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Isolation and characterization of low density detergent-insoluble membrane (LD-DIM) fraction from sea urchin sperm
1Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan.
Abstract:
The low density detergent-insoluble membrane (LD-DIM) fraction was obtained by a sucrose-density gradient centrifugation from sperm of three sea urchin species, Hemicentrotus pulcherrimus, Strongylocentrotus purpuratus, and Anthocidaris crassispina. These LD-DIM preparations were characterized by enriched glycosphingolipids (GSL) including gangliosides and sulfatide (SLF), having more than 50% of the total amount of GSL present in these sperm. Interestingly, a minor component of H. pulcherrimus sperm (HO3S-->8Neu5Acalpha2-->8Neu5Acalpha2-->6Glcbeta1++ +-->Cer) was shown to be even more enriched in the LD-DIM as revealed by using monoclonal antibody (mAb.3G9) speific to this ganglioside. In addition to the GSL, phosphatidyl-serine (PS) and diacylglcerol (DG) were enriched in the LD-DIM. On the other hand, cholesterol (CL) and sphingomyelin (SM) were not so enriched, which contrasted with the LD-DIM from Madin-Darby canine kidney (MDCK) cells, where CL and SM were reported to be abundant. Because mammalian somatic cell-derived DIMs have been proposed to be associated with functional signal transduction, it seems possible that the ganglioside-enriched LD-DIM in sea urchin sperm can participate in binding to eggs and the subsequent egg activation process. To our knowledge this is the chemical characterization of the LD-DIM fraction of a gametic cell.
Insights
Sea urchin sperm low-density detergent-insoluble membrane (LD-DIM) fractions are enriched in glycosphingolipids (GSL). This suggests a role for these GSL-rich LD-DIMs in sperm-egg interactions and egg activation.
Area of Science:
- Marine Biology
- Biochemistry
- Cell Biology
Background:
- Low-density detergent-insoluble membrane (LD-DIM) fractions are implicated in signal transduction in mammalian cells.
- Sea urchin sperm are model organisms for studying fertilization.
Purpose of the Study:
- To chemically characterize the LD-DIM fraction of sea urchin sperm.
- To investigate the potential role of LD-DIMs in sea urchin fertilization.
Main Methods:
- Sucrose-density gradient centrifugation was used to isolate LD-DIM fractions from three sea urchin species.
- Monoclonal antibody (mAb.3G9) specific to a particular ganglioside was employed for enrichment analysis.
- Lipid composition of LD-DIM fractions was analyzed.
Main Results:
- LD-DIM preparations from sea urchin sperm were enriched in glycosphingolipids (GSL), including gangliosides and sulfatides (SLF), comprising over 50% of total GSL.
- A specific ganglioside (HO3S-->8Neu5Acalpha2-->8Neu5Acalpha2-->6Glcbeta1++ +-->Cer) was found to be highly enriched in H. pulcherrimus sperm LD-DIM.
- Phosphatidylserine (PS) and diacylglycerol (DG) were also enriched, while cholesterol (CL) and sphingomyelin (SM) were not, contrasting with mammalian cell LD-DIMs.
Conclusions:
- Sea urchin sperm LD-DIMs are distinct from mammalian cell LD-DIMs in lipid composition.
- The ganglioside-enriched LD-DIM fraction in sea urchin sperm may play a role in egg binding and subsequent egg activation.