Spatio-temporal transcript profiling of rice roots and shoots in response to phosphate starvation and recovery.

Using rice (Oryza sativa) as a model crop species, we performed an in-depth temporal transcriptome analysis, covering the early and late stages of Pi deprivation as well as Pi recovery in roots and shoots, using next-generation sequencing. Analyses of 126 paired-end RNA sequencing libraries, spanning nine time points, provided a comprehensive overview of the dynamic responses of rice to Pi stress. Differentially expressed genes were grouped into eight sets based on their responses to Pi starvation and recovery, enabling the complex signaling pathways involved in Pi homeostasis to be untangled. A reference annotation-based transcript assembly was also generated, identifying 438 unannotated loci that were differentially expressed under Pi starvation. Several genes also showed induction of unannotated splice isoforms under Pi starvation. Among these, PHOSPHATE2 (PHO2), a key regulator of Pi homeostasis, displayed a Pi starvation-induced isoform, which was associated with increased translation activity. In addition, microRNA (miRNA) expression profiles after long-term Pi starvation in roots and shoots were assessed, identifying 20 miRNA families that were not previously associated with Pi starvation, such as miR6250. In this article, we present a comprehensive spatio-temporal transcriptome analysis of plant responses to Pi stress, revealing a large number of potential key regulators of Pi homeostasis in plants.

A membrane-bound NAC transcription factor, ANAC017, mediates mitochondrial retrograde signaling in Arabidopsis.

Plants require daily coordinated regulation of energy metabolism for optimal growth and survival and therefore need to integrate cellular responses with both mitochondrial and plastid retrograde signaling. Using a forward genetic screen to characterize regulators of alternative oxidase1a (rao) mutants, we identified RAO2/Arabidopsis NAC domain-containing protein17 (ANAC017) as a direct positive regulator of AOX1a. RAO2/ANAC017 is targeted to connections and junctions in the endoplasmic reticulum (ER) and F-actin via a C-terminal transmembrane (TM) domain. A consensus rhomboid protease cleavage site is present in ANAC017 just prior to the predicted TM domain. Furthermore, addition of the rhomboid protease inhibitor N-p-Tosyl-l-Phe chloromethyl abolishes the induction of AOX1a upon antimycin A treatment. Simultaneous fluorescent tagging of ANAC017 with N-terminal red fluorescent protein (RFP) and C-terminal green fluorescent protein (GFP) revealed that the N-terminal RFP domain migrated into the nucleus, while the C-terminal GFP tag remained in the ER. Genome-wide analysis of the transcriptional network regulated by RAO2/ANAC017 under stress treatment revealed that RAO2/ANAC017 function was necessary for >85% of the changes observed as a primary response to cytosolic hydrogen peroxide (H2O2), but only ~33% of transcriptional changes observed in response to antimycin A treatment. Plants with mutated rao2/anac017 were more stress sensitive, whereas a gain-of-function mutation resulted in plants that had lower cellular levels of H2O2 under untreated conditions.

Cyclin-dependent kinase E1 (CDKE1) provides a cellular switch in plants between growth and stress responses.

Plants must deal effectively with unfavorable growth conditions that necessitate a coordinated response to integrate cellular signals with mitochondrial retrograde signals. A genetic screen was carried out to identify regulators of alternative oxidase (rao mutants), using AOX1a expression as a model system to study retrograde signaling in plants. Two independent rao1 mutant alleles identified CDKE1 as a central nuclear component integrating mitochondrial retrograde signals with energy signals under stress. CDKE1 is also necessary for responses to general cellular stresses, such as H(2)O(2) and cold that act, at least in part, via anterograde pathways, and integrates signals from central energy/stress sensing kinase signal transduction pathways within the nucleus. Together, these results place CDKE1 as a central kinase integrating diverse cellular signals and shed light on a mechanism by which plants can effectively switch between growth and stress responses.

Effects of Unilateral Neuromuscular Electrical Stimulation with Illusionary Mirror Visual Feedback on the Contralateral Muscle: A Pilot Study.

We aim to examine the cross-education effects of unilateral muscle neuromuscular electrical stimulation (NMES) training combined with illusionary mirror visual feedback (MVF). Fifteen adults (NMES + MVF: 5; NMES: 5, Control: 5) completed this study. The experimental groups completed a 3-week NMES training on their dominant elbow flexor muscle. The NMES + MVF group had a mirror placed in the midsagittal plane between their upper arms, so a visual illusion was created in which their non-dominant arms appeared to be stimulated. Baseline and post-training measurements included both arms' isometric strength, voluntary activation level, and resting twitch. Cross-education effects were not observed from all dependent variables. For the unilateral muscle, both experimental groups showed greater strength increases when compared to the control (isometric strength % changes: NMES + MVF vs. NMES vs. Control = 6.31 ± 4.56% vs. 4.72 ± 8.97% vs. -4.04 ± 3.85%, p < 0.05). Throughout the training, even with the maximally tolerated NMES, the NMES + MVF group had greater perceived exertion and discomfort than the NMES. Additionally, the NMES-evoked force increased throughout the training for both groups. Our data does not support that NMES combined with or without MVF induces cross-education. However, the stimulated muscle becomes more responsive to the NMES and can become stronger following the training.

Wide-Pulse High-Frequency Neuromuscular Electrical Stimulation Evokes Greater Relative Force in Women Than in Men: A Pilot Study.

This study aimed to examine the potential sex differences in wide-pulse high-frequency neuromuscular electrical stimulation (WPHF NMES)-evoked force. Twenty-two subjects (10 women) completed this study. Prior to the stimulation, the visual analogue scale (VAS) for discomfort and the rating of perceived exertion (RPE) were measured, followed by the isometric strength of the dominant elbow flexor muscles. The subjects then completed ten, 10-s on 10-s off WPHF NMES (pulse width: 1 ms, frequency: 100 Hz) at maximum tolerable intensities. The subjects' RPE was recorded after each set, and the VAS was measured following the last stimulation. The stimulation induced significant increase in discomfort for both sexes, with women having greater discomfort than men (men: 22.4 ± 14.9 mm, women: 39.7 ± 12.7 mm). The stimulation amplitude was significantly greater in men than in women (men: 16.2 ± 6.3 mA, women: 12.0 ± 4.5 mA). For the evoked force, only the relative NMES-evoked force was found greater in women than in men (men: 8.96 ± 6.51%, women: 17.08 ± 12.61%). In conclusion, even at the maximum tolerable intensity, WPHF NMES evoked larger relative elbow flexion force in women than in men, with women experiencing greater discomfort.