The endoplasmic reticulum (ER) is the largest organelle in the cell, and its functions have been studied for decades. The past several years have provided novel insights into the existence of distinct domains between the ER and other organelles, known as membrane contact sites (MCSs). At these contact sites, organelle membranes are closely apposed and tethered, but do not fuse. Here, various protein complexes can work in concert to perform specialized functions such as binding, sensing and transferring molecules, as well as engaging in organelle biogenesis and dynamics. This Review describes the structure and functions of MCSs, primarily focusing on contacts of the ER with mitochondria and endosomes.

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Structure and function of ER membrane contact sites with other organelles.

TPC proteins are phosphoinositide- activated sodium-selective ion channels in endosomes and lysosomes.

Ebola virus causes sporadic outbreaks of lethal hemorrhagic fever in humans, but there is no currently approved therapy. Cells take up Ebola virus by macropinocytosis, followed by trafficking through endosomal vesicles. However, few factors controlling endosomal virus movement are known. Here we find that Ebola virus entry into host cells requires the endosomal calcium channels called two-pore channels (TPCs). Disrupting TPC function by gene knockout, small interfering RNAs, or small-molecule inhibitors halted virus trafficking and prevented infection. Tetrandrine, the most potent small molecule that we tested, inhibited infection of human macrophages, the primary target of Ebola virus in vivo, and also showed therapeutic efficacy in mice. Therefore, TPC proteins play a key role in Ebola virus infection and may be effective targets for antiviral therapy.

http://science.sciencemag.org/content/sci/347/6225/995.full.pdf

Two-pore channels control Ebola virus host cell entry and are drug targets for disease treatment

Endosomes, lysosomes and lysosome-related organelles are emerging as important Ca2+ storage cellular compartments with a central role in intracellular Ca2+ signalling. Endocytosis at the plasma membrane forms endosomal vesicles which mature to late endosomes and culminate in lysosomal biogenesis. During this process, acquisition of different ion channels and transporters progressively changes the endolysosomal luminal ionic environment (e.g. pH and Ca2+) to regulate enzyme activities, membrane fusion/fission and organellar ion fluxes, and defects in these can result in disease. In the present review we focus on the physiology of the inter-related transport mechanisms of Ca2+ and H+ across endolysosomal membranes. In particular, we discuss the role of the Ca2+-mobilizing messenger NAADP (nicotinic acid adenine dinucleotide phosphate) as a major regulator of Ca2+ release from endolysosomes, and the recent discovery of an endolysosomal channel family, the TPCs (two-pore channels), as its principal intracellular targets. Recent molecular studies of endolysosomal Ca2+ physiology and its regulation by NAADP-gated TPCs are providing exciting new insights into the mechanisms of Ca2+-signal initiation that control a wide range of cellular processes and play a role in disease. These developments underscore a new central role for the endolysosomal system in cellular Ca2+ regulation and signalling.

Molecular mechanisms of endolysosomal Ca2+ signalling in health and disease

Mutations in the leucine-rich repeat kinase-2 (LRRK2) gene cause late-onset Parkinson’s disease, but its physiological function has remained largely unknown. Here we report that LRRK2 activates a calcium-dependent protein kinase kinase-β (CaMKK-β)/adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathway which is followed by a persistent increase in autophagosome formation. Simultaneously, LRKR2 overexpression increases the levels of the autophagy receptor p62 in a protein synthesis-dependent manner, and decreases the number of acidic lysosomes. The LRRK2-mediated effects result in increased sensitivity of cells to stressors associated with abnormal protein degradation. These effects can be mimicked by the lysosomal Ca2+-mobilizing messenger nicotinic acid adenine dinucleotide phosphate (NAADP) and can be reverted by an NAADP receptor antagonist or expression of dominant-negative receptor constructs. Collectively, our data indicate a molecular mechanism for LRRK2 deregulation of autophagy and reveal previously unidentified therapeutic targets.

/pdf/leucine-rich-repeat-kinase-2-regulates-autophagy-through-a-57ag4a06c0.pdf

Leucine-rich repeat kinase 2 regulates autophagy through a calcium-dependent pathway involving NAADP

https://www.cell.com/current-biology/pdf/S0960-9822(10)00237-X.pdf

John A. Ward

Papers

Purified TPC Isoforms Form NAADP Receptors with Distinct Roles for Ca2+ Signaling and Endolysosomal Trafficking