Dyrk1A phosphorylates alpha-synuclein and enhances intracellular inclusion formation

Lewy bodies (LBs) are pathological hallmarks of Parkinson disease (PD) but also occur in Alzheimer disease (AD) and dementia of LBs. Alpha-synuclein, the major component of LBs, is observed in the brain of Down syndrome (DS) patients with AD. Dyrk1A, a dual specificity tyrosine-regulated kinase (Dyrk) family member, is the mammalian ortholog of the Drosophila minibrain (Mnb) gene, essential for normal postembryonic neurogenesis. The Dyrk1A gene resides in the human chromosome 21q22.2 region, which is associated with DS anomalies, including mental retardation. In this study, we examined whether Dyrk1A interacts with alpha-synuclein and subsequently affects intracellular alpha-synuclein inclusion formation in immortalized hippocampal neuronal (H19-7) cells. Dyrk1A selectively binds to alpha-synuclein in transformed and primary neuronal cells. Alpha-synuclein overexpression, followed by basic fibroblast growth factor-induced neuronal differentiation, resulted in cell death. We observed that accompanying cell death was increased alpha-synuclein phosphorylation and intracytoplasmic aggregation. In addition, the transfection of kinase-inactive Dyrk1A or Dyrk1A small interfering RNA blocked alpha-synuclein phosphorylation and aggregate formation. In vitro kinase assay of anti-Dyrk1A immunocomplexes demonstrated that Dyrk1A could phosphorylate alpha-synuclein at Ser-87. Furthermore, aggregates formed by phosphorylated alpha-synuclein have a distinct morphology and are more neurotoxic compared with aggregates composed of unmodified wild type alpha-synuclein. These findings suggest alpha-synuclein inclusion formation regulated by Dyrk1A, potentially affecting neuronal cell viability.     

Nanoporous protein matrix made of amyloid fibrils of β2‐microglobulin

Amyloid fibrils are considered as novel nanomaterials because of their nanoscale width, a regular constituting structure of cross β‐sheet conformation, and considerable mechanical strength. By using an amyloidogenic protein of β₂‐microglobulin (β₂M) related to dialysis‐related amyloidosis, nanoporous protein matrix has been prepared. The β₂M granules made of around 15 monomers showed an average size of 23.1 nm. They formed worm‐like fibrils at pH 7.4 in 20 mM sodium phosphate containing 0.15 M NaCl following vigorous nondirectional shaking incubation, in which they became laterally associated and interwound to generate the porous amyloid fibrillar matrix with an average pore size of 30–50 nm. This nanoporous protein matrix was demonstrated to be selectively disintegrated by reducing agents, such as tris‐(2‐carboxyethyl) phosphine. High surface area with nanopores on the surface has been suggested to make the matrix of β₂M amyloid fibrils particularly suitable for applications in the area of nanobiotechnology including drug delivery and tissue engineering. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

Osteoclast Derivation from Mouse Bone Marrow

Osteoclasts are highly specialized cells that are derived from the monocyte/macrophage lineage of the bone marrow. Their unique ability to resorb both the organic and inorganic matrices of bone means that they play a key role in regulating skeletal remodeling. Together, osteoblasts and osteoclasts are responsible for the dynamic coupling process that involves both bone resorption and bone formation acting together to maintain the normal skeleton during health and disease.As the principal bone-resorbing cell in the body, changes in osteoclast differentiation or function can result in profound effects in the body. Diseases associated with altered osteoclast function can range in severity from lethal neonatal disease due to failure to form a marrow space for hematopoiesis, to more commonly observed pathologies such as osteoporosis, in which excessive osteoclastic bone resorption predisposes to fracture formation.An ability to isolate osteoclasts in high numbers in vitro has allowed for significant advances in the understanding of the bone remodeling cycle and has paved the way for the discovery of novel therapeutic strategies that combat these diseases. Here, we describe a protocol to isolate and cultivate osteoclasts from mouse bone marrow that will yield large numbers of osteoclasts.     

Mutation of the lbp-5 gene alters metabolic output in Caenorhabditis elegans

Intracellular lipid-binding proteins (LBPs) impact fatty acid homeostasis in various ways, including fatty acid transport into mitochondria. However, the physiological consequences caused by mutations in genes encoding LBPs remain largely uncharacterized. Here, we explore the metabolic consequences of lbp-5 gene deficiency in terms of energy homeostasis in Caenorhabditis elegans. In addition to increased fat storage, which has previously been reported, deletion of lbp-5 attenuated mitochondrial membrane potential and increased reactive oxygen species levels. Biochemical measurement coupled to proteomic analysis of the lbp-5(tm1618) mutant revealed highly increased rates of glycolysis in this mutant. These differential expression profile data support a novel metabolic adaptation of C. elegans, in which glycolysis is activated to compensate for the energy shortage due to the insufficient mitochondrial β-oxidation of fatty acids in lbp-5 mutant worms. This report marks the first demonstration of a unique metabolic adaptation that is a consequence of LBP-5 deficiency in C. elegans. [BMB Reports 2014; 47(1): 15-20]     

Genome sequence of Lactobacillus salivarius GJ-24, a probiotic strain isolated from healthy adult intestine

The draft genome sequence of Lactobacillus salivarius GJ-24 isolated from the feces of healthy adults was determined. Its properties, including milk fermentation activity and bacteriocin production, suggest its potential uses as a probiotic lactic acid bacterium and start culture for dairy products.