The contribution of CK2 and spleen tyrosine kinase (SYK) to CFTR trafficking and PKA-induced activity.

Background: Regular assessment of exercise capacity in CF is a useful tool to facilitate exercise prescription and to monitor disease changes, in particular with aerobic tests that measure peak exercise (VO2). Indeed, aerobic fitness, measured by peak oxygen uptake during maximal exercise, is a strong predictor of survival. Aim: To investigate whether there were any correlations between the Cardio-Pulmonary Exercise Test (CPET), the Bruce Test (BT) and the Modified Shuttle Walking Test (MSWT). Methods: PATIENTS 18 CF patients (6 male, mean age 16 years, range 11-30), mean FEV1 80% predicted and mean VC 81% predicted. MEASURES All patients performed CPET on cycloergometer. All subjects performed BT that consisted of an incremental treadmill exercise test using Bruce protocol and with measurements of VO2. All patients performed MSWT in order to assess exercise tolerance, which was expressed in meters. A specific Armband was used for 3 days by each patient. A SenseWear\uaeArmband is a complex multisensory activity monitor that includes a 2-axis accelerometer, sensor for heat flux, galvanic skin response, and skin temperature. Moreover it is able to calculate the metabolic equivalent (MET) and the number of steps/day. To evaluate physical activity expressed in METs/week, the Minnesota Leisure Time Activity Survey (MLTAS) Questionnaire, adapted form of the Minnesota Leisure Activity Survey, was used. Results:We found a significant correlation between CPET and BT (r =0,935, p<0,0001) and between CPET and MSWT (r = 0,731, p=0,0006). However there was no correlation between exercise performance indexes and pulmonary function and nutritional status parameters. Finally there was a significant correlation between CPET (expressed in VO2) and MLTAS (expressed in METs/week) (r=0,83, p<0,0001). Conclusions: Our data showed that BT and MSWT are feasible, cheap, and easy to perform. These tests may be a viable alternative to CPET for assessing exercise capacity in CF patients. Moreover, analysis of the questionnaire (MLTAS) showed that physical activity has a real physiological impact on performance index. Clinical Relevance Statement: BT and MSWT are a feasible and cheap alternative to CPET and they may be used routinely to assess exercise tolerance in CF patients. Monitoring physical activity should be a primary goal in CF care

Finger stick blood test to assess postvaccination SARS-CoV-2 neutralizing antibody response against variants

10.1002/btm2.10293BIOENGINEERING & TRANSLATIONAL MEDICINE7

Nucleoside diphosphate kinase (NDPK, NM23, AWD): recent regulatory advances in endocytosis, metastasis, psoriasis, insulin release, fetal erythroid lineage and heart failure; translational medicine exemplified

The guest editor (AM) provides his perspective on the most recent advances on nucleoside diphosphate kinase (NDPK, otherwise known as AWD or NM23) showcasing phospho-histidine biochemistry and its impact on diverse pathology when disordered. His co-author (SO) provides state-of-the-art analyses from the European institute of Bioinformatics in an appendix to support the most recent advances made by the NDPK community. Unfortunately, to those outside the field, NDPK is often dismissed as a tiny ‘ancient housekeeper’ protein found in marine sponges, social amoebae, worms, fruit flies, rodents and humans but the state-of-the-art papers overviewed here show that NDPK does not act simply in mindless rote, inter-converting cellular ‘energy currencies’. That two NDPK isoforms regulate fetal erythroid lineage is a developmental case in point. Seminal Cancer Research UK support is gratefully acknowledged that generated additional resources to enable the NDPK community to meet in Dundee in 2007 (www.dundee.ac.uk/mchs/ndpk; next meeting is planned: 2010/Mannheim-Heidelberg). The presented papers illustrate the point that when scientists are left alone ‘shut up in the narrow cell of their laboratory’ (as the philosopher Ortega once said, a sentiment echoed by Erwin Schrödinger), then progress will ultimately occur bridging the gap between specialization and translation for human benefit. To aid translation, this overview initially introduces the NDPK family to the non-specialist, who serendipitously finds these proteins in their biology. This is immediately followed by examples of the diverse biology generated by this self-aggregating group of multi-functional proteins and finally capped by an emerging idea explaining how this diversity might arise

Understanding protein kinase CK2 mis-regulation upon F508del CFTR expression

We review areas of overlap between nucleoside diphosphate kinase (NDPK; nm23) and two proteins manifesting an equivalent diversity of action, each with many thousands of publications. The first is a constitutively active protein kinase, CK2 (formerly casein kinase 2), that includes NDPK amongst its hundreds of targets. The second is an enigmatic member of the ATP-binding cassette (ABC) family of membrane pumps that normally hydrolyse ATP to transport substrates. Yet our unusual family member (ABCC7) is not a pump but, uniquely, acts as a regulated anion channel. ABCC7 is the cystic fibrosis transmembrane conductance regulator (CFTR), and we discuss the highly prevalent CFTR mutation (F508del CFTR) in terms of the uncertainties surrounding the molecular basis of cystic fibrosis that cloud approaches to corrective therapy. Using lysates from cells stably expressing either wild-type or F508del CFTR, incubated with the CK2 substrate GTP, we show that the phosphoproteome of F508del CFTR-expressing cells both differs from wild-type CFTR-expressing cells and is significantly enhanced in intensity by ∼1.5-fold (p < 0.05, paired t test with Bonferroni correction, n = 4). Phosphorylation is about 50% attenuated with a specific CK2 inhibitor. We propose that a new function may exist for the CFTR region that is commonly mutated, noting that its sequence (PGTIKENIIF508GVSYDEYRYR) is not only highly conserved within the C sub-family of ABC proteins but also a related sequence is found in NDPK. We conclude that a latent path may exist between mutation of this conserved sequence, CK2 hyperactivity and disease pathogenesis that might also explain the heterozygote advantage for the common F508del CFTR mutant

Contribution of casein kinase 2 and spleen tyrosine kinase to CFTR trafficking and protein kinase A-induced activity

Previously, the pleiotropic "master kinase" casein kinase 2 (CK2) was shown to interact with CFTR, the protein responsible for cystic fibrosis (CF). Moreover, CK2 inhibition abolished CFTR conductance in cell-attached membrane patches, native epithelial ducts, and Xenopus oocytes. CFTR possesses two CK2 phosphorylation sites (S422 and T1471), with unclear impact on its processing and trafficking. Here, we investigated the effects of mutating these CK2 sites on CFTR abundance, maturation, and degradation coupled to effects on ion channel activity and surface expression. We report that CK2 inhibition significantly decreased processing of wild-type (wt) CFTR, with no effect on F508del CFTR. Eliminating phosphorylation at S422 and T1471 revealed antagonistic roles in CFTR trafficking: S422 activation versus T1471 inhibition, as evidenced by a severe trafficking defect for the T1471D mutant. Notably, mutation of Y512, a consensus sequence for the spleen tyrosine kinase (SYK) possibly acting in a CK2 context adjacent to the common CF-causing defect F508del, had a strong effect on both maturation and CFTR currents, allowing the identification of this kinase as a novel regulator of CFTR. These results reinforce the importance of CK2 and the S422 and T1471 residues for regulation of CFTR and uncover a novel regulation of CFTR by SYK, a recognized controller of inflammation