Nuclear presence of galectins suggests a role of these endogenous lectins in the regulation of transcription, pre-mRNA splicing and transport processes. Therefore, detection and localization of nuclear binding sites for galectins by a new methodological step, has potential to further functional analysis. In the first step of our model study we monitored the nuclear expression of galectins-1 and -3 in cultured stromal cells of human bone marrow and human/porcine keratinocytes. To enable detection and localization of galectin-specific binding sites, we used purified galectins biotinylated without loss of activity as cytochemical tool. The degree of labeling of the probes was determined by adapting two-dimensional gel electrophoresis and calculating pI changes in response to stepwise chemical modification of basic and acidic side chains by the biotinylation reagents. Binding studies revealed positivity for galectin-1, whereas galectins-3, -5, and -7 were not reactive with nuclear sites under identical conditions in bone marrow stromal cells and keratinocytes prepared from hair follicle enriched for stem cells. Inhibition by lactose indicated an involvement of the carbohydrate recognition domain in nuclear binding of galectin-1. Colocalization of the galectin-1-dependent signal with the SC35 splicing factor and sensitivity toward RNase treatment argued in favor of galectin binding in nuclear speckles, albeit only for a small fraction of the cells. Epidermal cells positive for galectin-1-binding sites expressed DNp63 known as a potential marker of stem cells. Based on cytokeratin expression cells with nuclear binding of labeled galectin-1 were basal and not suprabasal cells. Regarding proliferation, no relationship to the expression of a proliferation marker, Ki-67, was observed. The nucleolar signal colocalized with fibrillarin and nucleophosmin/B23 as representatives of nucleolar proteins in both types of studied cells. In conclusion, the application of labeled galectins to localize accessible binding sites adds a new aspect to the functional analysis of these lectins in the nucleus.

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A thorough characterization of the properties of squamous epithelial cells is necessary in order to improve our understanding of the functional aspects of normal development and malignant aberrations. Up to now, studies have focused almost exclusively on monitoring distinct protein markers. With our growing awareness of the coding function of glycan chains of cellular glycoconjugates and their interaction with receptors (lectins) in situ, defining the glycophenotype of these cells has become an important issue. Whereas the commonly applied plant lectins are tools used to map the presence and localization of biochemically defined saccharide epitopes, the introduction of endogenous (mammalian) lectins to this analysis enables us to take the step from monitoring the presence of glycan to understanding the functional implications by revealing ligand properties of the detected epitope for tissue lectin. Thus, in this study we investigated a distinct aspect of glycosylation using plant and mammalian lectins, i.e. the linkage type of sialylation. We first mapped the expression profile of the type of sialylation (alpha2,3- or alpha2,6-linked) by plant lectins. Based on the hypothesis that this factor regulates accessibility of ligands for endogenous lectins we introduced two labeled galectins to this study. Galectin-3 (but not galectin-1) binding was related to cell differentiation in normal adult and developing epithelia, cultured epidermal cells, and carcinomas derived from these epithelia. The presented data suggest that alpha2,6-linked N-acetyl-D-neuraminic acid moieties could serve to mask galectin-3-reactive glycoepitopes. As a consequence, monitoring of the linkage type of sialic acid in glycans by plant lectins therefore has implications for the extent of glycan reactivity with endogenous lectins, pointing to a potential function of changes in sialylation type beyond these cell and lectin systems.

Sporadic basal cell carcinoma (BCC) is the commonest human cancer. Although its aggressiveness is low and metastatic potential negligible, the increasing incidence of the tumor in the Western world drives attention to its pathogenesis. In 1996, germ-line mutations in the patched receptor of the Sonic hedgehog (Shh) signaling pathway were described in the Gorlin-Goltz syndrome in association with multiple nevoid BCCs. Later, the aberrant activation of the Shh was identified in sporadic BCCs as well. Recently, the role of other tumor suppressors and DNA repair gene mutations and their relationship with UV radiation-induced DNA damage have been elucidated.