Spermatid Perinuclear Ribonucleic Acid-Binding Protein Binds Microtubules In Vitro and Associates with Abnormal Manchettes In Vivo in Mice(3)

31 Jan

Vivo in Mice(3)

We have previously described an RNA-binding protein in mammalian spermatids whose association with the man-chette suggests that it may be involved in the transport or translation of selected mRNAs. The gene encoding the spermatid perinuclear RNA-binding protein (SPNR) was cloned in a screen for RNA-binding proteins that could bind to the 3′ untranslated region (UTR) of the testis-specific protamine 1 (Prm1) mRNA in vitro. The Prml gene encodes a small, highly basic protein that is involved in DNA packaging in sperm heads. Synthesis of the PRM1 protein is under posttranscriptional control in spermatids and is controlled by sequences in its 3′ UTR. Separate elements within the 3′ UTR appear to be involved in controlling both translational repression in round spermatids and translational activation of the Prml message in elongated spermatids. Translational delay of the Prml mRNA is essential for spermatid differentiation, as premature translation of the Prml message leads to precocious nuclear condensation and an arrest in spermatogenesis. ventolin inhalers

Immunolocalization studies have shown that SPNR is associated with the manchette and that its appearance temporally coincides with the translational activation of the Prml mRNA. SPNR contains two copies of a doublestranded RNA-binding motif that is found in several other proteins. Members of this protein family include Drosophila Staufen, murine protamine RNA-binding protein (PRBP), and human protein kinase activated by RNA. SPNR, and other family members that contain the double-stranded RNA-binding domain, bind double-stranded RNA and highly structured single-stranded RNAs in vitro. Despite the lack of sequence-specific binding in vitro, it is clear that certain members of the family do interact with specific RNA targets in vivo. We have suggested that SPNR interacts with the Prml mRNA in vivo, although in vitro RNA-binding studies with portions of the SPNR protein failed to reveal sequence-specific binding to any RNAs, including the Prml 3′ UTR.

It is possible that SPNR binds microtubules directly, or that its association with the manchette is indirect and that it binds to manchette microtubules via an MAP like a microtubule motor protein. Revealing the nature of SPNR’s interaction with the manchette may provide insight into its function during spermiogenesis. We sought to determine whether SPNR binds microtubules in vitro and whether the binding is direct or indirect. We also determined the sub-cellular localization of SPNR in several mouse mutants that have highly abnormal manchettes and unusual head morphology.