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

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

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

Keystone Symposia

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 line 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 (WB) while IgG glycosylation was analyzed in culture supernatants by lectin-ELISA and WB before and after endoglycosidase H, PNGase F or O-Sialoglycoprotein Endopeptidase digestion. We found that P4 induces a switch of STT3 isoform expression and increases Fab N-glycosylation. Neutralizing assays showed that these effects were dependent on the progesterone-induced blocking factor (PIBF), which was previously found to be an immunomodulatory molecule involved in fetomaternal tolerance. In order to investigate an in vivo modulation of OST isoform expression, we analyzed STT3-A/B expression at the maternofetal interface in a mouse abortion-prone model (CBA/JxDBA/2J). Sound stress was previously found to increase the abortion rate of these mice and to decrease serum levels of P4, PIBF and extra N-glycosylated blocking IgG antibodies. 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. 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.