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We tested the hypothesis that part of the lumenal amino acid segment between the two most C-terminal membrane segments of the skeletal muscle ryanodine receptor (RyR1) is important for channel activity and conductance. Eleven mutants were generated and expressed in HEK293 cells focusing on amino acid residue I4897 homologous to the selectivity filter of K(+) channels and six other residues in the M3-M4 lumenal loop. Mutations of amino acids not absolutely conserved in RyRs and IP(3)Rs (D4903A and D4907A) showed cellular Ca(2+) release in response to caffeine, Ca(2+)-dependent [(3)H]ryanodine binding, and single-channel K(+) and Ca(2+) conductances not significantly different from wild-type RyR1. Mutants with an I4897 to A, L, or V or D4917 to A substitution showed a decreased single-channel conductance, loss of high-affinity [(3)H]ryanodine binding and regulation by Ca(2+), and an altered caffeine-induced Ca(2+) release in intact cells. Mutant channels with amino acid residue substitutions that are identical in the RyR and IP(3)R families (D4899A, D4899R, and R4913E) exhibited a decreased K(+) conductance and showed a loss of high-affinity [(3)H]ryanodine binding and loss of single-channel pharmacology but maintained their response to caffeine in a cellular assay. Two mutations (G4894A and D4899N) were able to maintain pharmacological regulation both in intact cells and in vitro but had lower single-channel K(+) and Ca(2+) conductances than the wild-type channel. The results support the hypothesis that amino acid residues in the lumenal loop region between the two most C-terminal membrane segments constitute a part of the ion-conducting pore of RyR1.

Background: Cells respond to environmental stressors through several key pathways, including response to reactive oxygen species (ROS), nutrient and ATP sensing, DNA damage response (DDR), and epigenetic alterations. Mitochondria play a central role in these pathways not only through energetics and ATP production but also through metabolites generated in the tricarboxylic acid cycle, as well as mitochondria–nuclear signaling related to mitochondria morphology, biogenesis, fission/fusion, mitophagy, apoptosis, and epigenetic regulation.Objectives: We investigated the concept of bidirectional interactions between mitochondria and cellular pathways in response to environmental stress with a focus on epigenetic regulation, and we examined DNA repair and DDR pathways as examples of biological processes that respond to exogenous insults through changes in homeostasis and altered mitochondrial function.Methods: The National Institute of Environmental Health Sciences sponsored the Workshop on Mitochondria, Energetics, Epigenetics, Environment, and DNA Damage Response on 25–26 March 2013. Here, we summarize key points and ideas emerging from this meeting.Discussion: A more comprehensive understanding of signaling mechanisms (cross-talk) between the mitochondria and nucleus is central to elucidating the integration of mitochondrial functions with other cellular response pathways in modulating the effects of environmental agents. Recent studies have highlighted the importance of mitochondrial functions in epigenetic regulation and DDR with environmental stress. Development and application of novel technologies, enhanced experimental models, and a systems-type research approach will help to discern how environmentally induced mitochondrial dysfunction affects key mechanistic pathways.Conclusions: Understanding mitochondria–cell signaling will provide insight into individual responses to environmental hazards, improving prediction of hazard and susceptibility to environmental stressors.Citation: Shaughnessy DT, McAllister K, Worth L, Haugen AC, Meyer JN, Domann FE, Van Houten B, Mostoslavsky R, Bultman SJ, Baccarelli AA, Begley TJ, Sobol RW, Hirschey MD, Ideker T, Santos JH, Copeland WC, Tice RR, Balshaw DM, Tyson FL. 2014. Mitochondria, energetics, epigenetics, and cellular responses to stress. Environ Health Perspect 122:1271–1278; http://dx.doi.org/10.1289/ehp.1408418

Background:The term “exposome” was coined in 2005 to underscore the importance of the environment to human health and to bring research efforts in line with those on the human genome. The ability to characterize environmental exposures through biomonitoring is key to exposome research efforts.Objectives:Our objectives were to describe why traditional and nontraditional (exposomic) biomonitoring are both critical in studies aiming to capture the exposome and to make recommendations on how to transition exposure research toward exposomic approaches. We describe the biomonitoring needs of exposome research and approaches and recommendations that will help fill the gaps in the current science.Discussion:Traditional and exposomic biomonitoring approaches have key advantages and disadvantages for assessing exposure. Exposomic approaches differ from traditional biomonitoring methods in that they can include all exposures of potential health significance, whether from endogenous or exogenous sources. Issues of sample availability and quality, identification of unknown analytes, capture of nonpersistent chemicals, integration of methods, and statistical assessment of increasingly complex data sets remain challenges that must continue to be addressed.Conclusions:To understand the complexity of exposures faced throughout the lifespan, both traditional and nontraditional biomonitoring methods should be used. Through hybrid approaches and the integration of emerging techniques, biomonitoring strategies can be maximized in research to define the exposome.Citation:Dennis KK, Marder E, Balshaw DM, Cui Y, Lynes MA, Patti GJ, Rappaport SM, Shaughnessy DT, Vrijheid M, Barr DB. 2017. Biomonitoring in the era of the exposome. Environ Health Perspect 125:502–510; http://dx.doi.org/10.1289/EHP474

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