ABSTRACT
Long Interspersed Element-1 (LINE-1) is an oncogenic human retrotransposon that 'copies and pastes' DNA into new locations via reverse transcription. Given that enzymatically active LINE-1 can be exported in extracellular vesicles (EVs), and that LINE-1 mRNA and its two encoded proteins, ORF1p and ORF2p, are required for retrotransposition, the present study examined LINE-1 EV loading patterns relative to reverse transcriptase (RT) activity in vivo and in vitro. Density gradient ultracentrifugation identified conserved patterns of LINE-1 mRNA and protein distribution in EVs, with RT activity readily detected in EV fractions containing both LINE-1 mRNA and protein. Unlike whole cell and tissue lysates, the ORF1p in EVs was detected as a dimer. EVs from ostensibly healthy plasma donors showed variable but consistent ORF1p profiles, with residual levels of LINE-1 mRNA measured in some but not all samples. EVs from cancer cell lines had elevated mean LINE-1 levels and 5-85 times greater RT activity than EVs from normal cells or healthy plasma. EV RT activity was associated with EV LINE-1 mRNA content and was highest in cell lines that also expressed an elevated expression of ORF1p and ORF2p. Given that LINE-1 activation is a hallmark of many cancer types, our findings suggest that an EV LINE-1 'liquid biopsy' may be developed to monitor LINE-1 activity during the course of malignant progression.
Subject(s)
Extracellular Vesicles , Long Interspersed Nucleotide Elements , Lung Neoplasms , Endonucleases , Extracellular Vesicles/genetics , Extracellular Vesicles/metabolism , Humans , Lung Neoplasms/genetics , Proteins , RNA, Messenger/genetics , RNA-Directed DNA Polymerase/genetics , RNA-Directed DNA Polymerase/metabolism , Retroelements , Reverse TranscriptionABSTRACT
Efficient use of water, particularly in the western U.S., is an increasingly important aspect of many activities including agriculture, urban, and industry. As the population increases and agriculture and energy needs continue to rise, the pressure on water and other natural resources is expected to intensify. Recent advances in technology have stimulated growth in oil and gas development, as well as increasing the industry's need for water resources. This study provides an analysis of how efficiently water resources are used for unconventional shale development in Northeastern Colorado. The study is focused on the Wattenberg Field in the Denver-Julesberg Basin. The 2000 square mile field located in a semiarid climate with competing agriculture, municipal, and industrial water demands was one of the first fields where widespread use of hydraulic fracturing was implemented. The consumptive water intensity is measured using a ratio of the net water consumption and the net energy recovery and is used to measure how efficiently water is used for energy extraction. The water and energy use as well as energy recovery data were collected from 200 Noble Energy Inc. wells to estimate the consumptive water intensity. The consumptive water intensity of unconventional shale in the Wattenberg is compared with the consumptive water intensity for extraction of other fuels for other energy sources including coal, natural gas, oil, nuclear, and renewables. 1.4 to 7.5 million gallons is required to drill and hydraulically fracture horizontal wells before energy is extracted in the Wattenberg Field. However, when the large short-term total freshwater-water use is normalized to the amount of energy produced over the lifespan of a well, the consumptive water intensity is estimated to be between 1.8 and 2.7 gal/MMBtu and is similar to surface coal mining.
