Project description:The circadian profile of the transcriptomes in murine tissues was compared between groups of young and old male mice. The oscillatory phase and absolute expression levels were evaluated as a function of biological age. Distinct differences in multiple pathways were apparent based on biological and circadian time in a tissue specific manner. A total of 18 young (5 month) and old (24 month) C57BL/6 mice were acclimated to a set 12 hr light dark cycle and fed ad lib on standard lab chow for 2 weeks in the PBRC Comparative Biology Core Facility. Five days prior to the study, the mice were converted to constant darkness (red light) regimen. On the day of the study, mice were euthanized by CO2 asphyxiation after ad lib feeding overnight in groups of 3 animals per young or old cohort at 4 hr intervals beginning at 7 AM and ending at 3 AM on a single day (July, 2009). The body weight of each animal was recorded prior to dissection of the following tissues: brown adipose tissue (BAT), inguinal white adipose tissue (iWAT), and liver. Individual tissues were weighed prior to freezing in liquid nitrogen. Samples were stored at -80oC until use.Total RNA was isolated from the BAT, eWAT, and liver tissues using TriReagent (MRC, Cincinnati OH) in accordance with the manufacturer’s recommendations. The Illumina TotalPrep RNA Amplification Kit (Applied Biosystems Inc., Foster City, CA, Catalog #AMIL1791) was used to create labeled cRNA from 750ng of input total RNA according to the manufacturer’s protocol. The labeled cRNA samples were then assessed for quality and quantity using a NanoDrop and an Agilent Bioanalyzer. The MouseWG-6 v2 Beadchip (Illumina Sentrix Beadchip Array #11278593) contains 45,200 transcripts and allows 6 samples to be interrogated in parallel. 1.5ug of each labeled cRNA was hybridized to each array according to the manufacturer's protocol. Experimental group samples were distributed randomly across all beadchips. After an 18 hour hybridization at 58°C, the beadchips were processed according to manufacturer’s protocol and scanned using an Illumina BeadArray Reader (Illumina, Inc., San Diego, CA).

Project description:The efficacy of auxinic herbicides, a valuable weed control tool for growers worldwide, has been shown to vary with the time of day in which applications are made. However, little is known about the mechanisms causing this phenomenon. Investigating the differential in planta behavior of these herbicides across different times of application may grant an ability to advise which properties of auxinic herbicides are desirable when applications must be made around the clock. Radiolabeled herbicide experiments demonstrated a likely increase in ATP-binding cassette subfamily B (ABCB)-mediated 2,4-D and dicamba transport in Palmer amaranth (Amaranthus palmeri S. Watson) at simulated dawn compared to mid-day, as dose response models indicated that many orders of magnitude higher concentrations of N-1-naphthylphthalamic acid (NPA) and verapamil, respectively, are required to inhibit translocation by 50% at simulated sunrise compared to mid-day. Gas chromatographic analysis displayed that ethylene evolution in A. palmeri was higher when dicamba was applied during mid-day compared to sunrise. Furthermore, it was found that inhibition of translocation via 2,3,5-triiodobenzoic acid (TIBA) resulted in an increased amount of 2,4-D-induced ethylene evolution at sunrise, and the inhibition of dicamba translocation via NPA reversed the difference in ethylene evolution across time of application. Dawn applications of these herbicides were associated with increased expression of a putative 9-cis-epoxycarotenoid dioxygenase biosynthesis gene NCED1, while there was a notable lack of trends observed across times of day and across herbicides with ACS1, encoding 1-aminocyclopropane-1-carboxylic acid synthase. Overall, this research indicates that translocation is differentially regulated via specific protein-level mechanisms across times of application, and that ethylene release, a chief phytotoxic process involved in the response to auxinic herbicides, is related to translocation. Furthermore, transcriptional regulation of abscisic acid involvement in phytotoxicity and/or translocation are suggested.

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Cellular and physiological circadian mechanisms drive diurnal cell proliferation and expansion of white adipose tissue.

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