Specific but interdependent functions for Arabidopsis AGO4 and AGO6 in RNA-directed DNA methylation.

Argonaute (AGO) family proteins are conserved key components of small RNA-induced silencing pathways. In the RNA-directed DNA methylation (RdDM) pathway in Arabidopsis, AGO6 is generally considered to be redundant with AGO4. In this report, our comprehensive, genomewide analyses of AGO4- and AGO6-dependent DNA methylation revealed that redundancy is unexpectedly negligible in the genetic interactions between AGO4 and AGO6. Immunofluorescence revealed that AGO4 and AGO6 differ in their subnuclear co-localization with RNA polymerases required for RdDM. Pol II and AGO6 are absent from perinucleolar foci, where Pol V and AGO4 are co-localized. In the nucleoplasm, AGO4 displays a strong co-localization with Pol II, whereas AGO6 co-localizes with Pol V. These patterns suggest that RdDM is mediated by distinct, spatially regulated combinations of AGO proteins and RNA polymerases. Consistently, Pol II physically interacts with AGO4 but not AGO6, and the levels of Pol V-dependent scaffold RNAs and Pol V chromatin occupancy are strongly correlated with AGO6 but not AGO4. Our results suggest that AGO4 and AGO6 mainly act sequentially in mediating small RNA-directed DNA methylation.

Effects of five years conservation tillage for hedging against drought, stabilizing maize yield, and improving soil environment in the drylands of northern China.

Continuous tillage cultivation positioning trials can provide the basis for maintaining soil health, improving resource utilization efficiency and crop productivity, and achieving sustainable agricultural development. In this study, changes in soil stability and water-holding capacity characteristics were measured under different tillage cultivations from a multi-year microscopic perspective and analyzed to evaluate selected key indicators. Continuous monitoring of rainfall utilization efficiency and yield was carried out for five years. Here, we discuss the role of conservation tillage in buffering and stabilizing rainfall precipitation pattern on the fluctuation and uncertainty of soil water retention and water supply capacity and soil quality. The study was carried out on dryland areas of the Loess Plateau in northern China with eight tillage systems established in 2016: no-tillage (NT); no-tillage and straw (NTS); subsoiling (SU); subsoiling and straw (SUS); rotary tillage (RT); rotary tillage and straw (RTS); conventional tillage (CT); and conventional tillage and straw (CTS). All treatments were applied in conjunction with continuous cropping for five years. The evaluated soil parameters were mean weight diameter (MWD), geometric mean diameter (GMD), >0.25 mm aggregate content (R0.25) of water-stable aggregates (WSAs), soil moisture characteristic curve (SMCC), specific soil water capacity (Cθ), soil organic matter (SOM), rainfall utilization efficiency (RUE), and maize yields for five consecutive years. The MWD, GMD, and R0.25 of SUS were 27.38%, 17.57%, and 7.68% more than CTS (control), respectively. Overall, SOM, average annual RUE, and average annual yields increased by 14.64%, 11.89%, and 9.59%, respectively, compared with 2016. Our results strongly suggest that conservation tillage can considerably improve these characterization indicators. SUS was more effective than CTS in the 0-40 cm soil layer at hedging against drought in the area, stabilizing crop production, and achieving sustainable agricultural development.

Measuring spatial and temporal Ca2+ signals in Arabidopsis plants.

Developmental and environmental cues induce Ca(2+) fluctuations in plant cells. Stimulus-specific spatial-temporal Ca(2+) patterns are sensed by cellular Ca(2+) binding proteins that initiate Ca(2+) signaling cascades. However, we still know little about how stimulus specific Ca(2+) signals are generated. The specificity of a Ca(2+) signal may be attributed to the sophisticated regulation of the activities of Ca(2+) channels and/or transporters in response to a given stimulus. To identify these cellular components and understand their functions, it is crucial to use systems that allow a sensitive and robust recording of Ca(2+) signals at both the tissue and cellular levels. Genetically encoded Ca(2+) indicators that are targeted to different cellular compartments have provided a platform for live cell confocal imaging of cellular Ca(2+) signals. Here we describe instructions for the use of two Ca(2+) detection systems: aequorin based FAS (film adhesive seedlings) luminescence Ca(2+) imaging and case12 based live cell confocal fluorescence Ca(2+) imaging. Luminescence imaging using the FAS system provides a simple, robust and sensitive detection of spatial and temporal Ca(2+) signals at the tissue level, while live cell confocal imaging using Case12 provides simultaneous detection of cytosolic and nuclear Ca(2+) signals at a high resolution.

Aequorin-based luminescence imaging reveals stimulus- and tissue-specific Ca2+ dynamics in Arabidopsis plants.

Calcium ion is a versatile second messenger for diverse cell signaling in response to developmental and environmental cues. The specificity of Ca(2+)-mediated signaling is defined by stimulus-elicited Ca(2+) signature and downstream decoding processes. Here, an Aequorin-based luminescence recording system was developed for monitoring Ca(2+) in response to various stimuli in Arabidopsis. With the simple, highly sensitive, and robust Ca(2+) recording, this system revealed stimulus- and tissue-specific Ca(2+) signatures in seedlings. Cellular Ca(2+) dynamics and relationship to Aequorin-based Ca(2+) recording were explored using a GFP-based Ca(2+) indicator, which suggested that a synchronous cellular Ca(2+) signal is responsible for cold-induced Ca(2+) response in seedlings, whereas asynchronous Ca(2+) oscillation contributes to osmotic stress-induced Ca(2+) increase in seedlings. The optimized recording system would be a powerful tool for the identification and characterization of novel components in Ca(2+)-mediated stress-signaling pathways.