A Non-Canonical E-Box Within the \u3cem\u3eMyoD\u3c/em\u3e Core Enhancer is Necessary for Circadian Expression in Skeletal Muscle

The myogenic differentiation 1 (MyoD) gene is a master regulator of myogenesis. We previously reported that the expression of MyoD mRNA oscillates over 24 h in skeletal muscle and that the circadian clock transcription factors, BMAL1 (brain and muscle ARNT-like 1) and CLOCK (circadian locomotor output cycles kaput), were bound to the core enhancer (CE) of the MyoD gene in vivo. In this study, we provide in vivo and in vitro evidence that the CE is necessary for circadian expression of MyoD in adult muscle. Gel shift assays identified a conserved non-canonical E-box within the CE that is bound by CLOCK and BMAL1. Functional analysis revealed that this E-box was required for full activation by BMAL1/CLOCK and for in vitro circadian oscillation. Expression profiling of muscle of CEloxP/loxP mice found approximately 1300 genes mis-expressed relative to wild-type. Based on the informatics results, we analyzed the respiratory function of mitochondria isolated from wild-type and CEloxP/loxP mice. These assays determined that State 5 respiration was significantly reduced in CEloxP/loxP muscle. The results of this work identify a novel element in the MyoD enhancer that confers circadian regulation to MyoD in skeletal muscle and suggest that loss of circadian regulation leads to changes in myogenic expression and downstream mitochondrial function

Regulated deficit irrigation reduces water use of almonds without affecting yield

A plant-based regulated deficit irrigation (RDI) experiment in the northern Sacramento Valley determined that crop consumptive water use and irrigation could be reduced without significant detrimental effects on almond production. Tree stress was measured by recording midday stem water potential, a direct measure of tree water stress. With a water stress level of −14 to −18 bars during the hull-split period, average annual water savings were about 5 inches. Over 5 years, no significant yield reductions were observed, although average kernel weight was slightly lower. The results suggest that water savings can be achieved without affecting yield, even in soils with low water-holding capacity

Dissecting dual roles of MyoD during lineage conversion to mature myocytes and myogenic stem cells

The generation of myotubes from fibroblasts upon forced MyoD expression is a classic example of transcription factor-induced reprogramming. We recently discovered that additional modulation of signaling pathways with small molecules facilitates reprogramming to more primitive induced myogenic progenitor cells (iMPCs). Here, we dissected the transcriptional and epigenetic dynamics of mouse fibroblasts undergoing reprogramming to either myotubes or iMPCs using a MyoD-inducible transgenic model. Induction of MyoD in fibroblasts combined with small molecules generated Pax7+ iMPCs with high similarity to primary muscle stem cells. Analysis of intermediate stages of iMPC induction revealed that extinction of the fibroblast program preceded induction of the stem cell program. Moreover, key stem cell genes gained chromatin accessibility prior to their transcriptional activation, and these regions exhibited a marked loss of DNA methylation dependent on the Tet enzymes. In contrast, myotube generation was associated with few methylation changes, incomplete and unstable reprogramming, and an insensitivity to Tet depletion. Finally, we showed that MyoD's ability to bind to unique bHLH targets was crucial for generating iMPCs but dispensable for generating myotubes. Collectively, our analyses elucidate the role of MyoD in myogenic reprogramming and derive general principles by which transcription factors and signaling pathways cooperate to rewire cell identity

A non-canonical E-box within the MyoD core enhancer is necessary for circadian expression in skeletal muscle

The myogenic differentiation 1 (MyoD) gene is a master regulator of myogenesis. We previously reported that the expression of MyoD mRNA oscillates over 24 h in skeletal muscle and that the circadian clock transcription factors, BMAL1 (brain and muscle ARNT-like 1) and CLOCK (circadian locomotor output cycles kaput), were bound to the core enhancer (CE) of the MyoD gene in vivo. In this study, we provide in vivo and in vitro evidence that the CE is necessary for circadian expression of MyoD in adult muscle. Gel shift assays identified a conserved non-canonical E-box within the CE that is bound by CLOCK and BMAL1. Functional analysis revealed that this E-box was required for full activation by BMAL1/CLOCK and for in vitro circadian oscillation. Expression profiling of muscle of CEloxP/loxP mice found approximately 1300 genes mis-expressed relative to wild-type. Based on the informatics results, we analyzed the respiratory function of mitochondria isolated from wild-type and CEloxP/loxP mice. These assays determined that State 5 respiration was significantly reduced in CEloxP/loxP muscle. The results of this work identify a novel element in the MyoD enhancer that confers circadian regulation to MyoD in skeletal muscle and suggest that loss of circadian regulation leads to changes in myogenic expression and downstream mitochondrial function

Observing response processes with eye tracking in international large-scale assessments: evidence from the OECD PIAAC assessment

This paper reports on a pilot study that used eye tracking techniques to make detailed observations of item response processes in the OECD Programme for the International Assessment of Adult Competencies (PIAAC). The lab-based study also recorded physiological responses using measures of pupil diameter and electrodermal activity. The study tested 14 adult respondents as they individually completed the PIAAC computer-based assessment. The eye tracking observations help to fill an ‘explanatory gap’ by providing data on variation in item response processes that are not captured by other sources of process data such as think aloud protocols or computer-generated log files. The data on fixations and saccades provided detailed information on test item response strategies, enabling profiling of respondent engagement and response processes associated with successful performance. Much of that activity does not include the use of the keyboard and mouse, and involves ‘off-screen’ use of pen and paper (and calculator) that are not captured by assessment log-files. In conclusion, this paper points toward an important application of eye tracking in large-scale assessments. This includes insights into response processes in new domains such as adaptive problem-solving that aim to identify individuals’ ability to select and combine resources from the digital and physical environment

A regulated deficit irrigation strategy for hedgerow olive orchards with high plant density

Background & Aims There is not a consensus on the best irrigation approach for super-high density (SHD) olive orchards. Our aim was to design and test a regulated deficit irrigation (RDI) strategy for a sustainable balance between water saving, tree vigour and oil production. Methods We tested our RDI strategy for 3 years in an ‘Arbequina’ orchard with 1,667 trees ha−1. Two levels of irrigation reduction were applied, 60RDI and 30RDI, scaled to replacing 60 % and 30 %, respectively, of the of irrigation needs (IN). We also had a full irrigation (FI) treatment as control, with IN totalling 4,701 m3 ha−1 Results The 30RDI treatment showed the best balance between water saving, tree vigour and oil production. With a yearly irrigation amount (IA) of 1,366 m3 ha−1, which meant 72 % water saving as compared to FI, the reduction in oil yield was 26 % only. Conclusions Our results, together with recent knowledge on the effect of water stress on fruit development, allowed us to suggest a potentially improved RDI strategy for which a total IA of ca. 2,100 m3 ha−1 was calculated. Both some management details and the benefits of this suggested RDI strategy are still to be tested

The Msx1 Homeoprotein Recruits G9a Methyltransferase to Repressed Target Genes in Myoblast Cells

Although the significance of lysine modifications of core histones for regulating gene expression is widely appreciated, the mechanisms by which these modifications are incorporated at specific regulatory elements during cellular differentiation remains largely unknown. In our previous studies, we have shown that in developing myoblasts the Msx1 homeoprotein represses gene expression by influencing the modification status of chromatin at its target genes. We now show that genomic binding by Msx1 promotes enrichment of the H3K9me2 mark on repressed target genes via recruitment of G9a histone methyltransferase, the enzyme responsible for catalyzing this histone mark. Interaction of Msx1 with G9a is mediated via the homeodomain and is required for transcriptional repression and regulation of cellular differentiation, as well as enrichment of the H3K9me2 mark in proximity to Msx1 binding sites on repressed target genes in myoblast cells as well as the developing limb. We propose that regulation of chromatin status by Msx1 recruitment of G9a and other histone modifying enzymes to regulatory regions of target genes represents an important means of regulating the gene expression during development

The effects of applied water at various fractions of measured evapotranspiration on reproductive growth and water productivity of Thompson Seedless grapevines

The reproductive growth and water productivity (WPb) of Thompson Seedless grapevines were measured as a function of applied water amounts at various fractions of measured grapevine ETc for a total of eight irrigation treatments. Shoots were harvested numerous times during the growing season to calculate water productivity. Berry weight was maximized at the 0.6–0.8 applied water treatments across years. As applied water amounts increased soluble solids decreased. Berry weight measured at veraison and harvest was a linear function of the mean midday leaf water potential measured between anthesis and veraison and anthesis and harvest, respectively. As applied water amounts increased up to the 0.6–0.8 irrigation treatments there was a significant linear increase in yield. Yields at greater applied water amounts either leveled off or decreased. The reduction in yield on either side of the yearly maximum was due to fewer numbers of clusters per vine. Maximum yield occurred at an ETc ranging from 550 to 700 mm. Yield per unit applied water and WPb increased as applied water decreased. The results from this study demonstrated that Thompson Seedless grapevines can be deficit irrigated, increasing water use efficiency while maximizing yields