Biology subjects

Rivera-Kohr, D. A.

Publications and source records attributed to Rivera-Kohr, D. A..

4 recordsLinked to original sources

bioRxiv2025

Patterns of Microbial Succession and Niche Differentiation Across Depth and Age in a Landfill Have Implications for Management Strategies

Despite microorganisms being primarily responsible for landfill material decomposition, limited characterization has been performed across both landfill depth and age. Here, we investigated the microbial communities and physicochemical parameters in two active and two closed landfill wells from surface to bottom, as well as fill dirt and leachate at a sanitary landfill near Madison, Wisconsin, USA. Amplicon community sequencing fungi, bacteria and archaea revealed distinct microbial community structures across landfill sites. The observed patterns of microbial community succession by depth and age mirror the known phases of the landfill life cycle. Younger surface samples were dominated by aerobic fungi, which transitioned to fermentative bacteria and methanogenic archaea in older, deeper, layers. Simultaneously, high species richness was preserved across landfill ages, while reduced evenness at specific depths support spatial niche differentiation. In conjunction with the lack of trends found for physicochemical parameters by depth, this supports niche differentiation driven by the highly heterogenous waste inputs. This study provides the first comprehensive vertical profile of bacterial, fungal, and archaeal communities across landfill depths and ages, highlighting the influence of these parameters on physicochemical factors and microbial distribution within landfills. These results have implications for improving landfill management, including renewable gas energy production, minimizing emissions, and increasing degradation rates.

microbiology↗

bioRxiv2020

Reciprocal regulation of vacuolar calcium transport and V-ATPase activity, and the effects of Phosphatidylinositol 3,5-bisphosphate

Yeast vacuoles are acidified by the V-ATPase, a protein complex comprised of the membrane embedded VO complex and the soluble cytoplasmic V1 complex. The assembly of the V1-VO holoenzyme is required for the transfer of H+ into the vacuole lumen for acidification. The assembly of the V1-VO holoenzyme is stabilized by the lipid phosphatidylinositol 3,5-bisphospate (PI(3,5)P2) made by the PI3P 5-kinase Fab1/PIKfyve. The absence of PI(3,5)P2 leads to the dissociation of the V1 complex from the membrane. Separately, PI(3,5)P2 has been shown to modulate Ca2+ transport across the vacuole membrane during fission and fusion. Here we examined whether the regulation of H+ and Ca2+ by PI(3,5)P2 are interdependent. We show that modulating extraluminal Ca2+ concentrations inhibit V-ATPase activity. As extraluminal CaCl2 levels are raised, the activity of H+ pumping is reduced. Conversely, chelating free Ca2+ with EGTA stimulated vacuole acidification. Not only did Ca2+ levels affect H+ translocation, we also show that blocking V-ATPase activity inhibited Ca2+ transport into the vacuole lumen. Together, these data illustrate that Ca2+ transport and V-ATPase regulation are interconnected through the modulation of vacuolar lipid profiles. Summary StatementHere we show that Ca2+ and H+ transport across the vacuole membrane is reciprocally regulated and that it is linked to the production of Phosphatidylinositol 3,5-bisphoshpate.

biochemistry↗

bioRxiv2020

Sphingolipids with Very Long-chain Fatty Acids Regulate Vacuole Fusion During Tethering and Docking

Sphingolipids are essential in membrane trafficking and cellular homeostasis. Here, we show that sphingolipids containing very long-chain fatty acids (VLCFAs) promote robust homotypic vacuolar fusion in Saccharomyces cerevisiae. The elongase Elo3 adds two carbons to 24-carbon (C24) acyl chains to make C26 VLCFAs that are incorporated into sphingolipids. Vacuoles isolated from elo3{Delta} cells had increased fluidity relative to the wild-type and were attenuated for fusion. Upon further testing we found that vesicle the tethering stage was affected as elo3{Delta} vacuole clusters contained fewer vesicles versus the WT. Vacuole tethering requires the interactions of late endosomal Rab GTPase Ypt7 and the HOPS tethering complex. Pulldown assays using GST-Ypt7 showed that HOPS from elo3{Delta} vacuole extracts failed to bind Ypt7 while HOPS from WT extracts interacted with GST-Ypt7. Furthermore GFP-Ypt7 failed to localize at vertex microdomains of elo3{Delta} vacuoles relative to the WT, whereas HOPS and regulatory lipids did accumulate at vertices. Finally, we found that elo3{Delta} vacuoles had reduced V-ATPase. Together these data show that C26-VLCFA containing sphingolipids are important for Ytp7 function and vacuole homeostasis.

cell biology↗

bioRxiv2019

Copper Blocks V-ATPase Activity and SNARE Complex Formation to Inhibit Yeast Vacuole Fusion

The accumulation of Copper in organisms can lead to altered functions of various pathways, and become cytotoxic through the generation of reactive oxygen species. In yeast, cytotoxic metals such as Hg+, Cd2+, and Cu2+ are transported into the lumen of the vacuole through various pumps. Copper ions are initially transported into the cell by the copper transporter Ctr1 at the plasma membrane and sequestered by chaperones and other factors to prevent cellular damage by free cations. Excess copper ions can subsequently be transported into the vacuole lumen by an unknown mechanism. Transport across membranes requires the reduction of Cu2+ to Cu+. Labile copper ions can interact with membranes to alter fluidity, lateral phase separation and fusion. Here we found that CuCl2 potently inhibited vacuole fusion by blocking SNARE pairing. This was accompanied by the inhibition of V-ATPase H+ pumping. Deletion of the vacuolar reductase Fre6 had no effect on the inhibition of fusion by copper. This suggests that that Cu2+ is responsible for the inhibition of vacuole fusion and V-ATPase function. This notion is supported by the differential effects chelators. The Cu2+-specific chelator TETA rescued fusion, whereas the Cu+-specific chelator BCS had no effect on the inhibited fusion.

biochemistry↗