Oxygen consumption is oftentimes used as a proxy for metabolic rate. However, pupfish acclimated to ecologically relevant temperatures may employ extended periods of anaerobism despite the availability of oxygen-a process we called paradoxical anaerobism. In this study, we evaluated data from pupfish exhibiting stable oxygen consumption. Routine oxygen consumption ([Formula: see text]) of a refuge population derived from Cyprinodon spp. acclimated to 28 and 33 °C was evaluated at the ecologically relevant assay temperatures of between 25 and 38 °C. Different interpretations of the data are available depending on normalization. For instance, [Formula: see text] of smaller fish, measured per fish, was remarkably stable over a wide range of assay temperatures and was not different between acclimation groups. However, when measured on a mass-specific basis, [Formula: see text] in these same smaller fish increases more predictably as temperature increased. [Formula: see text] of refuge fish and the closely related pupfish, C. nevadensis mionectes, measured near their respective acclimation temperatures, were essentially identical. However, [Formula: see text] of 28 °C acclimated fish of both species, when measured at 34 °C, was greater than that of the 33 °C acclimated fish measured at 28 °C. We suggest that this observed 'efficiency' may result from significant anaerobic metabolism use. Experiments investigating factorial aerobic scope ([Formula: see text]/[Formula: see text]) yielded values less than 1 in 21-36% of the 33 °C acclimated fish. These values indicate a substantial contribution of anaerobic metabolism to energy utilization by these fish. However, muscle lactate levels are not elevated in exercising fish-a result that is consistent with paradoxical anaerobism use.

The habitat of the critically endangered Devils Hole Pupfish, Cyprinodon diabolis is marked by constant high temperatures and low oxygen availability. In order to explore the effects of these conditions on development and recruitment of eggs in Devils Hole, we tested the effects of two ecologically relevant temperatures on the development, hatch success, and oxygen consumption of eggs from a refuge population of pupfish derived from C. diabolis and eggs from its close sister species, Cyprinodon nevadensis mionectes. We developed a simple method to measure oxygen consumption in a single egg. Parent acclimation temperature, rather than incubation temperature, was the most important factor influencing hatch success. Eggs incubated at 33°C hatched more quickly compared to those incubated at 28°C. Despite this accelerated development, larvae from both temperatures were of similar size at hatch. Unexpectedly, eggs incubated at 33°C experience lower than expected oxygen consumption rates compared to those incubated at 28°C. Oxygen consumption rates would be limited at PO values that are much higher than environmental oxygen tensions. Oxygen consumption increased dramatically upon hatch, indicating that low oxygen conditions such as those present in Devils Hole may limit developing eggs.

Conventional protein kinase C (cPKC) isozymes tune the signaling output of cells, with loss-of-function somatic mutations associated with cancer and gain-of-function germline mutations identified in neurodegeneration. PKC with impaired autoinhibition is removed from the cell by quality-control mechanisms to prevent accumulation of aberrantly active enzyme. Here, we examine how a highly conserved residue in the C1A domain of cPKC isozymes permits quality-control degradation when mutated to histidine in cancer (PKCβ-R42H) and blocks downregulation when mutated to proline in the neurodegenerative disease spinocerebellar ataxia (PKCγ-R41P). Using FRET-based biosensors, we determined that mutation of R42 to any residue, including lysine, resulted in reduced autoinhibition as indicated by higher basal activity and faster agonist-induced plasma membrane translocation. R42 is predicted to form a stabilizing salt bridge with E655 in the C-tail and mutation of E655, but not neighboring E657, also reduced autoinhibition. Western blot analysis revealed that whereas R42H had reduced stability, the R42P mutant was stable and insensitive to activator-induced ubiquitination and downregulation, an effect previously observed by deletion of the entire C1A domain. Molecular dynamics (MD) simulations and analysis of stable regions of the domain using local spatial pattern (LSP) alignment suggested that P42 interacts with Q66 to impair mobility and conformation of one of the ligand-binding loops. Additional mutation of Q66 to the smaller asparagine (R42P/Q66N), to remove conformational constraints, restored degradation sensitivity. Our results unveil how disease-associated mutations of the same residue in the C1A domain can toggle between gain- or loss-of-function of PKC.

ii DISCLAIMER Recovery plans delineate reasonable actions required to recover and protect the species. The U.S. Fish and Wildlife Service (Service) prepares the plans, sometimes with the assistance of recovery teams, contractors, State and Federal Agencies, and others. Objectives are attained and any necessary funds made available subject to budgetary and other constraints affecting the parties involved, as well as the need to address other priorities. Time and costs provided for individual tasks are estimates only, and not to be taken as actual or budgeted expenditures. Recovery plans do not necessarily represent the views nor official positions or approval of any persons or agencies involved in the plan formulation, other than the Service. They represent the official position of the Service only after they have been signed by the Regional Director or Director as approved. Approved recovery plans are subject to modification as dictated by new findings, changes in species status, and the completion of recovery tasks. Gila Topminnow Revised Recovery PlanDecember 1998iii ACKNOWLEDGMENTSOriginal preparation of the revised Gila topminnow Recovery Plan (1994) was done