SUN domain proteins are integral proteins of the inner nuclear membrane and functions in linkage of the nuclear lamina to the cytoskeleton. Moreover, SUN domain proteins seem to mediate the tethering of the centrosome to the nuclear membrane, and they are involved in telomere attachment to the nuclear envelope in meiotic cells, and in germ cell development in invertebrates. In contrast to the widely expressed SUN domain proteins in mammals, SUN1 and SUN2, which have been analysed in great detail, there is virtually nothing known about testicular SUN domain proteins. Since a hallmark of male germ cell development is the profound remodelling of the nuclear envelope, emphasized, for example, by the reshaping of the nucleus during spermiogenesis, and the biogenesis of its tightly associated acrosome, SUN domain proteins might be engaged in these processes. We have isolated a novel SUN domain protein, SPAG4L-2, that differs from SPAG4L by an N-terminal insertion of 25 amino acids. Spag4l and Spag4l-2 are exclusively expressed in testis at about equimolar amounts, and show elevated transcription during ongoing spermiogenesis coincident with the appearance of round spermatids. Molecular dissection of the protein followed by cytological and biochemical investigations revealed that SPAG4L-2 and SPAG4L are transmembrane proteins that localize to the nuclear envelope. SPAG4L/4L-2 are restricted to the apical nuclear region of round spermatids that face the acrosomic vesicle, and thus are most probably involved in linkage of the acrosomic vesicle to the spermatid nucleus, and in acrosome biogenesis.