRG2 amino acid contacts and protein dynamics in a manner that could possibly explain the changes observed in eosinophil morphology. Our discoveries shed light on genetics of morphological traits. For the first time, we describe such large effect size on eosinophils morphology that is relatively frequent in Sardinian population.

The regular functioning of the nucleolus and nucleus-mitochondria crosstalk are considered unrelated processes, yet cytochrome c (Cc) migrates to the nucleus and even the nucleolus under stress conditions. Nucleolar liquid–liquid phase separation usually serves the cell as a fast, smart mechanism to control the spatial localization and trafcking of nuclear proteins. Actually, the alternative reading frame (ARF), a tumor suppressor protein sequestered by nucleophosmin (NPM) in the nucleoli, is shifted out from NPM upon DNA damage. DNA damage also triggers early translocation of respiratory Cc to nucleus before cytoplasmic caspase activation. Here, we show that Cc can bind to nucleolar NPM by triggering an extended-to-compact conformational change, driving ARF release. Such a NPM–Cc nucleolar interaction can be extended to a general mechanism for DNA damage in which the lysine-rich regions of Cc—rather than the canonical, arginine-rich stretches of membrane-less organelle components—controls the trafcking and availability of nucleolar proteins.

t has been recently shown that electron transfer between mitochondrial cytochrome c and the cytochrome c1 subunit of the cytochrome bc1 can proceed at long-distance through the aqueous solution. Cytochrome c is thought to adjust its activity by changing the affinity for its partners via Tyr48 phosphorylation, but it is unknown how it impacts the nanoscopic environment, interaction forces, and long-range electron transfer. Here, we constrain the orientation and separation between cytochrome c1 and cytochrome c or the phosphomimetic Y48pCMF cytochrome c, and deploy an array of single-molecule, bulk, and computational methods to investigate the molecular mechanism of electron transfer regulation by cytochrome c phosphorylation. We demonstrate that phosphorylation impairs long-range electron transfer, shortens the long-distance charge conduit between the partners, strengthens their interaction, and departs it from equilibrium. These results unveil a nanoscopic view of the interaction between redox protein partners in electron transport chains and its mechanisms of regulation.

Despite mitochondria being key for the control of cell homeostasis and fate, their role in DNA damage response is usually just regarded as an apoptotic trigger. However, growing evidence points to mitochondrial factors modulating nuclear functions. Remarkably, after DNA damage, cytochrome c (Cc) interacts in the cell nucleus with a variety of well-known histone chaperones, whose activity is competitively inhibited by the haem protein. As nuclear Cc inhibits the nucleosome assembly/disassembly activity of histone chaperones, it might indeed affect chromatin dynamics and histone deposition on DNA. Several histone chaperones actually interact with Cc Lys residues through their acidic regions, which are also involved in heterotypic interactions leading to liquid–liquid phase transitions responsible for the assembly of nuclear condensates, including heterochromatin. This relies on dynamic histone–DNA interactions that can be modulated by acetylation of specific histone Lys residues. Thus, Cc may have a major regulatory role in DNA repair by fine-tuning nucleosome assembly activity and likely nuclear condensate formation.

Cell signaling mechanisms modulate gene expression in response to internal and external stimuli. Cellular adaptation requires a precise and coordinated regulation of the transcription and translation processes. The post-transcriptional control of mRNA metabolism is mediated by the so-called RNA-binding proteins (RBPs), which assemble with specific transcripts forming messenger ribonucleoprotein particles of highly dynamic composition. RBPs constitute a class of trans-acting regulatory proteins with affinity for certain consensus elements present in mRNA molecules. However, these regulators are subjected to post-translational modifications (PTMs) that constantly adjust their activity to maintain cell homeostasis. PTMs can dramatically change the subcellular localization, the binding affinity for RNA and protein partners, and the turnover rate of RBPs. Moreover, the ability of many RBPs to undergo phase transition and/or their recruitment to previously formed membrane-less organelles, such as stress granules, is also regulated by specific PTMs. Interestingly, the dysregulation of PTMs in RBPs has been associated with the pathophysiology of many different diseases. Abnormal PTM patterns can lead to the distortion of the physiological role of RBPs due to mislocalization, loss or gain of function, and/or accelerated or disrupted degradation. This Mini Review offers a broad overview of the post-translational regulation of selected RBPs and the involvement of their dysregulation in neurodegenerative disorders, cancer and other pathologies.

Cell signaling mechanisms modulate gene expression in response to internal and external stimuli. Cellular adaptation requires a precise and coordinated regulation of the transcription and translation processes. The post-transcriptional control of mRNA metabolism is mediated by the so-called RNA-binding proteins (RBPs), which assemble with specific transcripts forming messenger ribonucleoprotein particles of highly dynamic composition. RBPs constitute a class of trans-acting regulatory proteins with affinity for certain consensus elements present in mRNA molecules. However, these regulators are subjected to post-translational modifications (PTMs) that constantly adjust their activity to maintain cell homeostasis. PTMs can dramatically change the subcellular localization, the binding affinity for RNA and protein partners, and the turnover rate of RBPs. Moreover, the ability of many RBPs to undergo phase transition and/or their recruitment to previously formed membrane-less organelles, such as stress granules, is also regulated by specific PTMs. Interestingly, the dysregulation of PTMs in RBPs has been associated with the pathophysiology of many different diseases. Abnormal PTM patterns can lead to the distortion of the physiological role of RBPs due to mislocalization, loss or gain of function, and/or accelerated or disrupted degradation. This Mini Review offers a broad overview of the post-translational regulation of selected RBPs and the involvement of their dysregulation in neurodegenerative disorders, cancer and other pathologies