Cystatin-related epididymal spermatogenic aggregates in the epididymis.

Cystatin-related epididymal spermatogenic (CRES) is the defining member of a reproductive subgroup within the family 2 cystatins of the cystatin superfamily of cysteine protease inhibitors. CRES is synthesized and secreted by the initial segment of the epididymis and is present in the sperm acrosome, suggesting roles in sperm maturation and fertilization. We have previously demonstrated that CRES is present within the epididymal lumen as monomeric (14 and N-glycosylated 19-kd forms) as well as sodium dodecyl sulfate-sensitive and sodium dodecyl sulfate-resistant high-molecular mass complexes. We have also shown that recombinant CRES protein will self-aggregate and form amyloid structures in vitro, raising the possibility that CRES might also form amyloid in vivo. Amyloid is a large protein aggregate with a specific cross-ß sheet structure, and its presence is usually associated with disease. This review discusses protein aggregation in the epididymis and provides a brief overview of amyloid formation, including recent studies in other organ systems identifying examples of amyloid that are nonpathologic and carry out biologic functions (ie, functional amyloid). Studies that were carried out to determine if amyloid is present in the epididymal lumen and if CRES is associated with these structures are also described. The presence of CRES amyloid in the mouse epididymal lumen and the absence of pathology suggest either the presence of mechanisms to neutralize the cytotoxicity associated with pathologic amyloid or that CRES is a new example of a functional amyloid with roles in epididymal function.

Extracellular quality control in the epididymis.

The epididymal lumen represents a unique extracellular environment because of the active sperm maturation process that takes place within its confines. Although much focus has been placed on the interaction of epididymal secretory proteins with spermatozoa in the lumen, very little is known regarding how the complex epididymal milieu as a whole is maintained, including mechanisms to prevent or control proteins that may not stay in their native folded state following secretion. Because some misfolded proteins can form cytotoxic aggregate structures known as amyloid, it is likely that control/surveillance mechanisms exist within the epididymis to protect against this process and allow sperm maturation to occur. To study protein aggregation and to identify extracellular quality control mechanisms in the epididymis, we used the cystatin family of cysteine protease inhibitors, including cystatin-related epididymal spermatogenic and cystatin C as molecular models because both proteins have inherent properties to aggregate and form amyloid. In this chapter, we present a brief summary of protein aggregation by the amyloid pathway based on what is known from other organ systems and describe quality control mechanisms that exist intracellularly to control protein misfolding and aggregation. We then present a summary of our studies of cystatin-related epididymal spermatogenic (CRES) oligomerization within the epididymal lumen, including studies suggesting that transglutaminase cross-linking may be one mechanism of extracellular quality control within the epididymis.