PIF4–Mediated Activation of YUCCA8 Expression Integrates Temperature into the Auxin Pathway in Regulating Arabidopsis Hypocotyl Growth

Higher plants adapt their growth to high temperature by a dramatic change in plant architecture. It has been shown that the transcriptional regulator phytochrome-interacting factor 4 (PIF4) and the phytohormone auxin are involved in the regulation of high temperature–induced hypocotyl elongation in Arabidopsis. Here we report that PIF4 regulates high temperature–induced hypocotyl elongation through direct activation of the auxin biosynthetic gene YUCCA8 (YUC8). We show that high temperature co-upregulates the transcript abundance of PIF4 and YUC8. PIF4–dependency of high temperature–mediated induction of YUC8 expression as well as auxin biosynthesis, together with the finding that overexpression of PIF4 leads to increased expression of YUC8 and elevated free IAA levels in planta, suggests a possibility that PIF4 directly activates YUC8 expression. Indeed, gel shift and chromatin immunoprecipitation experiments demonstrate that PIF4 associates with the G-box–containing promoter region of YUC8. Transient expression assay in Nicotiana benthamiana leaves support that PIF4 directly activates YUC8 expression in vivo. Significantly, we show that the yuc8 mutation can largely suppress the long-hypocotyl phenotype of PIF4–overexpression plants and also can reduce high temperature–induced hypocotyl elongation. Genetic analyses reveal that the shy2-2 mutation, which harbors a stabilized mutant form of the IAA3 protein and therefore is defective in high temperature–induced hypocotyl elongation, largely suppresses the long-hypocotyl phenotype of PIF4–overexpression plants. Taken together, our results illuminate a molecular framework by which the PIF4 transcriptional regulator integrates its action into the auxin pathway through activating the expression of specific auxin biosynthetic gene. These studies advance our understanding on the molecular mechanism underlying high temperature–induced adaptation in plant architecture

Reversible carotid artery narrowing in morning glory disc anomaly (vol 25, pg 198, 2005)

WOS: 00023402070001

A high-content endogenous GLUT4 trafficking assay reveals new aspects of adipocyte biology.

Funder: Wellcome-MRC, Institute of Metabolic Science, Metabolic Research Laboratories, Imaging CoreInsulin-induced GLUT4 translocation to the plasma membrane in muscle and adipocytes is crucial for whole-body glucose homeostasis. Currently, GLUT4 trafficking assays rely on overexpression of tagged GLUT4. Here we describe a high-content imaging platform for studying endogenous GLUT4 translocation in intact adipocytes. This method enables high fidelity analysis of GLUT4 responses to specific perturbations, multiplexing of other trafficking proteins and other features including lipid droplet morphology. Using this multiplexed approach we showed that Vps45 and Rab14 are selective regulators of GLUT4, but Trarg1, Stx6, Stx16, Tbc1d4 and Rab10 knockdown affected both GLUT4 and TfR translocation. Thus, GLUT4 and TfR translocation machinery likely have some overlap upon insulin-stimulation. In addition, we identified Kif13A, a Rab10 binding molecular motor, as a novel regulator of GLUT4 traffic. Finally, comparison of endogenous to overexpressed GLUT4 highlights that the endogenous GLUT4 methodology has an enhanced sensitivity to genetic perturbations and emphasises the advantage of studying endogenous protein trafficking for drug discovery and genetic analysis of insulin action in relevant cell types