(M361) THE POSTACROSOMAL ASSEMBLY OF SPERM HEAD PROTEIN, PAWP, IS INDEPENDENT OF ACROSOME FORMATION AND DEPENDENT ON MICROTUBULAR MANCHETTE TRANSPORT.
Wu, Alexander1, Sutovsky, Peter2, van der Spoel, Aarnoud 3, Oko, Richard1, 1 Queen's University, Kingston, ON, Canada2 University of Missouri-Columbia, Columbia, MO3 University of Oxford, Oxford, United Kingdom
ABSTRACT- PAWP (postacrosomal sheath WW domain binding protein), exclusively resides in the postacrosomal sheath (PAS) of the perinuclear theca (PT). Because of the importance of this region in initiating fertilization (Sutovsky et al. 2003), we were interested in resolving the origin and assembly of its proteins during spermatogenesis, utilizing PAWP as a model. Based on previous PT developmental studies we hypothesized that the assembly of PAWP is dependent on intramanchette protein transport and manchette descent and independent of subacrosomal PT formation. Utilizing specific antibodies, PAWP was first detected in the cytoplasmic lobe of spermatids beginning to undergo elongation and became most prominent in this region just prior to and during manchette descent. During this peak period, PAWP was concentrated over the apical half of the manchette and co-localized with alpha-tubulin. It was then assembled as part of the PAS in the wake of manchette descent. PAWP mRNA, on the other hand, was first detected in late pachytene spermatocytes, peaked in round spermatids, and declined during spermatid elongation. In order to confirm that PAWP-PAS assembly was independent of subacrosomal PT development, PAWP immunolocalization was performed on the testes of NB-DNJ treated mice who fail to form an acrosome and subacrosomal layer during spermiogenesis (van der Spoel et al., 2002 and unpublished data) but whose elongated spermatids still retain egg activating ability (Suganuma et. al., 2004). The same temporal and manchette based pattern of PAWP-PAS assembly during spermiogenesis was evident as in controls supporting our hypothesis that PAS assembly is independent of subacrosomal PT formation and that egg activating ability resides within the PAS. (Supported by CIHR, NIH, and USDA)
KEY WORDS: Perinuclear theca, Postacrosomal sheath, Intramanchette protein transport, Spermiogenesis