Autophagy is a process by which components of the cell are degraded to maintain essential activity and viability in response to nutrient limitation. Extensive genetic studies have shown that the yeast ATG1 kinase has an essential role in autophagy induction. Furthermore, autophagy is promoted by AMP activated protein kinase (AMPK), which is a key energy sensor and regulates cellular metabolism to maintain energy homeostasis. Conversely, autophagy is inhibited by the mammalian target of rapamycin (mTOR), a central cell-growth regulator that integrates growth factor and nutrient signals. Here we demonstrate a molecular mechanism for regulation of the mammalian autophagy-initiating kinase Ulk1, a homologue of yeast ATG1. Under glucose starvation, AMPK promotes autophagy by directly activating Ulk1 through phosphorylation of Ser 317 and Ser 777. Under nutrient sufficiency, high mTOR activity prevents Ulk1 activation by phosphorylating Ulk1 Ser 757 and disrupting the interaction between Ulk1 and AMPK. This coordinated phosphorylation is important for Ulk1 in autophagy induction. Our study has revealed a signalling mechanism for Ulk1 regulation and autophagy induction in response to nutrient signalling.

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AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1

The DNA Damage Response: Making It Safe to Play with Knives

Fluorogenic and chromogenic chemosensors and reagents for anions.

The brain is a singular organ of unique biological complexity that serves as the command center for cognitive and motor function. As such, this specialized system also possesses a unique chemical composition and reactivity at the molecular level. In this regard, two vital distinguishing features of the brain are its requirements for the highest concentrations of metal ions in the body and the highest per-weight consumption of body oxygen. In humans, the brain accounts for only 2% of total body mass but consumes 20% of the oxygen that is taken in through respiration. As a consequence of high oxygen demand and cell complexity, distinctly high metal levels pervade all regions of the brain and central nervous system. Structural roles for metal ions in the brain and the body include the stabilization of biomolecules in static (e.g., Mg2+ for nucleic acid folds, Zn2+ in zinc-finger transcription factors) or dynamic (e.g., Na+ and K+ in ion channels, Ca2+ in neuronal cell signaling) modes, and catalytic roles for brain metal ions are also numerous and often of special demand.

Metals in Neurobiology: Probing Their Chemistry and Biology with Molecular Imaging

Artificial Organic Host Molecules for Anions

https://ir.lib.hiroshima-u.ac.jp/files/public/1/17109/20141016130321626953/MCP_5_749.pdf

Phosphate-binding Tag, a New Tool to Visualize Phosphorylated Proteins

We provide a standard phosphate-affinity SDS-PAGE (Mn2+–Phos-tag SDS-PAGE) protocol, in which Phos-tag is used to analyze large phosphoproteins with molecular masses of more than 200 kDa. A previous protocol required a long electrophoresis time of 12 h for separation of phosphoisotypes of large proteins (∼150 kDa). This protocol, which uses a 3% (wt/vol) polyacrylamide gel strengthened with 0.5% (wt/vol) agarose, permits the separation of protein phosphoisotypes larger than 200 kDa within 2 h. In subsequent immunoblotting, phosphoisotypes of high-molecular-mass proteins, such as mammalian target of rapamycin (289 kDa), ataxia telangiectasia-mutated kinase (350 kDa) and p53-binding protein 1 (213 kDa), can be clearly detected as up-shifted migration bands on the improved Mn2+–Phos-tag SDS-PAGE gel. The procedure from the beginning of gel preparation to the end of electrophoresis requires about 4 h in this protocol.

Separation and detection of large phosphoproteins using Phos-tag SDS-PAGE

Zinc-fluorophores have recently been attracting much interest in biological and environmental applications. How to detect selectively trace Zn2+ with efficient signal transduction is the central problem. Following carbonic anhydrase-based biosensors with fluorescent aromatic sulfonamides, a chemosensor, Zinquin, is now extensively used to study the role of intracellular Zn2+ in cellular biology. New types of zinc-fluorophores, zinc-finger peptides attached with fluorescent dyes and a dansylamide-pendant macrocyclic polyamine, have been developed in 1996. The principles, properties and limitations of these are discussed.

Tohru Koike

Papers

Phosphate-binding Tag, a New Tool to Visualize Phosphorylated Proteins

Separation and detection of large phosphoproteins using Phos-tag SDS-PAGE

Recent development of zinc-fluorophores

A zinc(II) complex of 1,5,9-triazacyclododecane ([12]aneN3) as a model for carbonic anhydrase

Roles of zinc(II) ion in phosphatases. A model study with zinc(II)-macrocyclic polyamine complexes