Image-Based Quantification of Arabidopsis thaliana Stomatal Aperture from Leaf Images.

The quantification of stomatal pore size has long been a fundamental approach to understand the physiological response of plants in the context of environmental adaptation. Automation of such methodologies not only alleviates human labor and bias but also realizes new experimental research methods through massive analysis. Here, we present an image analysis pipeline that automatically quantifies stomatal aperture of Arabidopsis thaliana leaves from bright-field microscopy images containing mesophyll tissue as noisy backgrounds. By combining a You Only Look Once X-based stomatal detection submodule and a U-Net-based pore segmentation submodule, we achieved a mean average precision with an intersection of union (IoU) threshold of 50% value of 0.875 (stomata detection performance) and an IoU of 0.745 (pore segmentation performance) against images of leaf discs taken with a bright-field microscope. Moreover, we designed a portable imaging device that allows easy acquisition of stomatal images from detached/undetached intact leaves on-site. We demonstrated that this device in combination with fine-tuned models of the pipeline we generated here provides robust measurements that can substitute for manual measurement of stomatal responses against pathogen inoculation. Utilization of our hardware and pipeline for automated stomatal aperture measurements is expected to accelerate research on stomatal biology of model dicots.

Quercetin and HSC70 coregulate the anti-inflammatory action of the ubiquitin-like protein MNSFß.

BACKGROUND: Quercetin is a flavonol that modifies many cellular processes. Monoclonal nonspecific suppressor factor ß is a member of the ubiquitin-like family of proteins that are involved in various biological processes. It has been demonstrated that quercetin regulates the effect of MNSFß on tumor necrosis factor-α secretion in lipopolysaccharide (LPS)-stimulated macrophages. This study found that quercetin and the heat shock protein HSC70 coregulate the action of MNSFß. METHODS AND RESULTS: Quercetin dose-dependently suppressed the LPS/interferon γ-induced nitric oxide production without cytotoxicity in the macrophage-like cell line Raw264.7. SiRNA knockdown experiments showed that quercetin inhibited the MNSFß and HSC70 siRNA-mediated enhancement of TNFα and the production of RANTES, a member of C-C chemokine superfamily, in LPS-stimulated Raw264.7 cells. Western blot analysis showed that quercetin and HSC70 regulated ERK1/2 activation and LPS-stimulated IκBα degradation by affecting the complex formation of MNSFß and the proapoptotic protein Bcl-G. Moreover, MNSFß is implicated in TLR4/MyD88 signaling but not in TLR3 signaling. CONCLUSIONS: HSC70 is an important chaperone that facilitates the stabilization of MNSFß. Quercetin may negatively control the function of MNSFß by regulating the action of the molecular chaperone HSC70. MNSFß mediates TLR4/Myd88 signaling but not TLR3 signaling.

Ubiquitin-like protein MNSFß regulates glycolysis and promotes cell proliferation with HSC70 assistance.

Monoclonal non-specific suppressor factor ß (MNSFß) is a universally expressed ubiquitin-like protein that has multiple biological functions. MNSFß modifies its target molecules through covalent conjugation. Most recently, we identified a molecular chaperone, HSC70, that facilitates the stabilization of aggregable MNSFß. In the current study, we determined the role of HSC70 in stabilizing unstable MNSFß. HSC70 promoted the correct folding of MNSFß both in vitro and in vivo. We also examined the regulatory function of MNSFß in cell proliferation and glycolysis. MNSFß siRNA and HSC70 siRNA treatment attenuated lactate release from Raw264.7 macrophage-like cells. MNSFß siRNA inhibited glucose uptake in Raw264.7 cells. We found that glucose transporter 1 (GLUT1) is an important membrane protein involved in the regulatory function of MNSFß during glycolysis. MNSFß siRNA inhibited the increased GLUT1 expression in LPS-stimulated cells, suggesting that MNSFß controls the inflammatory response through GLUT1 regulation. We identified several important molecules, including lactate dehydrogenase A, which are regulated by MNSFß and involved in glucose metabolism. Here we firstly report that MNSFß regulates glycolysis and promotes cell proliferation.

Draft Genome Sequence of the Thermophilic Unicellular Cyanobacterium Synechococcus sp. Strain C9.

This study presents the genome sequence of Synechococcus sp. strain C9 (= CCMEE 5213 = ATCC 700244), a thermophilic unicellular cyanobacterium that was originally isolated from a thermal pool at Octopus Spring, Yellowstone National Park, USA. The genome consists of a 2,958,309-bp chromosome with a GC content of 52.9% and 2,854 protein-coding sequences.