Summary Pyruvate kinase isoform M2 (PKM2) is a glycolysis enzyme catalyzing conversion of phosphoenolpyruvate (PEP) to pyruvate with transferring a phosphate from PEP to ADP. We report here that PKM2 localizes to the cell nucleus. The levels of nuclear PKM2 correlate with cell proliferation. PKM2 activates transcription of MEK5 by phosphorylating stat3 at Y705. In vitro phosphorylation assays show that PKM2 is a protein kinase using PEP as phosphate donor. ADP competes with the protein substrate binding, indicating that the substrate may bind to the ADP site of PKM2. Our experiments suggest that PKM2 dimer is an active protein kinase, while the tetramer is an active pyruvate kinase. Expression a PKM2 mutant that exists as a dimer promotes cell proliferation, indicating that protein kinase activity of PKM2 plays a role in promoting cell proliferation. Our study reveals an important link between metabolism alteration and gene expression during tumor transformation and progression.

Ca2+ is one of the most universal and versatile signaling molecules and is involved in almost every aspect of cellular processes. Viruses are adept at utilizing the universal Ca2+ signal to create a tailored cellular environment that meets their own demands. This review summarizes most of the known mechanisms by which viruses perturb Ca2+ homeostasis and utilize Ca2+ and cellular Ca2+-binding proteins to their benefit in their replication cycles. Ca2+ plays important roles in virion structure formation, virus entry, viral gene expression, posttranslational processing of viral proteins and virion maturation and release. As part of the review, we introduce an algorithm to identify linear “EF-hand” Ca2+-binding motifs which resulted in the prediction of a total of 93 previously unrecognized Ca2+-binding motifs in virus proteins. Many of these proteins are nonstructural proteins, a class of proteins among which Ca2+ interactions had not been formerly appreciated. The presence of linear Ca2+-binding motifs in viral proteins enlarges the spectrum of Ca2+–virus interplay and expands the total scenario of viral calciomics.

The calcium-sensing receptor (CaSR) is a G-protein-coupled receptor that has an extracellular bilobed venus flytrap domain (VFTD) predicted to contain five calcium (Ca(2+))-binding sites. To elucidate the structure-function relationships of the VFTD, we investigated 294 unrelated probands with familial hypocalciuric hypercalcaemia (FHH), neonatal severe primary hyperparathyroidism (NSHPT) or autosomal dominant hypocalcaemic hypercalciuria (ADHH) for CaSR mutations and performed in vitro functional expression studies and three-dimensional modelling of mutations involving the VFTD. A total of 70 different CaSR mutations were identified: 35 in FHH, 10 in NSHPT and 25 in ADHH patients. Furthermore, a CaSR variant (Glu250Lys) was identified in FHH and ADHH probands and demonstrated to represent a functionally neutral polymorphism. NSHPT was associated with a large proportion of truncating CaSR mutations that occurred in the homozygous or compound heterozygous state. Thirty-four VFTD missense mutations were identified, and 18 mutations were located within 10 Å of one or more of the predicted Ca(2+)-binding sites, particularly at the VFTD cleft, which is the principal site of Ca(2+) binding. Mutations of residues 173 and 221, which are located at the entrance to the VFTD cleft binding site, were associated with both receptor activation (Leu173Phe and Pro221Leu) and inactivation (Leu173Pro and Pro221Gln), thereby highlighting the importance of these residues for entry and binding of Ca(2+) by the CaSR. Thus, these studies of disease-associated CaSR mutations have further elucidated the role of the VFTD cleft region in Ca(2+) binding and the function of the CaSR.

SummaryIn Arabidopsis thaliana, the etr1-2 mutation confers dominant ethylene insensitivity and results in a greater proportion of mature seeds that exhibit dormancy compared with mature seeds of the wild-type. We investigated the impact of the etr1-2 mutation on other plant hormones by analyzing the profiles of four classes of plant hormones and their metabolites by HPLC-ESI/MS/MS in mature seeds of wild-type and etr1-2 plants. Hormone metabolites were analyzed in seeds imbibed immediately under germination conditions, in seeds subjected to a 7-day moist-chilling (stratification) period, and during germination/early post-germinative growth. Higher than wild-type levels of abscisic acid (ABA) appeared to contribute, at least in part, to the greater incidence of dormancy in mature seeds of etr1-2. The lower levels of abscisic acid glucose ester (ABA-GE) in etr1-2 seeds compared with wild-type seeds under germination conditions (with and without moistchilling treatments) suggest that reduced metabolism of ABA to ABA-GE likely contributed to the accumulation of ABA during germination in the mutant. The mutant seeds exhibited generally higher auxin levels and a large build-up of indole-3-aspartate when placed in germination conditions following moistchilling. The mutant manifested increased levels of cytokinin glucosides through zeatin-O-glucosylation (Z-O-Glu). The resulting increase in Z-O-Glu was the largest and most consistent change associated with the ETR1 gene mutation. There were more gibberellins (GA) and at higher concentrations in the mutant than in wild-type. Our results suggest that ethylene signaling modulates the metabolism of all the other plant hormone pathways in seeds. Additionally, the hormone profiles of etr1-2 seed during germination suggest a requirement for higher than wild-type levels of GA to promote germination in the absence of a functional ethylene signaling pathway.

Structural and functional characterization of the extracellular domain of the human CaSR with bound Mg2+ and a tryptophan derivative.