Genetic association between human chitinases and lung function in COPD

Two primary chitinases have been identified in humans--acid mammalian chitinase (AMCase) and chitotriosidase (CHIT1). Mammalian chitinases have been observed to affect the host's immune response. The aim of this study was to test for association between genetic variation in the chitinases and phenotypes related to chronic obstructive pulmonary disease (COPD). Polymorphisms in the chitinase genes were selected based on previous associations with respiratory diseases. Polymorphisms that were associated with lung function level or rate of decline in the Lung Health Study (LHS) cohort were analyzed for association with COPD affection status in four other COPD case-control populations. Chitinase activity and protein levels were also related to genotypes. In the caucasian LHS population, the baseline forced expiratory volume in one second (FEV(1)) was significantly different between the AA and GG genotypic groups of the AMCase rs3818822 polymorphism. Subjects with the GG genotype had higher AMCase protein and chitinase activity compared with AA homozygotes. For CHIT1 rs2494303, a significant association was observed between rate of decline in FEV(1) and the different genotypes. In the African American LHS population, CHIT1 rs2494303 and AMCase G339T genotypes were associated with rate of decline in FEV(1). Although a significant effect of chitinase gene alleles was found on lung function level and decline in the LHS, we were unable to replicate the associations with COPD affection status in the other COPD study groups.     

Metabolic switching of human myotubes is improved by n-3 fatty acids

The aim of the present study was to examine whether pretreatment with different fatty acids, as well as the liver X receptor (LXR) agonist T0901317, could modify metabolic switching of human myotubes. The n-3 FA eicosapentaenoic acid (EPA) increased suppressibility, the ability of glucose to suppress FA oxidation. Substrate-regulated flexibility, the ability to increase FA oxidation when changing from a high glucose, low fatty acid condition (“fed”) to a high fatty acid, low glucose (“fasted”) condition, was increased by EPA and other n-3 FAs. Adaptability, the capacity to increase FA oxidation with increasing FA availability, was enhanced after pretreatment with EPA, linoleic acid (LA), and palmitic acid (PA). T0901317 counteracted the effect of EPA on suppressibility and adaptability, but it did not affect these parameters alone. EPA per se accumulated less, however, EPA, LA, oleic acid, and T0901317 treatment increased the number of lipid droplets (LD) in myotubes. LD volume and intensity, as well as mitochondrial mass, were independent of FA pretreatment. Microarray analysis showed that EPA regulated more genes than the other FAs and that specific pathways involved in carbohydrate metabolism were induced only by EPA. The present study suggests a favorable effect of n-3 FAs on skeletal muscle metabolic switching and glucose utilization.     

Structural basis of Blastomyces Endoglucanase-2 adjuvancy in anti-fungal and -viral immunity

The development of safe subunit vaccines requires adjuvants that augment immunogenicity of non-replicating protein-based antigens. Current vaccines against infectious diseases preferentially induce protective antibodies driven by adjuvants such as alum. However, the contribution of antibody to host defense is limited for certain classes of infectious diseases such as fungi, whereas animal studies and clinical observations implicate cellular immunity as an essential component of the resolution of fungal pathogens. Here, we decipher the structural bases of a newly identified glycoprotein ligand of Dectin-2 with potent adjuvancy, Blastomyces endoglucanase-2 (Bl-Eng2). We also pinpoint the developmental steps of antigen-specific CD4⁺ and CD8⁺ T responses augmented by Bl-Eng2 including expansion, differentiation and tissue residency. Dectin-2 ligation led to successful systemic and mucosal vaccination against invasive fungal infection and Influenza A infection, respectively. O-linked glycans on Bl-Eng2 applied at the skin and respiratory mucosa greatly augment vaccine subunit- induced protective immunity against lethal influenza and fungal pulmonary challenge.     

Association of the FAM46A Gene VNTRs and BAG6 rs3117582 SNP with Non Small Cell Lung Cancer (NSCLC) in Croatian and Norwegian Populations

We analyzed for associations between a variable number of tandem repeat (VNTR) polymorphism in the Family with sequence similarity 46, member A (FAM46A) gene and a single nucleotide polymorphism (rs3117582) in the BCL2-Associated Athanogene 6 (BAG6) with non small cell lung cancer in Croatian and Norwegian subjects. A total of 503 (262 Croatian and 241Norwegian) non small cell lung cancer patients and 897 controls (568 Croatian and 329 Norwegian) were analyzed. We found that the frequency of allele b (three VNTR repeats) of FAM46A gene was significantly increased in the patients compared to the healthy controls in the Croatian and the combined Croatian and Norwegian subjects. Genotype frequencies of cd (four and five VNTR repeats) and cc (four VNTR repeats homozygote) of the FAM46A gene were significantly decreased in the patients compared to the healthy controls in the Croatian and Norwegian subjects, respectively. Logistic regression analyses revealed FAM46A genotype cc to be an independent predictive factor for non small cell lung cancer risk in the Norwegian subjects after adjustment for age, gender and smoking status. This is the first study to suggest an association between the FAM46A gene VNTR polymorphisms and non small cell lung cancer. We found also that BAG6 rs3117582 SNP was associated with non small cell lung cancer in the Norwegian subjects and the combined Croatian-Norwegian subjects corroborating the earlier finding that BAG6 rs3117582 SNP was associated with lung cancer in Europeans. Logistic regression analyses revealed that genotypes and alleles of BAG6 were independent predictive factor for non small cell lung cancer risk in the Norwegian and combined Croatian-Norwegian subjects, after adjustment for age and gender.     

Protein Tyrosine Phosphatase 1B Regulates Pyruvate Kinase M2 Tyrosine Phosphorylation

Protein-tyrosine phosphatase 1B (PTP1B) is a physiological regulator of glucose homeostasis and adiposity and is a drug target for the treatment of obesity and diabetes. Here we identify pyruvate kinase M2 (PKM2) as a novel PTP1B substrate in adipocytes. PTP1B deficiency leads to increased PKM2 total tyrosine and Tyr¹⁰⁵ phosphorylation in cultured adipocytes and in vivo. Substrate trapping and mutagenesis studies identify PKM2 Tyr-105 and Tyr-148 as key sites that mediate PTP1B-PKM2 interaction. In addition, in vitro analyses illustrate a direct effect of Tyr-105 phosphorylation on PKM2 activity in adipocytes. Importantly, PTP1B pharmacological inhibition increased PKM2 Tyr-105 phosphorylation and decreased PKM2 activity. Moreover, PKM2 Tyr-105 phosphorylation is regulated nutritionally, decreasing in adipose tissue depots after high-fat feeding. Further, decreased PKM2 Tyr-105 phosphorylation correlates with the development of glucose intolerance and insulin resistance in rodents, non-human primates, and humans. Together, these findings identify PKM2 as a novel substrate of PTP1B and provide new insights into the regulation of adipose PKM2 activity.     

Ultrastructural characterization of giant endosomes induced by GTPase-deficient Rab5

The small GTPase Rab5 controls the fusogenic properties of early endosomes through GTP-dependent recruitment and activation of effector proteins. Expression of a GTPase-defective mutant, Rab5(Q79L), is known to cause formation of enlarged early endosomes. The ability of Rab5-GTP to recruit multiple effectors raises the question whether the Rab5(Q79L)-induced giant endosomes simply represent enlarged early endosomes or whether they have a more complex phenotype. In this report, we have addressed this issue by generating a HEp2 cell line with inducible expression of Rab5(Q79L) and performing ultrastructural analysis of Rab5(Q79L)-induced endosomes. We find that Rab5(Q79L) not only induces formation of enlarged early endosomes but also causes enlargement of later endocytic profiles. Most strikingly, Rab5(Q79L) causes formation of enlarged multivesicular endosomes with a large number of intraluminal vesicles, and endosomes that contain both early and late endocytic markers are frequently observed. In addition, we observe defects in the sorting of the EGF receptor and the transferrin receptor through this compartment.     

Re-routing of the invariant chain to the direct sorting pathway by introduction of an AP3-binding motif from LIMP II

AP3 is a heteromeric adaptor protein complex involved in the biogenesis of late endosomal/lysosomal structures. It recognizes tyrosine- and leucine-based sorting signals present in the cytoplasmic tails or loops of a number of proteins and is thought to be responsible for the direct transport of these proteins from the Golgi network to late endosomal/lysosomal structures. We have previously reported (Rodionov, H?ning, Silye, Kongsvik, von Figura, Bakke, 2002. Structural requirements for interactions between leucine-sorting signals and clathrin-associated adaptor protein complex AP3. J. Biol. Chem. 277, 47436-47443) that in vitro binding of AP3 to the leucine signals is dependent on the nature of three residues immediately upstream of the leucine signal and suggested that these three amino acids define whether the protein is sorted to endosomes via the plasma membrane (PM) or traffics directly to the late endosomes/lysosomes. In this paper, we show in vivo evidence that residues favoring AP3 binding introduced into a protein that is transported via the PM such as the invariant chain can re-route such protein into direct sorting to late endosomal/lysosomal structures.