title: Cullin3 is a KLHL10-interacting protein preferentially expressed during late spermiogenesis.
abstract: Kelch-like 10 (KLHL10) is a member of the BTB (Bric-a-brac, Tramtrack, and Broad-Complex)-kelch protein superfamily essential for spermiogenesis and male fertility. In a search for KLHL10-interacting proteins using a yeast two-hybrid assay, we identified Cullin3 (CUL3) as one of multiple KLHL10-interacting partners. Yeast cotransformation assays revealed that CUL3 bound the BTB/POZ domain of KLHL10. Northern blot and quantitative RT-PCR analyses demonstrated that Cul3 mRNA was preferentially expressed in the testis. In situ hybridization analysis localized Cul3 mRNA to spermatids in the adult testis. CUL3 protein was detected in elongating and elongated spermatids (steps 10-16) by immunofluorescent microscopy. The expression pattern of CUL3 resembles KLHL10. CUL3 was coimmunoprecipated with KLHL10, and KLHL10 was also detected in the CUL3 immunoprecipitants using testis lysates. These findings suggest that KLHL10, like other BTB/kelch proteins, interacts with CUL3 to form a CUL3-based ubiquitin E3 ligase that functions specifically in the testis to mediate protein ubiquitination during spermiogenesis.
authors.affiliation: Department of Physiology and Cell Biology, University of Nevada School of Medicine, Reno, NV 89557, USA.
authors.forename: Shouhua
authors.initials: S
authors.lastname: Wang
authors.forename: Huili
authors.initials: H
authors.lastname: Zheng
authors.forename: Yumiko
authors.initials: Y
authors.lastname: Esaki
authors.forename: Fiona
authors.initials: F
authors.lastname: Kelly
authors.forename: Wei
authors.initials: W
authors.lastname: Yan
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000378:metabolism*; D044842:Cullin Proteins / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009687:Nuclear Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013087:Spermatids / Q000378:metabolism*; D013091:Spermatogenesis / Q000502:physiology*; D013737:Testis / Q000378:metabolism; D020798:Two-Hybrid System Techniques
chemical_list: D018797:Cell Cycle Proteins; C503646:Cul3 protein, mouse; D044842:Cullin Proteins; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; C487517:Klhl10 protein, mouse; D009687:Nuclear Proteins; D012333:RNA, Messenger