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Defects in secretory pathway trafficking during sperm development in Adam2 knockout mice
Kathryn K Stein1, Jowell C Go, Paul Primakoff
1Section of Molecular and Cellular Biology, University of California, Davis, 95616, USA.
Abstract:
Adam2-null and Adam3-null male mice exhibit reduced levels of one or more ADAM proteins on mature sperm, in addition to the loss of the genetically targeted protein. ADAM protein loss was believed to occur posttranslationally, although the timing of loss and the mechanism by which the loss occurred were not explored. In this study we have found that in Adam3-null mice, fertilin beta (also known as ADAM2) is lost during the formation of testicular sperm. In Adam2-null males, most cyritestin (ADAM3) protein is also lost at this stage, but 25% of cyritestin is lost later, during sperm passage through the epididymis. Although normal levels of cyritestin are synthesized and acquire Endoglycosidase H resistance, indicating transit through the Golgi, the protein does not reach the cell surface. We also discovered that the majority of both fertilin beta and cyritestin are found in a Triton X-100 insoluble compartment on testicular sperm, when most of the cyritestin was observed on the cell surface. This insoluble compartment may represent a sorting platform, because in Adam2-knockout cells, only a small fraction of the cyritestin becomes Triton X-100 insoluble. Thus, it appears that cyritestin loss in Adam2-knockout mice may result, at least in part, from a disruption in protein trafficking.
Insights
In Adam3-null mice, ADAM2 protein is lost during sperm development. In Adam2-null mice, ADAM3 protein trafficking is disrupted, leading to its loss during epididymal passage.
Area of Science:
- Reproductive biology
- Spermatogenesis
- Protein trafficking
Background:
- ADAM (a disintegrin and metalloproteinase) proteins are crucial for sperm function.
- Previous studies indicated post-translational loss of ADAM proteins in null mice, but mechanisms remained unclear.
Purpose of the Study:
- To investigate the timing and mechanisms of ADAM2 and ADAM3 protein loss in Adam2-null and Adam3-null mice.
- To elucidate the role of protein trafficking in ADAM protein regulation during sperm maturation.
Main Methods:
- Analysis of ADAM2 and ADAM3 protein levels in testicular and epididymal sperm from Adam2-null and Adam3-null mice.
- Biochemical assays including Endoglycosidase H resistance and Triton X-100 solubility.
- Cellular localization studies of ADAM proteins.
Main Results:
- ADAM2 is lost during spermatogenesis in Adam3-null mice.
- In Adam2-null mice, most ADAM3 is lost during spermatogenesis, with the remainder lost during epididymal transit.
- ADAM3 in Adam2-null mice reaches the Golgi but fails to traffic to the cell surface, suggesting a trafficking defect.
- ADAM2 and ADAM3 are found in a Triton X-100 insoluble fraction in testicular sperm, potentially indicating a sorting platform.
Conclusions:
- ADAM protein loss in null mice occurs at specific stages of sperm development and epididymal transit.
- Disrupted protein trafficking, particularly for ADAM3 in Adam2-null mice, contributes to protein loss.
- The Triton X-100 insoluble compartment may play a role in ADAM protein sorting and cell surface localization.
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