Fluorescence intensity and lifetime imaging of lipofuscin-like autofluorescence for label-free predicting clinical drug response in cancer.

Conventional techniques for in vitro cancer drug screening require labor-intensive formalin fixation, paraffin embedding, and dye staining of tumor tissues at fixed endpoints. This way of assessment discards the valuable pharmacodynamic information in live cells over time. Here, we found endogenous lipofuscin-like autofluorescence acutely accumulated in the cell death process. Its unique red autofluorescence could report the apoptosis without labeling and continuously monitor the treatment responses in 3D tumor-culture models. Lifetime imaging of lipofuscin-like red autofluorescence could further distinguish necrosis from apoptosis of cells. Moreover, this endogenous fluorescent marker could visualize the apoptosis in live zebrafish embryos during development. Overall, this study validates that lipofuscin-like autofluorophore is a generic cell death marker. Its characteristic autofluorescence could label-free predict the efficacy of anti-cancer drugs in organoids or animal models.

The Role of Neuropeptide-Stimulated cAMP-EPACs Signalling in Cancer Cells.

Neuropeptides are autocrine and paracrine signalling factors and mainly bind to G protein-coupled receptors (GPCRs) to trigger intracellular secondary messenger release including adenosine 3', 5'-cyclic monophosphate (cAMP), thus modulating cancer progress in different kind of tumours. As one of the downstream effectors of cAMP, exchange proteins directly activated by cAMP (EPACs) play dual roles in cancer proliferation and metastasis. More evidence about the relationship between neuropeptides and EPAC pathways have been proposed for their potential role in cancer development; hence, this review focuses on the role of neuropeptide/GPCR system modulation of cAMP/EPACs pathways in cancers. The correlated downstream pathways between neuropeptides and EPACs in cancer cell proliferation, migration, and metastasis is discussed to glimmer the direction of future research.

The peptidyl-prolyl isomerases FKBP15-1 and FKBP15-2 negatively affect lateral root development by repressing the vacuolar invertase VIN2 in Arabidopsis.

MAIN CONCLUSION: The peptidyl-prolyl isomerases FKBP15-1 and FKBP15-2 negatively modulate lateral root development by repressing vacuolar invertase VIN2 activity. Lateral root (LR) architecture greatly affects the efficiency of nutrient absorption and the anchorage of plants. Although the internal phytohormone regulatory mechanisms that control LR development are well known, how external nutrients influence lateral root development remains elusive. Here, we characterized the function of two FK506-binding proteins, namely, FKBP15-1 and FKBP15-2, in Arabidopsis. FKBP15-1/15-2 genes were expressed prominently in the vascular bundles of the root basal meristem region, and the FKBP15-1/15-2 proteins were localized to the endoplasmic reticulum of the cells. Using IP-MS, Co-IP, and BiFC assays, we demonstrated that FKBP15-1 and FKBP15-2 interacted with vacuolar invertase 2 (VIN2). Compared to Col-0 and the single mutants, the fkbp15-1fkbp15-2 double mutant had more LRs, and presented higher sucrose catalytic activity. Moreover, genetic analysis showed genetic epistasis of VIN2 over FKBP15-1/FKBP15-2 in controlling LR development. Our results indicate that FKBP15-1 and FKBP15-2 participate in the control of LR number by inhibiting the catalytic activity of VIN2. Owing to the conserved peptidylprolyl cis-trans isomerase activity of FKBP family proteins, our results provide a clue for further analysis of the interplay between lateral root development and protein modification by FKBPs.

GhbHLH18 negatively regulates fiber strength and length by enhancing lignin biosynthesis in cotton fibers.

Cotton fibers are developed epidermal cells of the seed coat and contain large amounts of cellulose and minor lignin-like components. Lignin in the cell walls of cotton fibers effectively provides mechanical strength and is also presumed to restrict fiber elongation and secondary cell wall synthesis. To analyze the effect of lignin and lignin-like phenolics on fiber quality and the transcriptional regulation of lignin synthesis in cotton fibers, we characterized the function of a bHLH transcription factor, GhbHLH18, during fiber elongation stage. GhbHLH18 knock-down plants have longer and stronger fibers, and accumulate less lignin-like phenolics in mature cotton fibers than control plants. By mining public transcriptomic data for developing fibers, we discovered that GhbHLH18 is coexpressed with most lignin synthesis pathway genes. Furthermore, we showed that GhbHLH18 strongly binds to the E-box in the promoter region of GhPER8 and activates its expression. Transient over expression of GhPER8 protein in tobacco leaves significantly decreased the content of coniferyl alcohol and sinapic alcohol-the substrate respectively for G-lignin and S-lignin biosynthesis. These results suggest that GhbHLH18 is negatively associated with fiber quality by activating peroxidase-mediated lignin metabolism, thus the paper represents an alternative strategy to improve fiber quality.

Cotton fiber elongation requires the transcription factor GhMYB212 to regulate sucrose transportation into expanding fibers.

Cotton is white gold across the globe and composed of fiber cells derived from the outer integument of cotton ovules. Fiber elongation uses sucrose as a direct carbon source. The molecular mechanism transcriptionally controlling sucrose transport from ovules into the elongating fibers remains elusive. In this study the involvement of GhMYB212 in the regulation of sucrose transportion into expanding fibers was investigated. GhMYB212 RNAi plants (GhMYB212i) accumulated less sucrose and glucose in developing fibers, and had shorter fibers and a lower lint index. RNA-seq and protein-DNA binding assays revealed that GhMYB212 was closely linked to the pathways of sucrose and starch transportation and metabolism, directly controling the expression of a sucrose transporter gene GhSWEET12. GhSWEET12 RNAi plants (GhSWEET12i) possessed similar fiber phenotypes to those of GhMYB212i. Exogenous sucrose supplementation in ovule cultures did not rescue the shorter fiber phenotype of GhMYB212i and GhSWEET12i. This finding supported the idea that the attenuated rate of sucrose transport from the outer seed coat into the fibers is responsible for the retardation of fiber elongation. Current investigations support the idea that GhMYB212 functions as the main regulator of fiber elongation by controlling the expression of GhSWEET12, and therefore it is important to study cell expansion and sugar transportation during seed development.