Developmental expression of survivin during embryonic submandibular salivary gland development

Abstract

The regulation of programmed cell death is critical to developmental homeostasis and normal morphogenesis of embryonic tissues. Survivin, a member of the inhibitors of apoptosis protein (IAP) family primarily expressed in embryonic cells, is both an anti-apoptosis and a pro-survival factor. Since our previous studies have demonstrated the importance of apoptosis during embryonic submandibular salivary gland (SMG) development, we postulated that survivin is a likely mediator of SMG epithelial cell survival. Stage SMGs. Survivin is known to be a pro-survival and anti-apoptotic factor. Given that survivin translocation into the nucleus is required for the induction of entry into the cell cycle and the inhibition of apoptosis, our demonstration of nuclear-localized survivin protein in presumptive ductal and proacinar lumen-bounding cells suggests that survivin may be a key mediator of embryonic SMG epithelial cell survival.

Background

The regulation of programmed cell death is critical to developmental homeostasis and normal morphogenesis of embryonic tissues. Survivin, a member of the inhibitors of apoptosis protein (IAP) family primarily expressed in embryonic cells, is both an anti-apoptosis and a pro-survival factor. Since our previous studies have demonstrated the importance of apoptosis during embryonic submandibular salivary gland (SMG) development, we postulated that survivin is a likely mediator of SMG epithelial cell survival.

Results

Stage SMGs.

Conclusions

Survivin is known to be a pro-survival and anti-apoptotic factor. Given that survivin translocation into the nucleus is required for the induction of entry into the cell cycle and the inhibition of apoptosis, our demonstration of nuclear-localized survivin protein in presumptive ductal and proacinar lumen-bounding cells suggests that survivin may be a key mediator of embryonic SMG epithelial cell survival.

Introduction

]. However, although we have identified different apoptotic pathways which mediate duct and terminal bud lumen formation, little is known about which molecule(s) mediate the apoptotic stop signal such that these lumen-bounding epithelial cells survive.

].

Given the above, we postulated that survivin is an important pro-survival and anti-apoptotic molecule during embryonic ductal and proacinar formation. In this paper, we investigated the developmental expression of survivin transcripts and protein in embryonic SMGs. This is the first report of notable developmental changes in survivin expression and protein localization correlated with embryonic lumen formation.

Expression of Survivin Transcript

].

Stage SMGs.

Determination of Antibody Specificity

]. Based on the above results, we conclude that this anti-survivin antibody is specific for survivin protein.

Western blot analysis using whole antiserum shows the presence of survivin protein in 3 cancer cell lines (MO7E, HL-60, and HELA). Survivin protein was not detected in whole blood lysates or control preimmune serum.

Western blot analysis for survivin expression. Using affinity-purified anti-survivin antibody, survivin protein was identified as a single ~ 16 kDa band in HL-60 lysates. No band was seen in affinity column eluate of prebleed serum.

Spatiotemporal Distribution of Survivin

].

Stage ductal epithelial bilayer which consists of an inner columnar epithelia which surround the distinct lumen and an outer cuboidal epithelia which abuts the basement membrane. Nuclear localization (double arrowheads) of survivin protein is seen in the inner layer of columnar epithelial cells surrounding the lumen (*) and not in the outer cuboidal epithelia. A-C. Bar, 50 μm. D-E, Bar, 25 μm.

) suggests that survivin may be a key factor for the survival of epithelial cells surrounding forming ductal and terminal bud lumina.

Stage. What remains unclear is which factor or factors protect the initial bud epithelium from apoptosis. Further studies are needed to address this question.

Conclusions

Although apoptosis has been shown to mediate embryonic SMG ductal and proacinar lumen formation, little is known about which factor(s) mediate the anti-apoptosis/pro-survival signal. Our demonstration of a significant increase in survivin expression concomitant with SMG lumina formation, as well as survivin protein's nuclear localization in presumptive ductal and proacinar lumen-bounding cells, suggest that survivin is be a key mediator of embryonic SMG lumen-bounding epithelial cell survival.

Tissue Collection

]. E14-E18 SMGs and E15-E17 lungs were collected and stored at -70°C or processed for histology.

Analysis of RNA by Primer Extension

].

Immunohistochemistry

]. Immunohistochemical staining was performed using the Elite ABC kit (Vector Laboratories, Burlingham, CA).

Acknowledgements

This research was supported by NIH grant DE 11942.