Spatial differences in stoichiometry of EGR phosphatase and Microtubule-associated Stress Protein 1 control root meristem activity during drought stress.

During moderate severity drought and low water potential (ψw) stress, poorly understood signaling mechanisms restrict both meristem cell division and subsequent cell expansion. We found that the Arabidopsis thaliana Clade E Growth-Regulating 2 (EGR2) protein phosphatase and Microtubule-Associated Stress Protein 1 (MASP1) differed in their stoichiometry of protein accumulation across the root meristem and had opposing effects on root meristem activity at low ψw. Ectopic MASP1 or EGR expression increased or decreased, respectively, root meristem size and root elongation during low ψw stress. This, along with the ability of phosphomimic MASP1 to overcome the EGR-mediated suppression of root meristem size and the observation that ectopic EGR expression had no effect on unstressed plants, indicated that during low ψw EGR activation and attenuation of MASP1 phosphorylation in their overlapping zone of expression determines root meristem size and activity. Ectopic EGR expression also decreased root cell size at low ψw. Conversely, both the egr1-1 egr2-1 and egr1-1 egr2-1 masp1-1 mutants had similarly increased root cell size but only egr1-1egr2-1 had increased cell division. These observations demonstrated that EGRs affect meristem activity via MASP1 but affect cell expansion via other mechanisms. Interestingly, EGR2 was highly expressed in the root cortex, a cell type important for growth regulation and environmental response.

Insertion of YFP at P5CS1 and AFL1 shows the potential, and potential complications, of gene tagging for functional analyses of stress-related proteins.

Crispr/CAS9-enabled homologous recombination to insert a tag in frame with an endogenous gene can circumvent difficulties such as context-dependent promoter activity that complicate analysis of gene expression and protein accumulation patterns. However, there have been few reports examining whether such gene targeting/gene tagging (GT) can alter expression of the target gene. The enzyme encoded by Δ1-pyrroline-5-carboxylate synthetase 1 (P5CS1) is key for stress-induced proline synthesis and drought resistance, yet its expression pattern and protein localisation have been difficult to assay. We used GT to insert YFP in frame with the 5' or 3' ends of the endogenous P5CS1 and At14a-Like 1 (AFL1) coding regions. Insertion at the 3' end of either gene generated homozygous lines with expression of the gene-YFP fusion indistinguishable from the wild type allele. However, for P5CS1 this occurred only after selfing and advancement to the T5 generation allowed initial homozygous lethality of the insertion to be overcome. Once this was done, the GT-generated P5CS1-YFP plants revealed new information about P5CS1 localisation and tissue-specific expression. In contrast, insertion of YFP at the 5' end of either gene blocked expression. The results demonstrate that GT can be useful for functional analyses of genes that are problematic to properly express by other means but also show that, in some cases, GT can disrupt expression of the target gene.

An Intelligent Water Monitoring IoT System for Ecological Environment and Smart Cities.

Global precipitation is becoming increasingly intense due to the extreme climate. Therefore, creating new technology to manage water resources is crucial. To create a sustainable urban and ecological environment, a water level and water quality control system implementing artificial intelligence is presented in this research. The proposed smart monitoring system consists of four sensors (two different liquid level sensors, a turbidity and pH sensor, and a water oxygen sensor), a control module (an MCU, a motor, a pump, and a drain), and a power and communication system (a solar panel, a battery, and a wireless communication module). The system focuses on low-cost Internet of Things (IoT) devices along with low power consumption and high precision. This proposal collects rainfall from the preceding 10 years in the application region as well as the region's meteorological bureau's weekly weather report and uses artificial intelligence to compute the appropriate water level. More importantly, the adoption of dynamic adjustment systems can reserve and modify water resources in the application region more efficiently. Compared to existing technologies, the measurement approach utilized in this study not only achieves cost savings exceeding 60% but also enhances water level measurement accuracy by over 15% through the successful implementation of water level calibration decisions utilizing multiple distinct sensors. Of greater significance, the dynamic adjustment systems proposed in this research offer the potential for conserving water resources by more than 15% in an effective manner. As a result, the adoption of this technology may efficiently reserve and distribute water resources for smart cities as well as reduce substantial losses caused by anomalous water resources, such as floods, droughts, and ecological concerns.

3D-bioprinted alginate-based bioink scaffolds with ß-tricalcium phosphate for bone regeneration applications.

Background/purpose: 3D-printed bone tissue engineering is becoming recognized as a key approach in dentistry for creating customized bone regeneration treatments fitting patients bone defects requirements. 3D bioprinting offers an innovative method to fabricate detailed 3D structures, closely emulating the native bone micro-environment and better bone regeneration. This study aimed to develop an 3D-bioprintable scaffold using a combination of alginate and ß-tricalcium phosphate (ß-TCP) with the Cellink® BioX printer, aiming to advance the field of tissue engineering. Materials and methods: The physical and biological properties of the resulting 3D-printed scaffolds were evaluated at 10 %, 12 %, and 15 % alginate combined with 10 % ß-TCP. The scaffolds were characterized through printability, swelling behavior, degradability, and element analysis. The biological assessment included cell viability, alkaline phosphatase (ALP) activity. Results: 10 % alginate/ß-TCP 3D printed at 25 °C scaffold demonstrated the optimal condition for printability, swelling capability, and degradability of cell growth and nutrient diffusion. Addition of ß-TCP particles significantly improved the 3D printed material viscosity over only alginate (P < 0.05). 10 % alginate/ß-TCP enhanced MG-63 cell's proliferation (P < 0.05) and alkaline phosphatase activity (P < 0.001). Conclusion: This study demonstrated in vitro that 10 % alginate/ß-TCP bioink characteristic for fabricating 3D acellular bioprinted scaffolds was the best approach. 10 % alginate/ß-TCP bioink 3D-printed scaffold exhibited superior physical properties and promoted enhanced cell viability and alkaline phosphatase activity, showing great potential for personalized bone regeneration treatments.