Selective inhibition of cotranslational translocation of vascular cell adhesion molecule 1

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Abstract

Increased expression of vascular cell adhesion molecule 1 (VCAM1) is associated with a variety of chronic inflammatory conditions, making its expression and function a target for therapeutic intervention1,2,3. We have recently identified CAM741, a derivative of a fungus-derived cyclopeptolide that acts as a selective inhibitor of VCAM1 synthesis in endothelial cells. Here we show that the compound represses the biosynthesis of VCAM1 in cells by blocking the process of cotranslational translocation, which is dependent on the signal peptide of VCAM1. CAM741 does not inhibit targeting of the VCAM1 nascent chains to the translocon channel but prevents translocation to the luminal side of the endoplasmic reticulum (ER), through a process that involves the translocon component Sec61β. Consequently, the VCAM1 precursor protein is synthesized towards the cytosolic compartment of the cells, where it is degraded. Our results indicate that the inhibition of cotranslational translocation with low-molecular-mass compounds, using specificity conferred by signal peptides, can modulate the biosynthesis of certain secreted and/or membrane proteins. In addition, they highlight cotranslational translocation at the ER membrane as a potential target for drug discovery.

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Figure 1: CAM741 dose-dependently inhibits VCAM1 glycoprotein expression in transiently transfected cells.
Figure 2: Overexpressed VCAM1 is degraded by intracellular proteases in the presence of CAM741.
Figure 3: CAM741 dose-dependently inhibits cotranslational translocation of VCAM1.
Figure 4: VCAM1 SP is crucial for conferring cyclopeptolide sensitivity.
Figure 5: CAM741 inhibits VCAM1 translocation at a post-targeting step and enhances interaction of the VCAM1 nascent chains with Sec61β.

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Acknowledgements

We thank A. Rot for generation of the Sec61β antiserum, H. Jaksche for its purification, and B. Kappel, H. Pertl, L. Hofer and N. Lettner for technical assistance.

Author information

Correspondence to Jan E. de Vries.

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Competing interests

H.H., S.W., C.A.F., E.P.S., B.O., J.E.d.V. and I.J.D.L. are employed by Novartis. In addition, B.O., J.E.d.V. and I.J.D.L. own restricted stocks. J.B., K.M. and C.D.-N. left Novartis and do not have any conflicting financial interests.

Supplementary information

Supplementary Data

Data demonstrating that the VCAM-1 SP plus the first amino acid of the VCAM-1 mature region are required for full sensitivity to CAM741, while parts of the N-terminal region of the VCAM-1 SP are dispensable for translocation and sensitivity. (DOC 24 kb)

Supplementary Figure S1

In vitro translocation experiments described in Supplementary Data. (PDF 225 kb)

Supplementary Figure S1 Legend (DOC 23 kb)

Supplementary Table S1

Results from transient transfection experiments described in Supplementary Data. (DOC 32 kb)

Supplementary Methods

Methods for experiment shown in Supplementary Table S1. (DOC 19 kb)

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Besemer, J., Harant, H., Wang, S. et al. Selective inhibition of cotranslational translocation of vascular cell adhesion molecule 1. Nature 436, 290–293 (2005) doi:10.1038/nature03670

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