Progesterone induces a switch in oligosaccharyltransferase isoform expression: consequences on IgG N-glycosylation

PRADOS MB; ARCK PC; SZEKERES-BARTHO J; CARAMELO J; MIRANDA S

Congreso; New Frontiers at the Interface of Immunity and Glycobiology.; 2011

Keystone Symposia on Molecular and Cellular Biology

Progesterone induces a switch in oligosaccharyltransferase isoform expression: consequences on IgG N-glycosylation. 1Prados, MB; 1La Blunda, J; 2Szekeres-Bartho, J; 3Caramelo, J; 1Miranda, S. 1Inst. de Investigaciones Cardiológicas “Prof. Dr. Alberto C. Taquini”, CONICET- Universidad de Buenos Aires, Buenos Aires, Argentina. 2Department of Medical Microbiology and Immunology, Medical School, Pecs University, Hungary. 3Inst. de Investigaciones Bioquímicas de Buenos Aires, CONICET and Fundación Instituto Leloir, Buenos Aires, Argentina. The presence of N-glycans in the Fab region of IgG has shown to modify the properties of these molecules including changes in antibody affinity and stability. However, the underlying mechanism responsible for the presence or absence of these glycans remains unknown. N-linked glycosylation is catalyzed in the lumen of the endoplasmic reticulum by the oligosaccharyltransferase complex (OST). Mammalian cells can express two isoforms of the OST catalytic subunit, STT3-A and STT3-B, which are endowed with distinct enzymatic properties. In this work we employed a murine hybridoma cell culture to study whether STT3 isoform expression could be modulated by progesterone (P4), thus altering IgG N-glycosylation. STT3-A/B expression was studied by western blot while IgG glycosylation was analyzed in culture supernatants by lectin-ELISA before and after endoglycosidase H, PNGase F or O-Sialoglycoprotein Endopeptidase digestion. We found that P4 induces a switch of STT3 isoform expression, increasing IgG N-glycosylation. These effects were dependent on the progesterone-induced blocking factor (PIBF), which was previously found to be an immunomodulatory molecule relevant for the maintenance of pregnancy. In order to investigate an in vivo modulation of OST isoform expression, we analyzed STT3-A/B expression at maternofetal interface in non-stressed and stressed pregnant CBA/J female mice mated with DBA/2J males. Sound stress was previously found to increase the abortion rate of these mice and to decrease protective extra-N-glycosylated antibodies, P4 and PIBF levels. Mice were stressed by sound exposure (300 Hz in intervals of 15s) on gestation day 5.5 during 24hs and immediately sacrificed. Afterwards implantation units were removed and cryosectioned. STT3-A/B expression was analyzed by immunohistochemistry. STT3-A reactivity localized preferentially in glands and lumenal epithelium (LE), while STT3-B reactivity was weak. However, stress up-regulated STT3-B expression in the LE while STT3-A was down-regulated. In conclusion, in vitro experiments demonstrate that the STT3-B/STT3-A ratio modulates IgG N-glycosylation. What is more, this work provides the first evidence that STT3 isoforms expression can be modulated in vitro and in vivo. This fact provides a possible explanation for tissue-specific glycan heterogeneity. New Frontiers at the Interface of Immunity and Glycobiology. Keystone Symposia on Molecular and Cellular Biology. Internacional. 2011. Canadá. (oral presentation).