These authors equally contributed to this work.Keywords: cystic fibrosis, CFTR, autophagy, cysteamine, epigallocatechin gallate, sweat chloride Abbreviations: BECN1/Beclin 1, autophagy-related; CF, cystic fibrosis; CFTR, cystic fibrosis transmembrane conductance regulator; CHX, cycloheximide; CSNK2, casein kinase 2; CXCL2, chemokine (C-X-C motif) ligand 2; CXCL8, chemokine (C-X-C motif) ligand 8; EGCG, epigallocatechin gallate; FEV, forced expiratory volume; PM, plasma membrane; RPD, rectal potential difference; SQSTM1, sequestosome 1; TGM2, transglutaminase 2; TNF, tumor necrosis factor.Restoration of BECN1/Beclin 1-dependent autophagy and depletion of SQSTM1/p62 by genetic manipulation or autophagy-stimulatory proteostasis regulators, such as cystamine, have positive effects on mouse models of human cystic fibrosis (CF). These measures rescue the functional expression of the most frequent pathogenic CFTR mutant, F508del, at the respiratory epithelial surface and reduce lung inflammation in Cftr F508del homozygous mice. Cysteamine, the reduced form of cystamine, is an FDA-approved drug. Here, we report that oral treatment with cysteamine greatly reduces the mortality rate and improves the phenotype of newborn mice bearing the F508del-CFTR mutation. Cysteamine was also able to increase the plasma membrane expression of the F508del-CFTR protein in nasal epithelial cells from F508del homozygous CF patients, and these effects persisted for 24 h after cysteamine withdrawal. Importantly, this cysteamine effect after washout was further sustained by the sequential administration of epigallocatechin gallate (EGCG), a green tea flavonoid, both in vivo, in mice, and in vitro, in primary epithelial cells from CF patients. In a pilot clinical trial involving 10 F508del-CFTR homozygous CF patients, the combination of cysteamine and EGCG restored BECN1, reduced SQSTM1 levels and improved CFTR function from nasal epithelial cells in vivo, correlating with a decrease of chloride concentrations in sweat, as well as with a reduction of the abundance of TNF/ TNF-alpha (tumor necrosis factor) and CXCL8 (chemokine [C-X-C motif] ligand 8) transcripts in nasal brushing and TNF and CXCL8 protein levels in the sputum. Altogether, these results suggest that optimal schedules of cysteamine plus EGCG might be used for the treatment of CF caused by the F508del-CFTR mutation.

Intestinal handling of dietary proteins usually prevents local inflammatory and immune responses and promotes oral tolerance. However, in ~ 1% of the world population, gluten proteins from wheat and related cereals trigger an HLA DQ2/8‐restricted TH1 immune and antibody response leading to celiac disease. Prior epithelial stress and innate immune activation are essential for breaking oral tolerance to the gluten component gliadin. How gliadin subverts host intestinal mucosal defenses remains elusive. Here, we show that the α‐gliadin‐derived LGQQQPFPPQQPY peptide (P31–43) inhibits the function of cystic fibrosis transmembrane conductance regulator (CFTR), an anion channel pivotal for epithelial adaptation to cell‐autonomous or environmental stress. P31–43 binds to, and reduces ATPase activity of, the nucleotide‐binding domain‐1 (NBD1) of CFTR, thus impairing CFTR function. This generates epithelial stress, tissue transglutaminase and inflammasome activation, NF‐κB nuclear translocation and IL‐15 production, that all can be prevented by potentiators of CFTR channel gating. The CFTR potentiator VX‐770 attenuates gliadin‐induced inflammation and promotes a tolerogenic response in gluten‐sensitive mice and cells from celiac patients. Our results unveil a primordial role for CFTR as a central hub orchestrating gliadin activities and identify a novel therapeutic option for celiac disease.

The term 'casein kinase' has been widely used for decades to denote protein kinases sharing the ability to readily phosphorylate casein in vitro. These fall into three main classes: two of them, later renamed as protein kinases CK1 (casein kinase 1, also known as CKI) and CK2 (also known as CKII), are pleiotropic members of the kinome functionally unrelated to casein, whereas G-CK, or genuine casein kinase, responsible for the phosphorylation of casein in the Golgi apparatus of the lactating mammary gland, has only been identified recently with Fam20C [family with sequence similarity 20C; also known as DMP-4 (dentin matrix protein-4)], a member of the four-jointed family of atypical protein kinases, being responsible for the phosphorylation of many secreted proteins. In hindsight, therefore, the term 'casein kinase' is misleading in every instance; in the case of CK1 and CK2, it is because casein is not a physiological substrate, and in the case of G-CK/Fam20C/DMP-4, it is because casein is just one out of a plethora of its targets, and a rather marginal one at that. Strikingly, casein kinases altogether, albeit representing a minimal proportion of the whole kinome, appear to be responsible for the generation of up to 40-50% of non-redundant phosphosites currently retrieved in human phosphopeptides database. In the present review, a short historical explanation will be provided accounting for the usage of the same misnomer to denote three unrelated classes of protein kinases, together with an update of our current knowledge of these pleiotropic enzymes, sharing the same misnomer while playing very distinct biological roles.

Sixteen flavonoids and related compounds have been tested for their ability to inhibit three acidophilic Ser/Thr protein kinases: the Golgi apparatus casein kinase (G-CK) recently identified with protein FAM20C, protein kinase CK1, and protein kinase CK2. While G-CK is entirely insensitive to all compounds up to 40 μM concentration, consistent with the view that it is not a member of the kinome, and CK1 is variably inhibited in an isoform-dependent manner by fisetin and luteolin, and to a lesser extent by myricetin and quercetin, CK2 is susceptible to drastic inhibition by many flavonoids, displaying with six of them IC(50) values < 1 μM. A common denominator of these compounds (myricetin, quercetin, fisetin, kaempferol, luteolin, and apigenin) is a flavone scaffold with at least two hydroxyl groups at positions 7 and 4'. Inhibition is competitive with respect to the phospho-donor substrate ATP. The crystal structure of apigenin and luteolin in complex with the catalytic subunit of Zea mays CK2 has been solved, revealing their ability to interact with both the hinge region (Val116) and the positive area near Lys68 and the conserved water W1, the two main polar ligand anchoring points in the CK2 active site. Modeling experiments account for the observation that luteolin but not apigenin inhibits also CK1. The observation that luteolin shares its pyrocatechol moiety with tyrphostin AG99 prompted us to solve also the structure of this compound in complex with CK2. AG99 was found inside the ATP pocket, consistent with its mode of inhibition competitive with respect to ATP. As in the case of luteolin, the pyrocatechol group of AG99 is critical for binding, interacting with the positive area in the deepest part of the CK2 active site.