title: AKAP3 synthesis is mediated by RNA binding proteins and PKA signaling during mouse spermiogenesis.
abstract: Mammalian spermatogenesis is regulated by coordinated gene expression in a spatiotemporal manner. The spatiotemporal regulation of major sperm proteins plays important roles during normal development of the male gamete, of which the underlying molecular mechanisms are poorly understood. A-kinase anchoring protein 3 (AKAP3) is one of the major components of the fibrous sheath of the sperm tail that is formed during spermiogenesis. In the present study, we analyzed the expression of sperm-specific Akap3 and the potential regulatory factors of its protein synthesis during mouse spermiogenesis. Results showed that the transcription of Akap3 precedes its protein synthesis by about 2 wk. Nascent AKAP3 was found to form protein complex with PKA and RNA binding proteins (RBPs), including PIWIL1, PABPC1, and NONO, as revealed by coimmunoprecipitation and protein mass spectrometry. RNA electrophoretic gel mobility shift assay showed that these RBPs bind sperm-specific mRNAs, of which proteins are synthesized during the elongating stage of spermiogenesis. Biochemical and cell biological experiments demonstrated that PIWIL1, PABPC1, and NONO interact with each other and colocalize in spermatids' RNA granule, the chromatoid body. In addition, NONO was found in extracytoplasmic granules in round spermatids, whereas PIWIL1 and PABPC1 were diffusely localized in cytoplasm of elongating spermatids, indicating their participation at different steps of mRNA metabolism during spermatogenesis. Interestingly, type I PKA subunits colocalize with PIWIL1 and PABPC1 in the cytoplasm of elongating spermatids and cosediment with the RBPs in polysomal fractions on sucrose gradients. Further biochemical analyses revealed that activation of PKA positively regulates AKAP3 protein synthesis without changing its mRNA level in elongating spermatids. Taken together, these results indicate that PKA signaling directly participates in the regulation of protein translation in postmeiotic male germ cells, underscoring molecular mechanisms that regulate protein synthesis during mouse spermiogenesis.
authors.affiliation: Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China.
authors.forename: Kaibiao
authors.initials: K
authors.lastname: Xu
authors.affiliation: Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China.
authors.forename: Lele
authors.initials: L
authors.lastname: Yang
authors.affiliation: Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China.
authors.forename: Danyun
authors.initials: D
authors.lastname: Zhao
authors.affiliation: Department of Biology, University of Science and Technology of China, Hefei, China.
authors.forename: Yaoyao
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Key Laboratory of Regenerative Biology, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China qi_huayu@gibh.ac.cn.
authors.forename: Huayu
authors.initials: H
authors.lastname: Qi
mesh_terms: D054758:A Kinase Anchor Proteins / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D060565:Argonaute Proteins / Q000378:metabolism; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D008297:Male; D008815:Mice, Inbred Strains; D039102:Poly(A)-Binding Proteins / Q000378:metabolism; D014176:Protein Biosynthesis / Q000502:physiology; D012323:RNA Processing, Post-Transcriptional / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D013082:Sperm Tail / Q000502:physiology; D013087:Spermatids / Q000166:cytology / Q000502:physiology*; D013091:Spermatogenesis / Q000502:physiology*
chemical_list: D054758:A Kinase Anchor Proteins; C485611:Akap3 protein, mouse; D060565:Argonaute Proteins; D004268:DNA-Binding Proteins; C575837:EPAB protein, mouse; C500331:Nono protein, mouse; C448522:Piwil1 protein, mouse; D039102:Poly(A)-Binding Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: AKAP3; PKA; RNA binding protein; spermiogenesis; translation