We have identified KRP3, a novel kinesin-related protein expressed in the mammalian testis, and have examined the tissue distribution and subcellular localization of isoforms of this protein. Isolation of KRP3 clones, using the head domain identified in a previous PCR screen as probe, identified at least two KRP3 isoforms in the rat. We have isolated coding sequences of two highly related cDNAs from the rat testis that we have termed KRP3A and KRP3B (kinesin-related protein 3, A and B). Both cDNAs code for predicted polypeptides with the three-domain structure typical of kinesin superfamily members; namely a conserved motor domain, a region capable of forming a limited coiled-coil secondary structure, and a globular tail domain. Although almost identical in their head and stalk domains, these motors diverge in their tail domains. This group of motors is found in many tissues and cell types. The KRP3B motor contains DNA-binding motifs and an RCC1 (regulator of chromosome condensation 1) consensus sequence in its tail domain. Despite this similarity, KRP3B is not associated with the same structures as RCC1. Instead, KRP3 isoforms localize with the nuclei of developing spermatids, and their immunolocalization in the testis overlaps with that of the small GTPase Ran. Like Ran, KRP3 motors are associated in a polarized fashion with the nucleus of maturing spermatids at various stages of elongation. Our findings suggest a possible role for KRP3 motor isoforms in spermatid maturation mediated by possible interaction with the Ran GTPase.