Biological Chemistry最新文献

Tripterygium wilfordii has been used for a long time to treat autoimmune diseases. Its toxic side effects limit its clinical application. Mitophagy plays a protective role in various diseases. TANK-binding kinase 1 (TBK1) is a mitophagy-promoting molecule. This study aimed to investigate whether TBK1 could alleviate triptolide (TP)-induced nephrotoxicity by regulating mitophagy. To establish TP-induced nephrotoxic injury in animal model, 16 Sprague-Dawley rats were administered with TP by gavage, then renal tissues were collected for hematoxylin and eosin (HE) staining, western blotting and immunofluorescence analysis. To investigate whether up-regulation of TBK1 could alleviate TP-induced nephrotoxic injury and the specific mechanism, HK-2 cells were cultured in vitro, transfected with TBK1-overexpression recombinant lentivirus, then treated with TP. Western blotting, immunofluorescence, flow cytometry, multifunctional microplate detector were used to detect the relevant molecules. Here we found that TP caused kidney function damage, declined mitophagy levels, decreased the expression of TBK1 and mitophagy-related proteins in rats. TP stimulation decreased cell viability, mitochondrial membrane potential, mitophagy-protein, the formation of mito-autophagosomes and mito-autophagolysosomes in HK-2 cells. Upregulating TBK1 could reverse these damages. In summary, TP-induced cell injury had decreased mitophagy levels. Up-regulating TBK1 could increase mitophagy and further alleviate TP-induced cell injury.

Human papillomaviruses are causative agents in around 5 % of all cancers, and in a number of other human diseases. While prophylactic vaccines will alleviate the HPV disease burden on future generations, there are currently no therapeutic anti-viral strategies for combating HPV infections or lesions. HPV induce the proliferation of infected epithelial cells and modulate the host differentiation response, and both of these controls are required for a successful viral life cycle. Enhanced understanding of viral-host interactions during the viral life cycle will identify potential novel anti-viral strategies for therapeutic development. This minireview will summarize the critical role of the host enzyme CK2 in regulating the function of the viral proteins E1, E2 and E7; such control makes CK2 a critical enzyme for regulating HPV life cycles. Therapeutic strategies blocking CK2 function to combat HPV infections and treat HPV diseases will be described.

Poirier-Bienvenu neurodevelopmental syndrome is a neurodevelopmental disorder associated with de novo variants of the CSNK2B gene, characterized by intellectual disability, developmental delay, frequent seizures and more. While the majority of variants are nonsense variants leading to abortion of protein translation and no or truncated CK2β, many pathogenic missense variants also exist. We investigated the effect of four variants on CK2 holoenzyme formation and activity. We show that variants in the Zinc-finger region leads to reduced protein stability and altered subcellular localization. The instability is partly mediated by proteasomal and lysosomal degradation. We further show that homodimerization of these CK2β variants (p.Arg111Pro, p.Cys137Phe), localized within the Zinc-finger domain, is significantly reduced, while CK2α binding appears not affected. Other variants, p.Asp32Asn and p.Arg86Cys, did not affect stability or CK2β/α binding. For these mutants, the key to understanding the pathological mechanism may depend on external factors, such as altered protein-protein interaction. We conclude that Zinc-finger domain variants appear to destabilize the protein and affect holoenzyme formation, effectively reducing the pool of competent holoCK2. In the context of POBINDS, our findings suggest that Zinc-finger domain variants are likely to affect cells similarly to truncating and splicing variants with reduced translation of full-length CK2β.

Proteome-wide scale in a dose- and time-depending setting is crucial to fully understand the pharmacological mechanism of anticancer drugs as well as identification of candidates for drug response biomarkers. Here, we investigated the effect of the CIGB-300 anticancer peptide at IC₅₀ and IC₈₀ doses during 1 and 4 h of treatment on the squamous lung cancer cell (NCI-H226) proteome. An overwhelming dose-dependent inhibitory effect with minor up-regulated proteins was observed by increasing CIGB-300 dose level. Functional enrichment was also CIGB-300 dose-dependent with common or exclusively regulated proteins in each dose and time settings. A protein core involving small molecule biosynthesis, aldehyde metabolism and metabolism of nucleobases was regulated irrespectively to the dose or the treatment time. Importantly, a group of proteins linked to NSCLC tumor biology, poor clinical outcome and some Protein Kinase CK2 substrates, were significantly regulated by treating with both CIGB-300 doses. Likewise, we observed a consistent downregulation of different proteins that had been already reported to be inhibited by CIGB-300 in lung adenocarcinoma and acute myeloid leukemia. Overall, our proteomics-guided strategy based on time and drug dose served to uncover novel clues supporting the CIGB-300 cytotoxic effect and also to identify putative pharmacodynamic biomarkers in NSCLC.

Protein kinase CK2 (casein kinase 2) mainly exists as heterotetrameric holoenzyme with two catalytic subunits (CK2α or CK2α') bound to a homodimer of non-catalytic subunits (CK2β). With CSNK2A1 and CSNK2A2, the human genome contains two paralogs encoding catalytic CK2 subunits. Both gene products, called CK2α and CK2α', strongly interact with CK2β. An earlier report that CK2α' has a lower CK2β affinity than CK2α is confirmed via isothermal titration calorimetry in this study. Furthermore, we show with a fluorescence-anisotropy assay that a CK2β-competitive peptide binds less strongly to CK2α' than to CK2α. The reason for the reduced affinity of CK2α' to CK2β and CK2β competitors is puzzling: both isoenzymes have identical amino acid compositions at their CK2β interfaces, but the β4β5 loop, a component of this interface, is conformationally less adaptable in CK2α' than in CK2α due to intramolecular constraints. To release these constraints, we constructed a CK2α' mutant that was equalized to CK2α at the backside of the β4β5 loop. Concerning thermostability, affinity to CK2β or CK2β competitors and 3D-structure next to the β4β5 loop, this CK2α' mutant is more similar to CK2α than to its own wild-type, suggesting a critical role of the β4β5 loop adaptability for CK2β affinity.

Protein kinase CK2 is a promising therapeutic target, especially in oncology. Over the years, various inhibitors have been developed, with polyhalogenated scaffolds emerging as a particularly effective class. Halogens like bromine and chlorine enhance inhibitor stability by forming additional interactions within the ATP pocket. Among halogenated scaffolds, benzotriazole and benzimidazole have led to potent molecules such as 4,5,6,7-tetrabromo-1H-benzotriazole (IC₅₀ = 300 nM) and 4,5,6,7-tetrabromo-2-(dimethylamino)benzimidazole (IC₅₀ = 140 nM). Modifications, including 4,5,6-tribromo-7-ethyl-1H-benzotriazole (IC₅₀ = 160 nM), further improved activity. Changing scaffolds while retaining halogens has enabled design of new inhibitors. Flavonols, dibenzofuranones, and the indeno[1,2-b]indole scaffold are key examples. Halogenation of the reference molecule 5-isopropyl-5,6,7,8-tetrahydroindeno[1,2-b]indole-9,10-dione (4b, IC₅₀ = 360 nM) significantly boosted potency. The study focused on introducing four halogens, yielding to the compound 1,2,3,4-tetrabromo-5-isopropyl-5,6,7,8-tetrahydroindeno[1,2-b]indole-9,10-dione (MC11), with an IC₅₀ of 16 nM. Co-crystallography revealed how bromine atoms enhance binding, and MC11 demonstrated strong in cellulo activity, particularly against leukemic cell lines like IPC-Bcl2.

The glycosylphosphatidylinositol (GPI)-anchored serine proteases, prostasin and testisin, have essential roles in diverse physiological functions including development, reproduction, homeostasis and barrier function of epithelia, angiogenesis, coagulation, and fibrinolysis. Important functions in pathological conditions such as cancer, kidney disease and cardiovascular disease have also been reported. In this review, we summarize current knowledge of the cellular and in vivo roles of prostasin and testisin in physiology and pathophysiology and explore the underlying molecular mechanisms. We discuss how new insights of their role in cancer and cardiovascular disease may facilitate translation into clinical settings in the future.

Dipeptide production mediated by dipeptidyl-peptidase (DPP)4, DPP5, DPP7, and DPP11 plays a crucial role in growth of Porphyromonas gingivalis, a periodontopathic asaccharolytic bacterium. Given the particular P1-position specificity of DPPs, it has been speculated that DPP5 or DPP7 might be responsible for degrading refractory P1 amino acids, i.e., neutral (Thr, His, Gly, Ser, Gln) and hydrophilic (Asn) residues. The present results identified DPP7 as an entity that processes these residues, thus ensuring complete production of nutritional dipeptides in the bacterium. Activity enhancement by the P1' residue was observed in DPP7, as well as DPP4 and DPP5. Toward the refractory P1 residues, DPP7 uniquely hydrolyzed HX|LD-MCA (X = His, Gln, or Asn) and their hydrolysis was most significantly suppressed in dpp7 gene-disrupted cells. Additionally, hydrophobic P2 residue significantly enhanced DPP7 activity toward these substrates. The findings propose a comprehensive 20 P1 × 20 P2 amino acid matrix showing the coordination of four DPPs to achieve complete dipeptide production along with subsidiary peptidases. The present finding of a broad substrate specificity that DPP7 accounts for releasing 48 % (192/400) of N-terminal dipeptides could implicate its potential role in linking periodontopathic disease to related systemic disorders.

In the early days of CK2 research, it was already published that the affinity of CK2 for its substrate casein was affected by insulin. Subsequent to the discovery of inhibitors of CK2 kinase activity, it was shown that CK2 has an influence on hormones that regulate glucose homeostasis and on enzymes that influence glucose metabolism in pancreatic islet cells as well as in hormone-sensitive target cells. This regulation includes the influence on transcription factors and thereby, gene expression, as well as direct modulation of the catalytic activity. The used CK2 inhibitors, especially the older ones, show a broad range of specificity, selectivity and off-target effects. Recently applied methods to down-regulate the expression of individual CK2 subunits using siRNA or CRISPR/Cas9 technology have contributed to the improvement of specificity. It was shown that inhibition of CK2 kinase activity or knock-down or knock-out of CK2α leads to an elevated synthesis and secretion of insulin in pancreatic β-cells and a down-regulation of the synthesis and secretion of glucagon from pancreatic α-cells. In the present review CK2-dependent molecular mechanisms will be addressed which contribute to the maintenance of glucose homeostasis.