Abstract The transport of Ca2+across membranes precedes the fusion and fission of various lipid bilayers. Yeast vacuoles under hyperosmotic stress become fragmented through fission events that requires the release of Ca2+stores through the TRP channel Yvc1. This requires the phosphorylation of phosphatidylinositol‐3‐phosphate (PI3P) by the PI3P‐5‐kinase Fab1 to produce transient PI(3,5)P2pools. Ca2+is also released during vacuole fusion upontrans‐SNARE complex assembly, however, its role remains unclear. The effect of PI(3,5)P2on Ca2+flux during fusion was independent of Yvc1. Here, we show that while low levels of PI(3,5)P2were required for Ca2+uptake into the vacuole, increased concentrations abolished Ca2+efflux. This was as shown by the addition of exogenous dioctanoyl PI(3,5)P2or increased endogenous production of by the hyperactivefab1T2250Amutant. In contrast, the lack of PI(3,5)P2on vacuoles from the kinase deadfab1EEEmutant showed delayed and decreased Ca2+uptake. The effects of PI(3,5)P2were linked to the Ca2+pump Pmc1, as its deletion rendered vacuoles resistant to the effects of excess PI(3,5)P2. Experiments with Verapamil inhibited Ca2+uptake when added at the start of the assay, while adding it after Ca2+had been taken up resulted in the rapid expulsion of Ca2+. Vacuoles lacking both Pmc1 and the H+/Ca2+exchanger Vcx1 lacked the ability to take up Ca2+and instead expelled it upon the addition of ATP. Together these data suggest that a balance of efflux and uptake compete during the fusion pathway and that the levels of PI(3,5)P2can modulate which path predominates.
more »
« less
Resolving Ultrafast Photoinitiated Dynamics of the Hachimoji 5‐aza‐7‐deazaguanine Nucleobase: Impact of Synthetically Expanding the Genetic Alphabet †
- Award ID(s):
- 1800052
- PAR ID:
- 10436068
- Date Published:
- Journal Name:
- Photochemistry and Photobiology
- Volume:
- 99
- Issue:
- 2
- ISSN:
- 0031-8655
- Page Range / eLocation ID:
- 693 to 705
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
Abstract Oxygen and hydrogen isotopes were used in this study to detect a hydraulic connection between a sinkhole lake and a karst spring. In karst areas, surface water that flows to a lake can drain through sinkholes in the lakebed to the underlying aquifer, and then flows in karst conduits and through aquifer matrix. At the study site located in northwest Florida, USA, Lake Miccosukee immediately drains into two sinkholes. Results from a dye tracing experiment indicate that lake water discharges at Natural Bridge Spring, a first‐magnitude spring 32 km downgradient from the lake. By collecting weekly water samples from the lake, the spring, and a groundwater well 10 m away from the lake during the dry period between October 2019 and January 2020, it was found that, when rainfall effects on isotopic signature in spring water are removed, increased isotope ratios of spring water can be explained by mixing of heavy‐isotope‐enriched lake water into groundwater, indicating hydraulic connection between the lake and the spring. Such a detection of hydraulic connection at the scale of tens of kilometers and for a first‐magnitude spring has not been previously reported in the literature. Based on the isotope ratio data, it was estimated that, during the study period, about 8.5% the spring discharge was the lake water that drained into the lake sinkholes.more » « less
-
Comprehensive Summary We report an efficient and convergent strategy for the total synthesis of UCS1025A and its diastereomer tetra‐epi‐UCS1025A. UCS1025A is a representative member of the naturally occurring pyrrolizidinone polyketides, from which members with potent antibacterial, antifungal, and anticancer activities have been identified. Our approach features a tandem carbonylative Stille cross coupling and Diels‐Alder reaction to forge a key C—C bond and build thetrans‐decalin system. This tandem process utilizes carbon monoxide as a one‐carbon linchpin to stitch a vinyl triflate and a vinylstannane together and form the desired enone moiety for the subsequent intramolecular Diels‐Alder cyclization. Our synthesis also provides a versatile approach for the synthesis of other related pyrrolizidinone‐containing polyketides.more » « less
An official website of the United States government

