Loss of tuberin, the product of TSC2 gene, increases mammalian target of rapamycin (mTOR) signaling, promoting cell growth and tumor development. However, in cells expressing tuberin, it is not known how repression of mTOR signaling is relieved to activate this pathway in response to growth factors and how hamartin participates in this process. We show that hamartin colocalizes with hypophosphorylated tuberin at the membrane, where tuberin exerts its GTPase-activating protein (GAP) activity to repress Rheb signaling. In response to growth signals, tuberin is phosphorylated by AKT and translocates to the cytosol, relieving Rheb repression. Phosphorylation of tuberin at serines 939 and 981 does not alter its intrinsic GAP activity toward Rheb but partitions tuberin to the cytosol, where it is bound by 14-3-3 proteins. Thus, tuberin bound by 14-3-3 in response to AKT phosphorylation is sequestered away from its membrane-bound activation partner (hamartin) and its target GTPase (Rheb) to relieve the growth inhibitory effects of this tumor suppressor.

The phytohormone salicylic acid (SA) plays essential roles in biotic and abiotic responses, plant development, and leaf senescence. 2,5-Dihydroxybenzoic acid (2,5-DHBA or gentisic acid) is one of the most commonly occurring aromatic acids in green plants and is assumed to be generated from SA, but the enzymes involved in its production remain obscure. DMR6 (Downy Mildew Resistant6; At5g24530) has been proven essential in plant immunity of Arabidopsis (Arabidopsis thaliana), but its biochemical properties are not well understood. Here, we report the discovery and functional characterization of DMR6 as a salicylic acid 5-hydroxylase (S5H) that catalyzes the formation of 2,5-DHBA by hydroxylating SA at the C5 position of its phenyl ring in Arabidopsis. S5H/DMR6 specifically converts SA to 2,5-DHBA in vitro and displays higher catalytic efficiency (K cat /K m = 4. ) for SA. Interestingly, S5H/DMR6 displays a substrate inhibition property that may enable automatic control of its enzyme activities. The s5h mutant and s5hs3h double mutant overaccumulate SA and display phenotypes such as a smaller growth size, early senescence, and a loss of susceptibility to Pseudomonas syringae pv tomato DC3000. S5H/DMR6 is sensitively induced by SA/pathogen treatment and is expressed widely from young seedlings to senescing plants, whereas S3H is more specifically expressed at the mature and senescing stages. Collectively, our results disclose the identity of the enzyme required for 2,5-DHBA formation and reveal a mechanism by which plants fine-tune SA homeostasis by mediating SA 5-hydroxylation.

Chromatin structure is known to be a barrier to DNA repair and a large number of studies have now identified various factors that modify histones and remodel nucleosomes to facilitate repair. In response to ultraviolet (UV) radiation several histones are acetylated and this enhances the repair of DNA photoproducts by the nucleotide excision repair (NER) pathway. However, the molecular mechanism by which UV radiation induces histone acetylation to allow for efficient NER is not completely understood. We recently discovered that the E2F1 transcription factor accumulates at sites of UV-induced DNA damage and directly stimulates NER through a non-transcriptional mechanism. Here we demonstrate that E2F1 associates with the GCN5 acetyltransferase in response to UV radiation and recruits GCN5 to sites of damage. UV radiation induces the acetylation of histone H3 lysine 9 (H3K9) and this requires both GCN5 and E2F1. Moreover, as previously observed for E2F1, knock down of GCN5 results in impaired recruitment of NER factors to sites of damage and inefficient DNA repair. These findings demonstrate a direct role for GCN5 and E2F1 in NER involving H3K9 acetylation and increased accessibility to the NER machinery.

Eight commercially available organic Zn products and reagent-grade ZnSO4 x 7H2O (Zn Sulf) were evaluated by polarographic analysis, and solubility in .1 M K2HPO4-KH2PO4 buffer (pH 5), .2 M HCl-KCl buffer (pH 2), and deionized water. Fractions from these solubility tests were evaluated by gel filtration chromatography for structural integrity. Degree of chelation was generally positively related to chelation effectiveness determined by polarography. The organic sources were Zn methionine complex A (Zn MetA), Zn methionine complex B (Zn MetB), Zn polysaccharide complex (Zn Poly), Zn lysine complex (Zn Lys), Zn amino acid chelate (Zn AA), Zn proteinate A (Zn ProA), Zn proteinate B (Zn ProB), and Zn proteinate C (Zn ProC). Three experiments were conducted to estimate the relative bioavailability of Zn from the organic Zn supplements for chicks and lambs when added at high dietary levels to practical diets. Bone Zn concentration increased (P < .001) as dietary Zn increased in both experiments. When Zn Sulf was assigned a value of 100% as the standard, multiple linear regression slope ratios of bone Zn from chicks fed 3 wk regressed on dietary Zn intake gave estimated relative bioavailability values of 83 +/- 14.6 and 139 +/- 16.9 for Zn AA and Zn ProA, respectively, in Exp. 1 and 94 +/- 11.6, 99 +/- 8.8, and 108 +/- 11.4 for Zn Poly, Zn ProB, and Zn ProC, respectively, in Exp. 2. In Exp. 3, 42 lambs were fed diets containing Zn Sulf, Zn ProA, Zn AA, or Zn MetB for 21 d. Based on multiple linear regression slope ratios of liver, kidney, and pancreas Zn and liver metallothionein concentrations on added dietary Zn, bioavailability estimates relative to 100% for Zn Sulf were 130, 110, and 113 for Zn ProA, Zn AA, and Zn MetB, respectively. Except for Zn ProA, which was greater, the organic Zn supplements had bioavailability values similar to that of Zn Sulf for chicks and lambs. Bioavailability of organic Zn products was inversely related to solubility of Zn in pH 5 buffer in chicks (r2 = .91) and pH 2 buffer in lambs (r2 = .91), but not to an estimate of degree of chelation.

The levels of microRNAs (miRNAs) are altered under different conditions such as cancer, senescence, and aging. Here, we have identified differentially expressed miRNAs in skeletal muscle from young and old rhesus monkeys using RNA sequencing. In old muscle, several miRNAs were upregulated, including miR-451, miR-144, miR-18a and miR-15a, while a few miRNAs were downregulated, including miR-181a and miR-181b. A number of novel miRNAs were also identified, particularly in old muscle. We also examined the impact of caloric restriction (CR) on miRNA abundance by reverse transcription (RT) followed by real-time, quantitative (q)PCR analysis and found that CR rescued the levels of miR-181b and chr1:205580546, and also dampened the age-induced increase in miR-451 and miR-144 levels. Our results reveal that there are changes in expression of known and novel miRNAs with skeletal muscle aging and that CR may reverse some of these changes to a younger phenotype.