The hedgehog (hh) family members control many aspects of development in both vertebrates and invertebrates. Hh signaling is associated with important biological phenomena such as patterning, cell growth, and morphogenesis. Abnormal activation of this pathway has been observed in several types of human cancers. The seven-pass transmembrane protein Smoothened (Smo) is required in both insects and mammals for transduction of the Hh signal. Our strategy is to use Drosophila as a simple and genetically tractable model system to explore the mechanisms of Hh signal transduction. The long-term goal of our research is to elucidate how Hh signals are sensed and transmitted to control downstream biological events that ultimately govern cell growth and patterning. We showed that Smo transduces the Hh signals by directly recruiting a Costal2-Fused (Cos2-Fu) complex. We also showed that Smo activation requires phosphorylation by protein kinase A (PKA) and casein kinase I (CKI), and that phosphorylation leads to increased Smo cell-surface levels and signaling activity. We recently uncovered a feedback mechanism by which Fu promotes Smo hyperphosphorylation and cell-surface accumulation by antagonizing Cos2. These findings provide new tools and hypotheses to investigate the mechanisms of Hh signaling at the cell membrane. In this project, our central hypothesis is that Smo cell-surface accumulation and signaling activity are regulated by multiple kinases, while phosphatase(s) act as inhibitory components to attenuate Smo activation. To test our hypothesis, we will use a combination of genetic and biochemical approaches in three Specific Aims: 1) To further determine if differential Smo phosphorylation transduces gradient Hh activity;2) to investigate the molecular mechanisms by which G protein-coupled receptor kinase 2 (GRK2) is involved in Hh signaling;and 3) to determine the role of phosphatase in Smo dephosphorylation and inactivation.

Public Health Relevance

Abnormal Smoothened (Smo) activation results cancers such as basal cell carcinoma (BCC) and medulloblastoma. Investigation of the Smo signaling mechanisms will provide both insights into fundamental developmental problems and new avenues for developing diagnostic tools and therapeutical treatments of cancers. PHS 398/2590 (Rev. 09/04, Reissued 4/2006) Page Project Narrative

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM079684-04
Application #
8090442
Study Section
Development - 1 Study Section (DEV1)
Program Officer
Haynes, Susan R
Project Start
2008-07-01
Project End
2013-06-30
Budget Start
2011-07-01
Budget End
2012-06-30
Support Year
4
Fiscal Year
2011
Total Cost
$283,813
Indirect Cost
Name
University of Kentucky
Department
Biochemistry
Type
Schools of Medicine
DUNS #
939017877
City
Lexington
State
KY
Country
United States
Zip Code
40506
Li, Jing; Song, Jun; Zaytseva, Yekaterina Y et al. (2016) An obligatory role for neurotensin in high-fat-diet-induced obesity. Nature 533:411-5
Li, Tongchao; Fan, Junkai; Blanco-Sánchez, Bernardo et al. (2016) Ubr3, a Novel Modulator of Hh Signaling Affects the Degradation of Costal-2 and Kif7 through Poly-ubiquitination. PLoS Genet 12:e1006054
Jiang, Kai; Liu, Yajuan; Fan, Junkai et al. (2016) PI(4)P Promotes Phosphorylation and Conformational Change of Smoothened through Interaction with Its C-terminal Tail. PLoS Biol 14:e1002375
Jiang, Kai; Jia, Jianhang (2015) Analysis of Smoothened Phosphorylation and Activation in Cultured Cells and Wing Discs of Drosophila. Methods Mol Biol 1322:45-60
Shi, Jiandang; Liu, Yajuan; Xu, Xuehe et al. (2015) Deubiquitinase USP47/UBP64E Regulates β-Catenin Ubiquitination and Degradation and Plays a Positive Role in Wnt Signaling. Mol Cell Biol 35:3301-11
Jiang, Kai; Jia, Jianhang (2015) Smoothened regulation in response to Hedgehog stimulation. Front Biol (Beijing) 10:475-486
Jiang, Kai; Liu, Yajuan; Fan, Junkai et al. (2014) Hedgehog-regulated atypical PKC promotes phosphorylation and activation of Smoothened and Cubitus interruptus in Drosophila. Proc Natl Acad Sci U S A 111:E4842-50
Zhang, Yan; Fu, Lin; Qi, Xiaolong et al. (2013) Structural insight into the mutual recognition and regulation between Suppressor of Fused and Gli/Ci. Nat Commun 4:2608
Fan, Junkai; Jiang, Kai; Liu, Yajuan et al. (2013) Hrs promotes ubiquitination and mediates endosomal trafficking of smoothened in Drosophila hedgehog signaling. PLoS One 8:e79021
Schwend, Tyler; Jin, Zhigang; Jiang, Kai et al. (2013) Stabilization of speckle-type POZ protein (Spop) by Daz interacting protein 1 (Dzip1) is essential for Gli turnover and the proper output of Hedgehog signaling. J Biol Chem 288:32809-20

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