An omega-3 fatty acid, docosahexaenoic acid (DHA), is enriched in testicular membrane phospholipids, but its function is not well understood. The Fads2 gene encodes an enzyme required for the endogenous synthesis of DHA. Using Fads2-null mice (Fads2-/-), we found in our preceding studies that DHA deficiency caused the arrest of spermiogenesis and male infertility, both of which were reversed by dietary DHA. In this study, we investigated a cellular mechanism underlying the DHA essentiality in spermiogenesis. Periodic acid-Schiff staining and acrosin immunohistochemistry revealed the absence of acrosomes in Fads2-/- round spermatids. Acrosin, an acrosomal marker, was scattered throughout the cytoplasm of the Fads2-/- spermatids, and electron microscopy showed that proacrosomal granules were formed on the trans-face of the Golgi. However, excessive endoplasmic reticulum and vesicles were present on the cis-face of the Golgi in Fads2-/- spermatids. The presence of proacrosomal vesicles but lack of a developed acrosome in Fads2-/- spermatids suggested failed vesicle fusion. Syntaxin 2, a protein involved in vesicle fusion, colocalized with acrosin in the acrosome of wild-type mice. In contrast, syntaxin 2 remained scattered in reticular structures and showed no extensive colocalization with acrosin in the Fads2-/- spermatids, suggesting failed fusion with acrosin-containing vesicles or failed transport and release of syntaxin 2 vesicles from Golgi. Dietary supplementation of DHA in Fads2-/- mice restored an intact acrosome. In conclusion, acrosome biogenesis under DHA deficiency is halted after release of proacrosomal granules. Misplaced syntaxin 2 suggests an essential role of DHA in proper delivery of membrane proteins required for proacrosomal vesicle fusion.

Spermatid maturation and retention in the testis depend on the establishment and maintenance of a cell adhesion complex formed between spermatids and Sertoli cells. Deficiency in docosahexaenoic acid (DHA) was recently shown to disrupt postmeiotic germ cell maturation and result in infertility; however a role for DHA in germ cell adhesion has yet to be defined. Here, we use Fads2−/− mice to investigate the effect of a DHA deficiency on this adhesion process. We show impaired retention of immature spermatids in the seminiferous epithelium and their abnormal appearance in the nearest distal tissue, the epididymis. With immunohistochemistry we show that an essential adhesion protein expressed in spermatids (nectin‐3), and its binding partner in Sertoli cells (nectin‐2), each fail to organize at the sites of spermatid‐Sertoli contact, whereas both proteins organize normally in other regions of these cells. Further, vesicular transport proteins golgin‐97 and VAMP4, normally redistributed to the spermatid‐Sertoli adhesion at the initiation of its assembly, failed to do so under a DHA deficiency. In conclusion, spermatid retention is impaired under DHA deficiency due to a failure of spermatid‐Sertoli adhesion construction; the selective mislocalization of nectin‐2 and nectin‐3 suggests a role for DHA in certain specialized systems of complex adhesion assembly.Grant Funding Source: Hatch project ILLU‐971–354 through the Division of Nutritional Sciences Vision 20/20 program