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The subcellular localization of SF-1. Fu Huang. Lab Introduction. Lab 429, Institute of Molecular Biology, Academia Sinica. Research interests I. Steroid Function and Regulation II. Zebrafish Development. PI: Bon-chu Chung, Ph.D. Sumoylation regulates nuclear distribution of SF-1 - PowerPoint PPT Presentation
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The subcellular localization of SF-1
Fu Huang
Lab Introduction
Lab 429, Institute of Molecular Biology, Academia Sinica
PI: Bon-chu Chung, Ph.D.
Research interests
I. Steroid Function and Regulation
II. Zebrafish Development
Sumoylation regulates nuclear distribution of SF-1
Cytoplasmic distribution of SF-1.
Schematic overview of the biosynthetic pathways for steroid hormones
SF1 regulates the expression of P450 steroid hydroxylases.
Steroidogenic factor 1 (SF-1)
The Ftz-F1 box of SF-1 contains NLS sequence. (Li et al., 1999)
SF-1 is an orphan nuclear receptor.SF-1 was always found in nucleus in previous stu
dies.
SF-1
SUMO-1
SUMO-1 is a 101 amino acid polypeptide and only 1-97 a.a. can be attached to target proteins.
SF-1 has two sumoylation sites, K-119 and K-194.SUMO-1 can trigger the formation of PML nuclear
bodies.
SF-1
Cloning of FLAG-RFP-SUMO-1 fusion protein
SUMO-1 WT: 94 95 96 97CAA ACG GGG GGT Gly Gly
SUMO-1 AA (mutant): 94 95 96 97CAA ACG GCG GCT Ala Ala
Western blot of FLAG-RFP-SUMO-1
FLAG-RFP-SUMO-1
Sumoylated proteins
WT AA
Sumoylation regulates nuclear distribution of SF-1
RFP-SUMO-1 WT SF-1-GFP Merge
RFP-SUMO-1 AA SF-1-GFP Merge
WT
AA
Sumoylation regulates nuclear distribution of SF-1
Cytoplasmic distribution of SF-1.
Cytoplasmic distribution of SF-1
SF-1 Ab SF-1-GFP
Phase Merge
Cytoplasmic distribution of SF-1
SF-1-GFP RFP
Merge 3D view
Cytoplasmic localization of SF-1
Actin Ab SF-1-GFP
Merge 3D view
Cytoplasmic localization of SF-1
Tublin Ab SF-1-GFP
Merge 3D view
Cytoplasmic localization of SF-1
SCC Ab SF-1-GFP
Merge
LMB does not affect cytoplasmic distribution of SF-1
15min 45min 1h15min 1h45min
2h15min 2h45min 3h15min 3h45min
4h15min 4h45min 5h15min 5h45min
6h15min 6h45min (After LMB treatment)
SF-1 contains LMB- insensitive NES homologue SF-1
FF1bAR
SF-1
FF1b
AR
(743-817)
(743-817)
(306-381)
(306-381)
(359-436)
(359-436)
AR
AR
MRER
MRER
(743-817)
(743-817)(809-883)(386-455)
(809-883)(386-455)
(Saporita et al., 2003)
The LBD domain of SF-1 contains LMB-independent NES homologue
SF-1
Future work
Examine the interaction of SF-1 and SUMO-1 in living cell under different stimulus.
Examine the candidate NES fragment of SF-1.
Use electro-microscopy to find out the exact location of cytoplasmic SF-1.
Acknowledgments
Dr. Bon-chu ChungMr. Wei-yi ChenMr. Wen-chih LeeMs. LeeAll the lab members