Normal Ventricular and Regional Blood Flow Volumes and Native T1 Values in Healthy Japanese Children Obtained from Comprehensive Cardiovascular Magnetic Resonance Imaging.

Age-related mean and reference ranges for ventricular volumes and mass, regional blood flow measurements, and T1 values using cardiovascular magnetic resonance (CMR) imaging are yet to be established for the pediatric population. Especially in infants and toddlers, no consistent flow volume sets or T1 values have been reported. The purpose of this study was to determine the relevant normal values.Twenty-three children (aged 0.1-15.3 years) without cardiovascular diseases were included. Comprehensive CMR imaging including cine, 2-dimensional phase-contrast, and native T1 mapping, were performed. Ventricular volumes and masses, 11 sets of regional blood flow volumes, and myocardial and liver T1 values were measured. All intraclass correlation coefficient values were > 0.94, except for the right ventricular mass (0.744), myocardial (0.868) and liver T1 values (0.895), reflecting good to excellent agreement between rates.Regression analysis showed an exponential relationship between body surface area (BSA) and ventricular volumes, mass, and regional blood flow volumes (normal value = a*BSAb). Left ventricular myocardial T1 values were regressed on linear regression with age (normal value = -7.39*age + 1091), and hepatic T1 values were regressed on a quadratic function of age (normal value = 0.923*age2 -18.012*age + 613).Comparison of the 2 different methods for the same physical quantities by Bland-Altman plot showed no difference except that the right ventricular stroke volume was 1.5 mL larger than the main pulmonary trunk flow volume.This study provides the normal values for comprehensive CMR imaging in Japanese children.

Phosphoproteomic and proteomic profiling of serine/threonine protein kinase PkaE of Streptomyces coelicolor A3(2) and its role in secondary metabolism and morphogenesis.

This study aimed to investigate the role of serine/threonine kinase PkaE in Streptomyces coelicolor A3(2). Liquid chromatography tandem mass spectrometry was performed for comparative phosphoproteome and proteome analyses of S. coelicolor A3(2), followed by an in vitro phosphorylation assay. Actinorhodin production in the pkaE deletion mutant was lower than that in wild-type S. coelicolor A3(2), and the spores of the pkaE deletion mutant were damaged. Furthermore, phosphoproteome analysis revealed that 6 proteins were significantly differentially hypophosphorylated in pkaE deletion mutant (p < 0.05, fold-change ≤ 0.66), including BldG and FtsZ. In addition, the in vitro phosphorylation assay revealed that PkaE phosphorylated FtsZ. Comparative proteome analysis revealed 362 differentially expressed proteins (p < 0.05) and six downregulated proteins in the pkaE deletion mutant involved in actinorhodin biosynthesis. Gene ontology enrichment analysis revealed that PkaE participates in various biological and cellular processes. Hence, S. coelicolor PkaE participates in actinorhodin biosynthesis and morphogenesis.

Expression and characterization of Streptomyces coelicolor serine/threonine protein kinase PkaE.

We identified and characterized a new eukaryotic-type protein kinase (PkaE) from Streptomyces coelicolor A3 (2) M145. PkaE, consisting of 510 amino acid residues, is a cytoplasmic protein kinase and contains the catalytic domain of eukaryotic protein kinases in the N-terminal region. Recombinant PkaE was found to be autophosphorylated at threonine residues only. The disruption of chromosomal pkaE resulted in the overproduction of the actinorhodin-related blue pigment antibiotics. pkaE was expressed during the late growth phase in S. coelicolor A3 (2) M145, which corresponded to the production time of blue pigments. This result indicated that PkaE acts as a negative regulator for production of the secondary metabolites. In addition, PkaE was able to phosphorylate KbpA, a regulator involved in the AfsK-AfsR regulatory pathway.

Expression and characterization of the Streptomyces coelicolor serine/threonine protein kinase PkaD.

We identified and characterized the gene encoding a new eukaryotic-type protein kinase from Streptomyces coelicolor A3(2) M145. PkaD, consisting of 598 amino acid residues, contained the catalytic domain of eukaryotic protein kinases in the N-terminal region. A hydrophobicity plot indicated the presence of a putative transmembrane spanning sequence downstream of the catalytic domain, suggesting that PkaD is a transmembrane protein kinase. The recombinant PkaD was found to be phosphorylated at the threonine and tyrosine residues. In S. coelicolor A3(2), pkaD was transcribed as a monocistronic mRNA, and it was expressed constitutively throughout the life cycle. Disruption of chromosomal pkaD resulted in a significant loss of actinorhodin production. This result implies the involvement of pkaD in the regulation of secondary metabolism.

Peroxisome proliferator-activated receptor alpha and its response element are required but not sufficient for transcriptional activation of the mouse heart-type fatty acid binding protein gene.

Expression of heart-type fatty acid binding protein is restricted mainly to the skeletal and cardiac muscles and further regulated by peroxisome proliferator-activated receptor alpha. The molecular basis for the muscle-restricted peroxisome proliferator-activated receptor alpha action on the fatty acid binding gene was analyzed using normal and the receptor-null mice and the cultured cells. Two possible peroxisome proliferator-response elements were found in the promoter region of the mouse gene. A gel shift assay showed that both elements were functional. However, neither the tandem repeats of the elements nor the cloned promoter sequence could be activated by peroxisome proliferator-activated receptor alpha and its ligand in the reporter gene assay using cultured cells. The cloned promoter responded to the ligand only in the muscle when the reporter gene was introduced into the mouse muscle. Using a chimeric receptor with the activation domain of herpes virus VP16 protein and the tandem repeats of the elements with or without mutation, the upstream element was finally demonstrated to be potentially involved in the receptor-dependent transcriptional activation. These results suggest that the peroxisome proliferator-response element of the mouse gene is atypical and there is a muscle-specific mechanism to enhance the weak binding of the receptor to the response element to ensure the muscle-specific action of peroxisome proliferator-activated receptor alpha on the heart-type fatty acid binding protein gene promoter.

Sequences and evolutionary analyses of eukaryotic-type protein kinases from Streptomyces coelicolor A3(2).

Four eukaryotic-type protein serine/threonine kinases from Streptomyces coelicolor A3(2) were cloned and sequenced. To explore evolutionary relationships between these and other protein kinases, the distribution of protein serine/threonine kinase genes in prokaryotes was examined with the TFASTA program. Genes of this type were detected in only a few species of prokaryotes and their distribution was uneven; Streptomyces, Mycobacterium, Synechocystis and Myxococcus each contained more than three such genes. Homology analyses by GAP and Rdf2 programs suggested that some kinases from one species were closely related, whilst others were only remotely related. This was confirmed by examining phylogenetic trees constructed by the neighbour-joining and other methods. For each species, analysis of the coding regions indicated that the G+C content of protein kinase genes was similar to that of other genes. Considered with the fact that in phylogenetic trees the amino acid sequences of STPK from Aquifex aeolicus and some other eukaryotic-type protein kinases in prokaryotes form a cluster with protein kinases from eukaryotes, this suggests that the eukaryotic-type protein kinases were present originally in both prokaryotes and eukaryotes, but that most of these genes have been lost during the evolutionary process in prokaryotes because they are not needed. This conclusion is supported by the observation that the prokaryotes retaining several of these kinases undergo complicated morphological and/or biochemical differentiation.