ST3GAL6 Antibody

What is ST3GAL6 and what are its key biological functions?

ST3GAL6 is a sialyltransferase enzyme that transfers N-acetylneuraminic acid to galactose present on glycolipids (gangliosides) or the N- or O-linked sugar chains of glycoproteins. It plays a crucial role in the formation of Sialyl-Lewis X, a carbohydrate structure critical for selectin-induced cell adhesion and a blood group antigen . In cellular contexts, ST3GAL6 contributes to glycosylation patterns that influence cell-cell interactions, particularly in immune and cancer cell biology. Functionally, it participates in protein-protein interactions with other glyco-metabolism-related proteins, including FUT family members and B4GALTs, and is involved in processes like keratan sulfate metabolism, glycosaminoglycan biosynthesis, and mucopolysaccharide metabolism .

While the calculated molecular weight of ST3GAL6 is 38 kDa (based on 331 amino acids), the observed molecular weight in experimental conditions typically ranges from 50-60 kDa . This difference is likely due to post-translational modifications, particularly glycosylation, which is common for glycosyltransferases. Researchers should anticipate this higher apparent molecular weight when analyzing Western blot results.

What positive controls are recommended for ST3GAL6 antibody experiments?

Based on antibody validation data, several positive controls have been confirmed:

• Cell lines: MCF-7, Sp2/0 cells

• Tissues: Human heart tissue, human placenta tissue, human liver tissue

• Pathological samples: Human urothelial carcinoma tissue

For immunohistochemistry studies specifically, human liver tissue shows reliable positive staining and can serve as an appropriate control.

How does ST3GAL6 expression correlate with cancer prognosis?

ST3GAL6 expression shows context-dependent prognostic value across different cancer types:

This differential prognostic pattern highlights the tissue-specific roles of ST3GAL6 and suggests that its altered glycosylation effects may have distinct consequences in different tumor microenvironments.

What is the relationship between ST3GAL6 and the EGFR signaling pathway?

In lung adenocarcinoma, ST3GAL6 has been identified as a negative regulator of EGFR signaling. Protein-protein interaction network analysis through GeneMANIA revealed that ST3GAL6 is associated with the EGFR signaling pathway . Functional studies demonstrated that knockdown of ST3GAL6 in LUAD cells results in:

• Elevated phosphorylation levels of EGFR at Y1068 and Y1173

• Increased phosphorylation of downstream ERK at T202/T204

• Significantly increased expression of MMP2 and MMP9 (matrix metalloproteinases)

• Enhanced invasion capabilities of LUAD cells

This suggests that ST3GAL6 may influence cancer progression by modulating EGFR-mediated signaling, potentially through alterations in receptor glycosylation patterns that affect receptor activation and downstream pathways.

How is ST3GAL6 expression regulated at the molecular level?

Research has identified a long non-coding RNA (lncRNA), ST3GAL6-AS1, that overlaps with the first exon of the ST3GAL6 gene and serves as an upstream regulator . Key findings include:

• ST3GAL6-AS1 is downregulated in LUAD samples compared to normal lung tissues

• Expression levels of ST3GAL6 and ST3GAL6-AS1 are positively correlated in LUAD samples

• Depletion of ST3GAL6-AS1 in A549 and HCC827 cells results in:

  • Downregulation of ST3GAL6 at both mRNA and protein levels

  • Elevated phosphorylation of EGFR and ERK

  • Increased MMP2 and MMP9 expression

  • Enhanced invasion capabilities

This regulatory mechanism provides important insights for researchers studying transcriptional control of glycosyltransferases and opens potential therapeutic avenues targeting ST3GAL6-AS1 to modulate ST3GAL6 expression.

What are the optimal protocols for ST3GAL6 detection by immunohistochemistry?

For successful immunohistochemical detection of ST3GAL6:

• Recommended antibody dilution: 1:50-1:500 (optimized based on sample type)

• Antigen retrieval: Two options validated:

  • TE buffer pH 9.0 (recommended primary option)

  • Alternative: Citrate buffer pH 6.0

• Positive control tissues: Human liver tissue, human urothelial carcinoma tissue

• Detection system: Standard secondary antibody approach with chromogenic detection

For differential analysis between normal and pathological samples, it's valuable to note that ST3GAL6 protein is strongly detected in the cytosol of normal lung epithelial cells, while showing reduced expression in LUAD specimens, with progressive reduction correlating with advanced disease stages .

How can I design appropriate knockdown experiments for ST3GAL6 functional studies?

Based on published research, effective ST3GAL6 knockdown experiments should include:

• Cell line selection: Multiple myeloma cell lines (for hematological malignancy studies) or lung adenocarcinoma cell lines (A549, HCC827) show functional consequences upon ST3GAL6 modulation

• Readouts to assess:

  • Adhesion capabilities (especially to bone marrow stromal cells)

  • In vivo homing capabilities (in xenograft models)

  • EGFR pathway activation (phosphorylation status)

  • Invasion capabilities (transwell assays)

  • Expression of downstream targets (MMP2, MMP9)

• Controls: Include scrambled shRNA controls and validate knockdown efficiency at both mRNA and protein levels

• Complementary approach: Consider parallel knockdown of ST3GAL6-AS1, which regulates ST3GAL6 expression, to confirm the specificity of observed effects

How should Western blot protocols be optimized for ST3GAL6 detection?

For optimal Western blot detection of ST3GAL6:

• Recommended dilution range: 1:1000-1:8000

• Expected molecular weight: 50-60 kDa (rather than the calculated 38 kDa)

• Positive controls: MCF-7 cells, human heart tissue, Sp2/0 cells, human placenta tissue

• Loading controls: Anti-actin or anti-α-tubulin are suitable protein-loading controls

• Membrane type: PVDF membranes have been successfully used

• Sample preparation: Whole-cell lysates subjected to standard SDS-PAGE protocols are appropriate

How should I interpret differential ST3GAL6 expression between normal and diseased states?

When analyzing ST3GAL6 expression patterns:

• Multiple Myeloma: ST3GAL6 is expressed at significantly higher levels in CD138+ cells from MM patients compared to healthy donors, with the highest expression in secondary plasma cell leukemia. Immunohistochemistry shows ST3GAL6 expression in MGUS and MM cases not apparent in healthy donors .

• Lung Adenocarcinoma: ST3GAL6 is significantly downregulated in LUAD compared to normal lung tissue. IHC validation confirms that ST3GAL6 protein level is significantly lower in LUAD tissues than in paired normal lung tissues (n = 88) .

• Progressive changes: In LUAD, ST3GAL6 expression progressively decreases with advanced disease stages, with ST3GAL6 slightly reduced in stage 1 and barely detectable in stage 3 .

These contrasting patterns highlight the context-dependent roles of ST3GAL6 in different malignancies and suggest that alterations in glycosylation may have tissue-specific consequences.

What functional pathways are associated with ST3GAL6 in disease contexts?

GO enrichment and KEGG analyses of ST3GAL6-associated genes in LUAD revealed involvement in multiple pathways:

Biological processes (BP):

• Immune system regulation (antigen processing and presentation)

• Neutrophil activation in immune response

• Mitotic nuclear division

Cellular components (CC):

• MHC class II protein complex

• Mitotic spindle

• Condensed chromosome, centromeric region

Molecular function (MF):

• MHC class II receptor activity

• Carbohydrate binding

• Rho GTPase binding

KEGG pathways:

• Intestinal immune network for IgA production

• Cell adhesion molecules

• NOD-like receptor signaling

• Antigen processing and presentation

• Inflammatory pathways

Understanding these associated pathways provides researchers with potential mechanisms through which ST3GAL6 may influence disease progression beyond its canonical glycosylation functions.

How does ST3GAL6 function in the tumor microenvironment and immune response?

ST3GAL6 contributes to the tumor microenvironment through several mechanisms:

• Formation of selectin ligands: ST3GAL6 is involved in creating Sialyl-Lewis X, critical for selectin-mediated cell adhesion . In multiple myeloma, this may influence interactions between malignant plasma cells and the bone marrow microenvironment .

• Immune modulation: GO and KEGG analyses reveal ST3GAL6 association with immune functions including antigen processing, presentation, and MHC class II activity . This suggests ST3GAL6-mediated glycosylation may influence immune recognition of tumor cells.

• EGFR signaling regulation: In LUAD, ST3GAL6 negatively regulates EGFR signaling, influencing tumor cell behavior and potentially response to EGFR-targeted therapies .

• Correlation with immune markers: In some MM cases with high ST3GAL6 levels, corresponding high levels of HECA-452 staining (which binds to sLex and reacts with both cutaneous lymphocyte antigen and hematopoietic cell E-/L-selectin ligand) were observed .

These functions indicate that ST3GAL6 may be an important mediator at the interface between tumor cells and their microenvironment, particularly in contexts involving immune cell interactions.

What are the therapeutic implications of targeting ST3GAL6 in cancer?

Given the demonstrated roles of ST3GAL6 in different cancers, several therapeutic implications emerge:

• Multiple Myeloma: ST3GAL6 knockdown inhibits in vivo homing and prolongs survival in xenograft mice. High expression correlates with inferior survival, particularly in patients not receiving high-dose therapy or thalidomide . This suggests ST3GAL6 inhibition could complement existing MM therapies.

• Lung Adenocarcinoma: ST3GAL6 negatively regulates EGFR signaling, and its downregulation promotes invasion. This implies that strategies to restore ST3GAL6 expression might help suppress EGFR-driven lung cancer progression .

• Upstream targeting: The identification of ST3GAL6-AS1 as a regulator of ST3GAL6 provides a potential target for modulating ST3GAL6 expression. In LUAD, enhancing ST3GAL6-AS1 might increase ST3GAL6 levels and thereby suppress EGFR signaling .

These diverse implications highlight the importance of context-specific approaches when considering ST3GAL6 as a therapeutic target.

How might ST3GAL6 antibodies be used in biomarker development?

The prognostic significance of ST3GAL6 in different cancers suggests potential biomarker applications:

Development of standardized IHC protocols using validated ST3GAL6 antibodies would be critical for translating these findings to clinical biomarker applications.