Here, we directly tested the hypothesis that Zip14 transports free zinc, iron, and other metal ions by using the Xenopus laevis oocyte heterologous expression system, and use of this approach also allowed us to characterize the functional properties of Zip14. Expression of mouse Zip14 in RNAinjected oocytes stimulated the uptake of 55 Fe in the presence of L-ascorbate but not nitrilotriacetic acid, indicating that Zip14 is an iron transporter specific for ferrous ion (Fe 2ϩ ) over ferric ion (Fe 3ϩ uptake also was saturable (K0.5 Ϸ 2 M) but, notably, the metal-ion inhibition profile and Ca 2ϩ dependence of Zn 2ϩ transport differed from those of Fe 2ϩ transport, and we propose a model to account for these observations. Our data reveal that Zip14 is a complex, broad-scope metal-ion transporter. Whereas zinc appears to be a preferred substrate under normal conditions, we found that Zip14 is capable of mediating cellular uptake of NTBI characteristic of iron-overload conditions. cadmium transport; hereditary hemochromatosis; iron; homeostasis iron transport; Xenopus laevis oocyte; SLC39A14; thalassemia; zinc transport IRON-OVERLOAD CONDITIONS (e.g., thalassemia, hereditary hemochromatosis) are characterized by the appearance in plasma of nontransferrin-bound iron (NTBI) and result in cardiomyopathy, diabetes, hepatic cancer, and cirrhosis. Identification of the routes of cellular NTBI uptake will therefore provide novel targets for therapeutics.Zrt-and Irt-like protein-14 (Zip14) is a member of a large family of mammalian metal-ion transporters, the SLC39 gene family (6,11,12,22,26). Zip14 (synonyms SLC39A14, KIAA0062) is strongly expressed in the intestine (25, 35) but its subcellular localization there is not yet clear. Notably, Zip14 is abundantly expressed in the liver, heart, and pancreas (25, 35, 42), the major sites of organ damage in iron overload. Our previous data identified Zip14 as a candidate route for NTBI uptake since overexpression of Zip14 in human embryonic kidney (HEK)293, SF9, or HeLa cell lines stimulated NTBI uptake (14, 25), whereas small interfering RNA (siRNA) suppression of endogenous Zip14 in AML12 mouse hepatocytes decreased NTBI uptake (25).We have expressed mouse Zip14 in RNA-injected Xenopus oocytes, an efficient heterologous expression system ideal for direct assays of membrane transport and tolerant of broad manipulation of experimental conditions. We used radiotracer assays to test the hypothesis that Zip14 transports free iron and to examine the functional properties and metal-ion substrate profile of Zip14. MATERIALS AND METHODSReagents. Restriction enzymes were obtained from New England Biolabs (Ipswich, MA). All other reagents were obtained from SigmaAldrich (St. Louis, MO) or Research Products International (Prospect, IL) unless otherwise indicated.Expression of mouse Zip14 and human DMT1 in Xenopus oocytes. We performed laparotomy and ovariectomy on adult female Xenopus laevis frogs (Nasco, Fort Atkinson, WI) under 3-aminoethylbenzoate methanesulfonate anesthesia...

We analyzed Cassini Imaging Science Subsystem (ISS) images of the plume of Enceladus to derive particle number densities for the purpose of comparing our results with those obtained from other Cassini instrument investigations. Initial discrepancies in the results from different instruments, as large as factors of 10–20, can be reduced to ∼2 to 3 by accounting for the different times and geometries at which measurements were taken. We estimate the average daily ice production rate, between 2006 and 2010, to be 29 ± 7 kg/s, and a solid-to-vapor ratio, S/V > 0.06. At 50 km altitude, the plume's peak optical depth during the same time period was τ ∼ 10−3; by 2015, it was ∼10−4. Our inferred differential size distribution at 50 km altitude has an exponent q = 3. We estimate the average geothermal flux into the sea beneath Enceladus' south polar terrain to be comparable to that of the average Atlantic, of order 0.1 W/m2. Should microbes be present on Enceladus, concentrations at hydrothermal vents on Enceladus could be comparable to those on Earth, ∼105 cells/mL. We suggest the well-known process of bubble scrubbing as a means by which oceanic organic matter and microbes may be found in the plume in significantly enhanced concentrations: for the latter, as high as 107 cells/mL, yielding as many as 103 cells on a 0.04 m2 collector in a single 50 km altitude transect of the plume. Mission design can increase these numbers considerably. A lander mission, for example, catching falling plume particles on the same collector, could net, over 100 Enceladus days without bubble scrubbing, at least 105 cells; and, if bubble scrubbing is at work, up to 108 cells. Key Words: Enceladus—Microbe—Organic matter—Life detection. Astrobiology 17, 876–901.

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We use images acquired by the Cassini Imaging Science Subsystem (ISS) to investigate the temporal variation of the brightness and height of the south polar plume of Enceladus. The plume's brightness peaks around the moon's apoapse, but with no systematic variation in scale height with either plume brightness or Enceladus' orbital position. We compare our results, both alone and supplemented with Cassini near-infrared observations, with predictions obtained from models in which tidal stresses are the principal control of the eruptive behavior. There are three main ways of explaining the observations: (1) the activity is controlled by right-lateral strike slip motion; (2) the activity is driven by eccentricity tides with an apparent time delay of about 5 hr; (3) the activity is driven by eccentricity tides plus a 1:1 physical libration with an amplitude of about 0. • 8 (3.5 km). The second hypothesis might imply either a delayed eruptive response, or a dissipative, viscoelastic interior. The third hypothesis requires a libration amplitude an order of magnitude larger than predicted for a solid Enceladus. While we cannot currently exclude any of these hypotheses, the third, which is plausible for an Enceladus with a subsurface ocean, is testable by using repeat imaging of the moon's surface. A dissipative interior suggests that a regional background heat source should be detectable. The lack of a systematic variation in plume scale height, despite the large variations in plume brightness, is plausibly the result of supersonic flow; the details of the eruption process are yet to be understood.