TPC2 is a novel NAADP-sensitive Ca2+ release channel, operating as a dual sensor of luminal pH and Ca2+

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a molecule capable of initiating the release of intracellular Ca2+ required for many essential cellular processes. Recent evidence links two-pore channels (TPCs) with NAADP-induced release of Ca2+ from lysosome-like acidic organelles; however, there has been no direct demonstration that TPCs can act as NAADP-sensitive Ca2+-release channels. Controversial evidence also proposes ryanodine receptors as the primary target of NAADP. We show that TPC2, the major lysosomal targeted isoform, is a cation channel with selectivity for Ca2+ that will enable it to act as a Ca2+ release channel in the cellular environment. NAADP opens TPC2 channels in a concentration-dependent manner, binding to high affinity activation and low affinity inhibition sites. At the core of this process is the luminal environment of the channel. The sensitivity of TPC2 to NAADP is steeply dependent on the luminal [Ca2+] allowing extremely low levels of NAADP to open the channel. In parallel, luminal pH controls NAADP affinity for TPC2 by switching from reversible activation of TPC2 at low pH to irreversible activation at neutral pH. Further evidence earmarking TPCs as the likely pathway for NAADP-induced intracellular Ca2+ release is obtained from the use of Ned-19, the selective blocker of cellular NAADP-induced Ca2+ release. Ned-19 antagonizes NAADP-activation of TPC2 in a non-competitive manner at 1 μM but potentiates NAADP activation at nanomolar concentrations. This single-channel study provides a long awaited molecular basis for the peculiar mechanistic features of NAADP signaling and a framework for understanding how NAADP can mediate key physiological events.Publisher PDFPeer reviewe

An emerging role for NAADP-mediated Ca2+ signaling in the pancreatic beta-cell

Several recent reports, including one in this journal, have reignited the debate about whether the calcium-mobilizing messenger, nicotinic adenine nucleotide diphosphate (NAADP) plays a central role in the regulation of calcium signalling in pancreatic β-cell. These studies have highlighted a role for NAADP-induced Ca(2+) mobilization not only in mediating the effects of the incretin, GLP-1 and the autocrine proliferative effects of insulin, but also possibly a fundamental role in glucose-mediated insulin secretion in the pancreatic β-cell

Estimating Wildfire-Generated Ozone over North America Using Ozonesonde Profiles and a Differential Back Trajectory Technique

An objective method, employing HYSPLIT back-trajectories and Moderate Resolution Imaging Spectroradiometer (MODIS) fire observations, is developed to estimate ozone enhancement in air transported from regions of active forest fires at 18 ozone sounding sites located across North America. The Differential Back Trajectory (DBT) method compares mean differences between ozone concentrations associated with fire-affected and fire-unaffected parcels. It is applied to more than 1100 ozonesonde profiles collected from these sites during the summer months June to August 2006, 2008, 2010 and 2011. Layers of high ozone associated with low humidity were first removed from the ozonesonde profiles to minimize the potential effects of stratospheric intrusions on the calculations. No significant influence on average ozone levels by North American fires was found for stations located at Arctic latitudes. The ozone enhancement for stations nearer large fires, such as Trinidad Head and Bratt\u27s Lake, was up to 4.8% of the TTOC (Total Tropospheric Ozone Column). Fire ozone accounted for up to 8.3% of TTOC at downwind sites such as Yarmouth, Sable Island, Narragansett, and Walsingham. The results are consistent with other studies that have reported an increase in ozone production with the age of the smoke plume

PLCζ is the physiological trigger of the Ca2+ oscillations that induce embryogenesis in mammals but conception can occur in its absence

Activation of the egg by the sperm is the first, vital stage of embryogenesis. The sperm protein PLC zeta has been proposed as the physiological agent that triggers the Ca2+ oscillations that normally initiate embryogenesis. Consistent with this, recombinant PLC zeta induces Ca2+ oscillations in eggs and debilitating mutations in the PLCZ1 gene are associated with infertility in men. However, there has been no evidence that knockout of the gene encoding PLC. abolishes the ability of sperm to induce Ca2+ oscillations in eggs. Here, we show that sperm derived from Plcz1(-/-) male mice fail to trigger Ca2+ oscillations in eggs, cause polyspermy and thus demonstrate that PLC zeta is the physiological trigger of these Ca2+ oscillations. Remarkably, some eggs fertilized by PLC zeta-null sperm can develop, albeit at greatly reduced efficiency, and after a significant time-delay. In addition, Plcz1(-/-) males are subfertile but not sterile, suggesting that in the absence of PLC zeta, spontaneous egg activation can eventually occur via an alternative route. This is the first demonstration that in vivo fertilization without the normal physiological trigger of egg activation can result in offspring. PLC zeta-null sperm now make it possible to resolve long-standing questions in fertilization biology, and to test the efficacy and safety of procedures used to treat human infertility

Both RyRs and TPCs are required for NAADP-induced intracellular Ca2+ release

Intracellular Ca2+ release is mostly mediated by inositol trisphosphate, but intracellular cyclic-ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP) are important messengers in many systems. Whereas cADPR generally activates type 2 ryanodine receptors (RyR2s), the NAADP-activated Ca2+ release mechanism is less clear. Using knockouts and antibodies against RyRs and Two-Pore Channels (TPCs), we have compared their relative importance for NAADP-induced Ca2+ release from two-photon permeabilized pancreatic acinar cells. In these cells, cholecystokinin-elicited Ca2+ release is mediated by NAADP. TPC2-KO reduced NAADP-induced Ca2+ release by 64%, but the combination of TPC2-KO and an antibody against TPC1, significantly reduced Ca2+ release by 86% (64% vs. 86%, p TPC2 > RyR3 > TPC1 >> RyR2. However, when assessing NAADP-induced Ca2+ release solely from the acidic stores (granules/endosomes/lysosomes), antibodies against TPC2 and TPC1 virtually abolished the Ca2+ liberation as did antibodies against RyR1 and RyR3. Our results indicate that the primary, but very small, NAADP-elicited Ca2+ release via TPCs from endosomes/lysosomes triggers the detectable Ca2+-induced Ca2+ release via RyR1 and RyR3 occurring from the granules and the ER

Identification of a Novel Gene for Diabetic Traits in Rats, Mice, and Humans

The genetic basis of type 2 diabetes remains incompletely defined despite the use of multiple genetic strategies. Multiparental populations such as heterogeneous stocks (HS) facilitate gene discovery by allowing fine mapping to only a few megabases, significantly decreasing the number of potential candidate genes compared to traditional mapping strategies. In the present work, we employed expression and sequence analysis in HS rats (Rattus norvegicus) to identify Tpcn2 as a likely causal gene underlying a 3.1-Mb locus for glucose and insulin levels. Global gene expression analysis on liver identified Tpcn2 as the only gene in the region that is differentially expressed between HS rats with glucose intolerance and those with normal glucose regulation. Tpcn2 also maps as a cis-regulating expression QTL and is negatively correlated with fasting glucose levels. We used founder sequence to identify variants within this region and assessed association between 18 variants and diabetic traits by conducting a mixed-model analysis, accounting for the complex family structure of the HS. We found that two variants were significantly associated with fasting glucose levels, including a nonsynonymous coding variant within Tpcn2. Studies in Tpcn2 knockout mice demonstrated a significant decrease in fasting glucose levels and insulin response to a glucose challenge relative to those in wild-type mice. Finally, we identified variants within Tpcn2 that are associated with fasting insulin in humans. These studies indicate that Tpcn2 is a likely causal gene that may play a role in human diabetes and demonstrate the utility of multiparental populations for positionally cloning genes within complex loci

The state of the Martian climate

60°N was +2.0°C, relative to the 1981–2010 average value (Fig. 5.1). This marks a new high for the record. The average annual surface air temperature (SAT) anomaly for 2016 for land stations north of starting in 1900, and is a significant increase over the previous highest value of +1.2°C, which was observed in 2007, 2011, and 2015. Average global annual temperatures also showed record values in 2015 and 2016. Currently, the Arctic is warming at more than twice the rate of lower latitudes