Submitted on June 10, 2005
Accepted on March 31, 2006
Translocon Pores in the Endoplasmic Reticulum Are Permeable to Small Anions
Beata Lizak1, Ibolya Czegle1, Miklos Csala2, Angelo Benedetti3, Jozsef Mandl2, and Gabor Banhegyi4*
1 Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, Budapest, Hungary
2 Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, Budapest, Hungary; Endoplasmic Reticulum Research Group, Hungarian Academy of Sciences and Semmelweis University, Budapest, Hungary
3 Dipartimento di Fisiopatologia, Medicina Sperimentale e Sanita Pubblica, Universita di Siena, Siena, Italy
4 Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, Budapest, Hungary; Endoplasmic Reticulum Research Group, Hungarian Academy of Sciences and Semmelweis University, Budapest, Hungary; Dipartimento di Fisiopatologia, Medicina Sperimentale e Sanita Pubblica, Universita di Siena, Siena, Italy
* To whom correspondence should be addressed. E-mail: banhegyi{at}puskin.sote.hu.
Contribution of translocon peptide channel to the permeation of low molecular mass anions was investigated in rat liver microsomes. Puromycin, which purges translocon pores of nascent polypeptides creating additional empty pores raised the microsomal uptake of radiolabeled UDP-glucuronic acid, while did not increase the uptake of glucose-6-phosphate or glutathione. The role of translocon pore in the transport of small anions was also envisaged by measuring the effect of puromycin on the activity of microsomal enzymes with intraluminal active sites. The mannose-6-phosphatase activity of glucose-6-phosphatase and the activity of UDP-glucuronosyltransferase were elevated upon addition of puromycin, but glucose-6-phosphatase and 
-glucuronidase activities were not changed. The increase in enzyme activities was due to a better access of the substrates to the luminal compartment rather than to activation of the enzymes. Antibody against Sec61 translocon component decreased the activity of UDP-glucuronosyltransferase and antagonized the effect of puromycin. Similarly, the addition of the puromycin antagonist anisomycin or treatments of microsomes resulting in the release of attached ribosomes prevented the puromycin-dependent increase in the activity. Mannose-6-phosphatase and UDP-glucuronosyltransferase activities of smooth microsomal vesicles showed higher basal latencies, which were not affected by puromycin. In conclusion, translationally inactive, ribosome-bound translocons allow small anions to cross the endoplasmic reticulum membrane. This pathway can contribute to the non-specific substrate supply of enzymes with intraluminal active centers.
