PMID:16688858

Citation	Saitoh, N, Uchimura, Y, Tachibana, T, Sugahara, S, Saitoh, H and Nakao, M (2006) In situ SUMOylation analysis reveals a modulatory role of RanBP2 in the nuclear rim and PML bodies. Exp. Cell Res. 312:1418-30
Abstract	SUMO modification plays a critical role in a number of cellular functions including nucleocytoplasmic transport, gene expression, cell cycle and formation of subnuclear structures such as promyelocytic leukemia (PML) bodies. In order to identify the sites where SUMOylation takes place in the cell, we developed an in situ SUMOylation assay using a semi-intact cell system and subsequently combined it with siRNA-based knockdown of nucleoporin RanBP2, also known as Nup358, which is one of the known SUMO E3 proteins. With the in situ SUMOylation assay, we found that both nuclear rim and PML bodies, besides mitotic apparatuses, are major targets for active SUMOylation. The ability to analyze possible SUMO conjugation sites would be a valuable tool to investigate where SUMO E3-like activities and/or SUMO substrates exist in the cell. Specific knockdown of RanBP2 completely abolished SUMOylation along the nuclear rim and dislocated RanGAP1 from the nuclear pore complexes. Interestingly, the loss of RanBP2 markedly reduced the number of PML bodies, in contrast to other, normal-appearing nuclear compartments including the nuclear lamina, nucleolus and chromatin, suggesting a novel link between RanBP2 and PML bodies. SUMOylation facilitated by RanBP2 at the nuclear rim may be a key step for the formation of a particular subnuclear organization. Our data imply that SUMO E3 proteins like RanBP2 facilitate spatio-temporal SUMOylation for certain nuclear structure and function.
Links	PubMed
Keywords	Active Transport, Cell Nucleus/physiology; Biological Assay/methods; Cell Compartmentation/physiology; Cell Nucleus/genetics; Cell Nucleus/metabolism; Down-Regulation/physiology; GTPase-Activating Proteins/genetics; GTPase-Activating Proteins/metabolism; HeLa Cells; Humans; Intracellular Membranes/metabolism; Mitosis/physiology; Molecular Chaperones/genetics; Molecular Chaperones/metabolism; Neoplasm Proteins/genetics; Neoplasm Proteins/metabolism; Nuclear Envelope/genetics; Nuclear Envelope/metabolism; Nuclear Pore Complex Proteins/genetics; Nuclear Pore Complex Proteins/metabolism; Nuclear Proteins/genetics; Nuclear Proteins/metabolism; Organelles/genetics; Organelles/metabolism; Promyelocytic Leukemia Protein; Protein Transport/physiology; RNA Interference; Small Ubiquitin-Related Modifier Proteins/genetics; Small Ubiquitin-Related Modifier Proteins/metabolism; Transcription Factors/genetics; Transcription Factors/metabolism; Tumor Suppressor Proteins/genetics; Tumor Suppressor Proteins/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
HUMAN:RBP2	enables	GO:0019789: SUMO transferase activity	ECO:0000269: experimental evidence used in manual assertion		F		Seeded From UniProt	complete

References

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