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Intracellular modifications of human apolipoprotein E
Abstract:
We have used pulse-chase techniques to study the synthesis, intracellular modification, and secretion of human apolipoprotein E by cultures of HepG2 cells and peripheral blood human monocyte macrophages. We have found that modified apoE isoproteins are detectable intracellularly after 14 min of pulse, and their relative concentration increases linearly over a 2-h pulse-chase period. At the same time, the relative concentration of unmodified apoE decreases. All the major modified apoE isoproteins appear simultaneously, and they correspond to the sialo apoE forms apooEs2, apoEs4, and apoEs6. ApoE secretion is first detected after a 30-min pulse. Secreted apoE consists of 92% of the same sialated apoE forms observed intracellularly. ApoE sialation and secretion were not affected by treatment of the HepG2 cultures with tunicamycin, whereas monensin inhibited both the intracellular sialation and secretion of this protein. These findings suggest that apoE is secreted in the form of three major isoproteins generated by intracellular modification of this protein with one or more O-linked oligosaccharide chains containing sialic acid.
Insights
Human apolipoprotein E (apoE) undergoes intracellular modification into three major sialylated isoproteins. These modified apoE forms are then secreted by HepG2 cells and macrophages.
Area of Science:
- Biochemistry
- Cell Biology
- Molecular Biology
Background:
- Apolipoprotein E (apoE) plays a crucial role in lipid metabolism and transport.
- Understanding the post-translational modifications and secretion pathways of apoE is essential for elucidating its function.
Purpose of the Study:
- To investigate the synthesis, intracellular modification, and secretion of human apolipoprotein E (apoE).
- To identify the specific forms of apoE that are secreted and the mechanisms regulating these processes.
Main Methods:
- Pulse-chase experiments were conducted using HepG2 cells and human monocyte-derived macrophages.
- Cells were treated with tunicamycin and monensin to assess their effects on apoE sialylation and secretion.
Main Results:
- Modified apoE isoproteins were detected intracellularly within 14 minutes and increased over a 2-hour period.
- Three major sialylated apoE isoproteins (apoEs2, apoEs4, apoEs6) were identified intracellularly and in secreted fractions.
- ApoE secretion was observed after 30 minutes, with secreted apoE predominantly consisting of the same sialylated forms.
- Tunicamycin did not affect apoE sialylation or secretion, while monensin inhibited both processes.
Conclusions:
- Human apolipoprotein E is secreted as three major isoproteins.
- These isoproteins are generated through intracellular modification involving O-linked oligosaccharide chains containing sialic acid.
- The findings suggest a specific pathway for apoE modification and secretion in HepG2 cells and macrophages.