TAILIEUCHUNG - Báo cáo khoa học: The A-subunit of surface-bound Shiga toxin stimulates clathrin-dependent uptake of the toxin

Shiga toxin can be internalized by clathrin-dependent endocytosis in differ-ent cell lines, although it binds specifically to the glycosphingolipid Gb3. It has been demonstrated previously that the toxin can induce recruitment of the toxin–receptor complex to clathrin-coated pits, but whether this process is concentration-dependent or which part of the toxin molecule is involved in this process, have so far been unresolved issues. | ềFEBS Journal The A-subunit of surface-bound Shiga toxin stimulates clathrin-dependent uptake of the toxin Maria L. Torgersen1 Silje U. Lauvrak1 and Kirsten Sandvig1 2 1 Institute for Cancer Research The Norwegian Radium Hospital Montebello Oslo Norway 2 Department of Molecular Biosciences University of Oslo Norway Keywords clathrin endocytosis Shiga toxin Correspondence K. Sandvig Institute for Cancer Research The Norwegian Radium Hospital Montebello 0310 Oslo Norway Fax 47 22508692 Tel 47 22934294 E-mail ksandvig@ Received 12 May 2005 revised 14 June 2005 accepted 24 June 2005 doi Shiga toxin can be internalized by clathrin-dependent endocytosis in different cell lines although it binds specifically to the glycosphingolipid Gb3. It has been demonstrated previously that the toxin can induce recruitment of the toxin-receptor complex to clathrin-coated pits but whether this process is concentration-dependent or which part of the toxin molecule is involved in this process have so far been unresolved issues. In this article we show that the rate of Shiga toxin uptake is dependent on the toxin concentration in several cell lines HEp-2 HeLa Vero and baby hamster kidney BHK and that the increased rate observed at higher concentrations is strictly dependent on the presence of the A-subunit of cell surface-bound toxin. Surface-bound B-subunit has no stimulatory effect. Furthermore this increase in toxin endocytosis is dependent on functional clathrin as it did not occur in BHK cells after induction of antisense to clathrin heavy chain thereby blocking clathrin-dependent endocytosis. By immunofluorescence we show that there is an increased colocalization between Alexa-labeled Shiga toxin and Cy5-labeled transferrin in HeLa cells upon addition of unlabeled toxin. In conclusion the data indicate that the Shiga toxin A-subunit of cell surface-bound toxin stimulates clathrin-dependent uptake of the toxin. Possible explanations for .

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• Shiga toxin can be internalized by clathrin dependent endocytosis in differ ent cell lines
• although it binds specifically to the glycosphingolipid Gb3
• It has been demonstrated previously that the toxin can induce recruitment of the toxin–receptor complex to clathrin coated pits
• but whether this process is concentration dependent or which part of the toxin molecule is involved in this process
• have so far been unresolved issues
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