Reversible amyloids of pyruvate kinase couple cell metabolism and stress granule disassembly.

Cells respond to stress by blocking translation, rewiring metabolism and forming transient messenger ribonucleoprotein assemblies called stress granules (SGs). After stress release, re-establishing homeostasis and disassembling SGs requires ATP-consuming processes. However, the molecular mechanisms whereby cells restore ATP production and disassemble SGs after stress remain poorly understood. Here we show that upon stress, the ATP-producing enzyme Cdc19 forms inactive amyloids, and that their rapid re-solubilization is essential to restore ATP production and disassemble SGs in glucose-containing media. Cdc19 re-solubilization is initiated by the glycolytic metabolite fructose-1,6-bisphosphate, which directly binds Cdc19 amyloids, allowing Hsp104 and Ssa2 chaperone recruitment and aggregate re-solubilization. Fructose-1,6-bisphosphate then promotes Cdc19 tetramerization, which boosts its activity to further enhance ATP production and SG disassembly. Together, these results describe a molecular mechanism that is critical for stress recovery and directly couples cellular metabolism with SG dynamics via the regulation of reversible Cdc19 amyloids.

Loss of Paid Employment up to 4 Years after Colorectal Cancer Diagnosis-A Nationwide Register-Based Study with a Population-Based Reference Group.

Cancer survivors consider work as a key aspect of cancer survivorship while previous research indicated that cancer survivors have a higher risk of unemployment. The objectives were to assess: (1) whether colorectal cancer survivors less often have paid employment at diagnosis compared to a population-based reference group, (2) whether colorectal cancer survivors with paid work have a higher risk of loss of employment up to 4 years after diagnosis compared to a population-based reference group and (3) which colorectal cancer survivors are at highest risk of loss of paid employment. In a nationwide register-based study, persons diagnosed with colorectal cancer (N = 12,007) as registered in the Netherlands Cancer Registry, were compared on loss of paid employment with a sex and age-matched population-based reference group (N = 48,028) from Statistics Netherlands. Cox regression analyses were conducted. Colorectal cancer survivors had a higher risk of loss of paid employment (HR 1.56 [1.42, 1.71]). Within the group of colorectal cancer survivors, risk of loss of paid employment was lower for older survivors (>60 vs. 45-55) (HR 0.64 [0.51, 0.81]) and higher for those with a more advanced cancer stage (IV vs. I) (HR 1.89 [1.33, 2.70]) and those receiving radiotherapy (HR 1.37 [1.15, 1.63]). Colorectal cancer survivors at high risk of loss of paid employment may benefit from work support interventions as part of cancer survivorship.

Conservation of metabolic regulation by phosphorylation and non-covalent small-molecule interactions.

Here, we review extant observations of protein phosphorylation and small-molecule interactions in metabolism and ask which of their specific regulatory functions are conserved in Escherichia coli and Homo sapiens. While the number of phosphosites is dramatically higher in humans, the number of metabolite-protein interactions remains largely constant. Moreover, we found the regulatory logic of metabolite-protein interactions, and in many cases also the effector molecules, to be conserved. Post-translational regulation through phosphorylation does not appear to replace this regulation in human but rather seems to add additional opportunities for fine-tuning and more complex responses. The abundance of metabolite-protein interactions in metabolism, their conserved cross-species abundance, and the apparent conservation of regulatory logic across enormous phylogenetic distance demonstrate their relevance for maintaining cellular homeostasis in these ancient biological processes.

Stress-induced growth rate reduction restricts metabolic resource utilization to modulate osmo-adaptation time.

A near-constant feature of stress responses is a downregulation or arrest of the cell cycle, resulting in transient growth slowdown. To investigate the role of growth slowdown in the hyperosmotic shock response of S. cerevisiae, we perturbed the G1/S checkpoint protein Sic1 to enable osmo-stress response activation with diminished growth slowdown. We document that in this mutant, adaptation to stress is accelerated rather than delayed. This accelerated recovery of the mutant proceeds by liquidation of internal glycogen stores, which are then shunted into the osmo-shock response. Therefore, osmo-adaptation in wild-type cells is delayed because growth slowdown prevents full accessibility to cellular glycogen stores. However, faster adaptation comes at the cost of acute sensitivity to subsequent osmo-stresses. We suggest that stress-induced growth slowdown acts as an arbiter to regulate the resources devoted to osmo-shock, balancing short-term adaptation with long-term robustness.

Predicting thromboembolic complications in COVID-19 ICU patients using machine learning.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic is a challenge for intensive care units (ICU) in part due to the failure to identify risks for patients early and the inability to render an accurate prognosis. Previous reports suggest a strong association between hypercoagulability and poor outcome. Factors related to hemostasis may, therefore, serve as tools to improve the management of COVID-19 patients. AIM: The purpose of this report is to develop a model to determine whether it is possible to early identify COVID-19 patients at risk for thromboembolic complications (TCs). METHODS: We analyzed electronic health record data of 108 consecutive COVID-19 patients admitted to the adult ICU of the Erasmus University Medical Center between February 27 and May 20, 2020. By training a decision tree classifier on 66% of the available data, a model for the prediction of TCs was developed. RESULTS: The median (interquartile range) age was 62 (53-70) years and 73% were male. Forty-three patients (40%) developed a TC during their ICU stay. Mortality was higher for patients in the TCs group compared to the control group (26% vs. 8%, P=0.03). Lactate dehydrogenase, standardized bicarbonate, albumin, and leukocytes were identified by the Decision Tree classifier as the most powerful predictors for TCs 2 days before the onset of the TC, with a sensitivity of 73% and a positive likelihood ratio of 2.7 on the test dataset. CONCLUSIONS: Clinically relevant TCs frequently occur in critically ill COVID-19 patients. These can successfully be predicted using a decision tree model. Although this model could be of special importance to aid clinical decision making, its generalizability and clinical impact should be determined in a larger population. RELEVANCE FOR PATIENTS: Recently, severe TCs were observed in COVID-19 patients with progressive respiratory failure warranting ICU treatment. Timely identification of patients at risk of developing TCs is critical inasmuch as it would enable clinicians to initiate potentially salvaging therapeutic anticoagulation.

Systematic mapping of protein-metabolite interactions in central metabolism of Escherichia coli.

Metabolite binding to proteins regulates nearly all cellular processes, but our knowledge of these interactions originates primarily from empirical in vitro studies. Here, we report the first systematic study of interactions between water-soluble proteins and polar metabolites in an entire biological subnetwork. To test the depth of our current knowledge, we chose to investigate the well-characterized Escherichia coli central metabolism. Using ligand-detected NMR, we assayed 29 enzymes towards binding events with 55 intracellular metabolites. Focusing on high-confidence interactions at a false-positive rate of 5%, we detected 98 interactions, among which purine nucleotides accounted for one-third, while 50% of all metabolites did not interact with any enzyme. In contrast, only five enzymes did not exhibit any metabolite binding and some interacted with up to 11 metabolites. About 40% of the interacting metabolites were predicted to be allosteric effectors based on low chemical similarity to their target's reactants. For five of the eight tested interactions, in vitro assays confirmed novel regulatory functions, including ATP and GTP inhibition of the first pentose phosphate pathway enzyme. With 76 new candidate regulatory interactions that have not been reported previously, we essentially doubled the number of known interactions, indicating that the presently available information about protein-metabolite interactions may only be the tip of the iceberg.

Towards detecting regulatory protein-metabolite interactions.

New mapping approaches have greatly expanded our view on the cellular landscape of protein-metabolite interactions. These methods either identify proteins interacting with a selected metabolite or vice versa. By reviewing recent developments, we found that comprehensive mapping of the protein-metabolite interaction space can be achieved eventually using existing methods, amongst which proteomics techniques to assess cell wide protein property changes in response to metabolite treatment currently offer the highest potential. Since we expect major advances in mapping protein-metabolite interactions in the near future, the challenge shifts to the identification of interaction functionality, for which currently only few specialized methods are available.

Histamine-functionalized copolymer micelles as a drug delivery system in 2D and 3D models of breast cancer.

Histamine functionalized block copolymers based on poly(allyl glycidyl ether)-b-poly(ethylene oxide) (PAGE-b-PEO) were prepared with different ratios of histamine and octyl or benzyl groups using UV-initiated thiol-ene click chemistry. At neutral pH, the histamine units are uncharged and hydrophobic, while in acidic environments, such as in the endosome, lysosomes, or extracellular sites of tumours, the histamine groups are positively charged and hydrophilic. pH responsible polymer drug delivery systems is a promising route to site specific delivery of drugs and offers the potential to avoid side effects of systemic treatment. Our detailed in vitro experiments of the efficacy of drug delivery and the intracellular localization characteristics of this library of NPs in 2D and 3D cultures of breast cancer revealed that the 50% histamine-modified polymer loaded with DOX exhibited rapid accumulation in the nucleus of free DOX within 2 h. Confocal studies showed enhanced mitochondrial localization and lysosomal escape when compared to controls. From these combined studies, it was shown that by accurately tuning the structure of the initial block copolymers, the resulting self-assembled NPs can be designed to exploit histamine as an endosomal escape trigger and the octyl/benzyl units give rise to a hydrophobic core resulting in highly efficacious drug delivery systems (DDS) with control over intracellular localization. Optimization and rational control of the intracellular localization of both DDS and the parent drug can give nanomedicines a substantial increase in efficacy and should be explored in future studies.

Biochemical evaluation of virtual screening methods reveals a cell-active inhibitor of the cancer-promoting phosphatases of regenerating liver.

Computationally supported development of small molecule inhibitors has successfully been applied to protein tyrosine phosphatases in the past, revealing a number of cell-active compounds. Similar approaches have also been used to screen for small molecule inhibitors for the cancer-related phosphatases of regenerating liver (PRL) family. Still, selective and cell-active compounds are of limited availability. Since especially PRL-3 remains an attractive drug target due to its clear role in cancer metastasis, such compounds are highly demanded. In this study, we investigated various virtual screening approaches for their applicability to identify novel small molecule entities for PRL-3 as target. Biochemical evaluation of purchasable compounds revealed ligand-based approaches as well suited for this target, compared to docking-based techniques that did not perform well in this context. The best hit of this study, a 2-cyano-2-ene-ester and hence a novel chemotype targeting the PRLs, was further optimized by a structure-activity-relationship (SAR) study, leading to a low micromolar PRL inhibitor with acceptable selectivity over other protein tyrosine phosphatases. The compound is active in cells, as shown by its ability to specifically revert PRL-3 induced cell migration, and exhibits similar effects on PRL-1 and PRL-2. It is furthermore suitable for fluorescence microscopy applications, and it is commercially available. These features make it the only purchasable, cell-active and acceptably selective PRL inhibitor to date that can be used in various cellular applications.