1. Shanghai Institute of Hematology, State Key Laboratory for Medical Genomics, National Research Center for Translational Medicine at Shanghai, Collaborative Innovation Center of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China 2. Key Laboratory of Laparoscopic Technology of Zhejiang Province, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China 3. Department of General Surgery, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China
Cullin-RING E3 ubiquitin ligase (CRL)-4 is a member of the large CRL family in eukaryotes. It plays important roles in a wide range of cellular processes, organismal development, and physiological and pathological conditions. DDB1- and CUL4-associated factor 8 (DCAF8) is a WD40 repeat-containing protein, which serves as a substrate receptor for CRL4. The physiological role of DCAF8 is unknown. In this study, we constructed Dcaf8 knockout mice. Homozygous mice were viable with no noticeable abnormalities. However, the fertility of Dcaf8-deficient male mice was markedly impaired, consistent with the high expression of DCAF8 in adult mouse testis. Sperm movement characteristics, including progressive motility, path velocity, progressive velocity, and track speed, were significantly lower in Dcaf8 knockout mice than in wild-type (WT) mice. However, the total motility was similar between WT and Dcaf8 knockout sperm. More than 40% of spermatids in Dcaf8 knockout mice showed pronounced morphological abnormalities with typical bent head malformation. The acrosome and nucleus of Dcaf8 knockout sperm looked similar to those of WT sperm. In vitro tests showed that the fertilization rate of Dcaf8 knockout mice was significantly reduced. The results demonstrated that DCAF8 plays a critical role in spermatogenesis, and DCAF8 is a key component of CRL4 function in the reproductive system.
N Schultz, FK Hamra, DL Garbers. A multitude of genes expressed solely in meiotic or postmeiotic spermatogenic cells offers a myriad of contraceptive targets. Proc Natl Acad Sci USA 2003; 100(21): 12201–12206 https://doi.org/10.1073/pnas.1635054100
pmid: 14526100
8
CC Hou, WX Yang. New insights to the ubiquitin-proteasome pathway (UPP) mechanism during spermatogenesis. Mol Biol Rep 2013; 40(4): 3213–3230 https://doi.org/10.1007/s11033-012-2397-y
pmid: 23268313
9
WM Baarends, R van der Laan, JA Grootegoed. Specific aspects of the ubiquitin system in spermatogenesis. J Endocrinol Invest 2000; 23(9): 597–604 https://doi.org/10.1007/BF03343782
pmid: 11079455
10
D Kopanja, N Roy, T Stoyanova, RA Hess, S Bagchi, P Raychaudhuri. Cul4A is essential for spermatogenesis and male fertility. Dev Biol 2011; 352(2): 278–287 https://doi.org/10.1016/j.ydbio.2011.01.028
pmid: 21291880
11
T Ma, JA Keller, X Yu. RNF8-dependent histone ubiquitination during DNA damage response and spermatogenesis. Acta Biochim Biophys Sin (Shanghai) 2011; 43(5): 339–345 https://doi.org/10.1093/abbs/gmr016
pmid: 21444325
12
S Wang, H Zheng, Y Esaki, F Kelly, W Yan. Cullin3 is a KLHL10-interacting protein preferentially expressed during late spermiogenesis. Biol Reprod 2006; 74(1): 102–108 https://doi.org/10.1095/biolreprod.105.045484
pmid: 16162871
13
LY Lu, J Wu, L Ye, GB Gavrilina, TL Saunders, X Yu. RNF8-dependent histone modifications regulate nucleosome removal during spermatogenesis. Dev Cell 2010; 18(3): 371–384 https://doi.org/10.1016/j.devcel.2010.01.010
pmid: 20153262
14
MD Petroski, RJ Deshaies. Function and regulation of Cullin-RING ubiquitin ligases. Nat Rev Mol Cell Biol 2005; 6(1): 9–20 https://doi.org/10.1038/nrm1547
pmid: 15688063
W Zhong, H Feng, FE Santiago, ET Kipreos. CUL-4 ubiquitin ligase maintains genome stability by restraining DNA-replication licensing. Nature 2003; 423(6942): 885–889 https://doi.org/10.1038/nature01747
pmid: 12815436
J Cheng, J Guo, BJ North, K Tao, P Zhou, W Wei. The emerging role for Cullin 4 family of E3 ligases in tumorigenesis. Biochim Biophys Acta Rev Cancer 2019; 1871(1): 138–159 https://doi.org/10.1016/j.bbcan.2018.11.007
pmid: 30602127
20
H Wang, L Zhai, J Xu, HY Joo, S Jackson, H Erdjument-Bromage, P Tempst, Y Xiong, Y Zhang. Histone H3 and H4 ubiquitylation by the CUL4-DDB-ROC1 ubiquitin ligase facilitates cellular response to DNA damage. Mol Cell 2006; 22(3): 383–394 https://doi.org/10.1016/j.molcel.2006.03.035
pmid: 16678110
S Angers, T Li, X Yi, MJ MacCoss, RT Moon, N Zheng. Molecular architecture and assembly of the DDB1-CUL4A ubiquitin ligase machinery. Nature 2006; 443(7111): 590–593 https://doi.org/10.1038/nature05175
pmid: 16964240
23
Y Yin, L Liu, C Yang, C Lin, GM Veith, C Wang, P Sutovsky, P Zhou, L Ma. Cell autonomous and nonautonomous function of CUL4B in mouse spermatogenesis. J Biol Chem 2016; 291(13): 6923–6935 https://doi.org/10.1074/jbc.M115.699660
pmid: 26846852
24
CY Lin, CY Chen, CH Yu, IS Yu, SR Lin, JT Wu, YH Lin, PL Kuo, JC Wu, SW Lin. Human X-linked intellectual disability factor CUL4B is required for post-meiotic sperm development and male fertility. Sci Rep 2016; 6(1): 20227 https://doi.org/10.1038/srep20227
pmid: 26832838
25
Y Yin, C Lin, ST Kim, I Roig, H Chen, L Liu, GM Veith, RU Jin, S Keeney, M Jasin, K Moley, P Zhou, L Ma. The E3 ubiquitin ligase Cullin 4A regulates meiotic progression in mouse spermatogenesis. Dev Biol 2011; 356(1): 51–62 https://doi.org/10.1016/j.ydbio.2011.05.661
pmid: 21624359
26
A Ali, BV Mistry, HA Ahmed, R Abdulla, HA Amer, A Prince, AM Alazami, FS Alkuraya, A Assiri. Deletion of DDB1- and CUL4-associated factor-17 (Dcaf17) gene causes spermatogenesis defects and male infertility in mice. Sci Rep 2018; 8(1): 9202 https://doi.org/10.1038/s41598-018-27379-0
pmid: 29907856
27
Y Wu, L Zhou, X Wang, J Lu, R Zhang, X Liang, L Wang, W Deng, YX Zeng, H Huang, T Kang. A genome-scale CRISPR-Cas9 screening method for protein stability reveals novel regulators of Cdc25A. Cell Discov 2016; 2(1): 16014 https://doi.org/10.1038/celldisc.2016.14
pmid: 27462461
28
M Nowak, B Suenkel, P Porras, R Migotti, F Schmidt, M Kny, X Zhu, EE Wanker, G Dittmar, J Fielitz, T Sommer. DCAF8, a novel MuRF1 interaction partner, promotes muscle atrophy. J Cell Sci 2019; 132(17): jcs233395 https://doi.org/10.1242/jcs.233395
pmid: 31391242
29
G Li, T Ji, J Chen, Y Fu, L Hou, Y Feng, T Zhang, T Song, J Zhao, Y Endo, H Lin, X Cai, Y Cang. CRL4DCAF8 ubiquitin ligase targets histone H3K79 and promotes H3K9 methylation in the liver. Cell Rep 2017; 18(6): 1499–1511 https://doi.org/10.1016/j.celrep.2017.01.039
pmid: 28178526
30
D Huang, C Liu, X Sun, X Sun, Y Qu, Y Tang, G Li, T Tong. CRL4DCAF8 and USP11 oppositely regulate the stability of myeloid leukemia factors (MLFs). Biochem Biophys Res Commun 2020; 529(2): 127–132 https://doi.org/10.1016/j.bbrc.2020.05.186
pmid: 32703400
31
CJ Klein, Y Wu, P Vogel, HH Goebel, C Bönnemann, K Zukosky, MV Botuyan, X Duan, S Middha, EJ Atkinson, G Mer, PJ Dyck. Ubiquitin ligase defect by DCAF8 mutation causes HMSN2 with giant axons. Neurology 2014; 82(10): 873–878 https://doi.org/10.1212/WNL.0000000000000206
pmid: 24500646
32
LT Gou, P Dai, JH Yang, Y Xue, YP Hu, Y Zhou, JY Kang, X Wang, H Li, MM Hua, S Zhao, SD Hu, LG Wu, HJ Shi, Y Li, XD Fu, LH Qu, ED Wang, MF Liu. Pachytene piRNAs instruct massive mRNA elimination during late spermiogenesis. Cell Res 2015; 25(2): 266 https://doi.org/10.1038/cr.2015.14
pmid: 25645811
33
LT Gou, JY Kang, P Dai, X Wang, F Li, S Zhao, M Zhang, MM Hua, Y Lu, Y Zhu, Z Li, H Chen, LG Wu, D Li, XD Fu, J Li, HJ Shi, MF Liu. Ubiquitination-deficient mutations in human Piwi cause male infertility by impairing histone-to-protamine exchange during spermiogenesis. Cell 2017; 169(6):1090–1104.e13 https://doi.org/10.1016/j.cell.2017.04.034
pmid: 28552346
34
KJ Livak, TD Schmittgen. Analysis of relative gene expression data using real-time quantitative PCR and the 2(−ΔΔC(T)) method. Methods 2001; 25(4): 402–408 https://doi.org/10.1006/meth.2001.1262
pmid: 11846609
P Quinn, JF Kerin, GM Warnes. Improved pregnancy rate in human in vitro fertilization with the use of a medium based on the composition of human tubal fluid. Fertil Steril 1985; 44(4): 493–498 https://doi.org/10.1016/S0015-0282(16)48918-1
pmid: 3902512
37
Q Chen, H Peng, L Lei, Y Zhang, H Kuang, Y Cao, QX Shi, T Ma, E Duan. Aquaporin3 is a sperm water channel essential for postcopulatory sperm osmoadaptation and migration. Cell Res 2011; 21(6): 922–933 https://doi.org/10.1038/cr.2010.169
pmid: 21135872
P Sutovsky. Ubiquitin-dependent proteolysis in mammalian spermatogenesis, fertilization, and sperm quality control: killing three birds with one stone. Microsc Res Tech 2003; 61(1): 88–102 https://doi.org/10.1002/jemt.10319
pmid: 12672125
41
M Kanatsu-Shinohara, I Onoyama, KI Nakayama, T Shinohara. Skp1-Cullin-F-box (SCF)-type ubiquitin ligase FBXW7 negatively regulates spermatogonial stem cell self-renewal. Proc Natl Acad Sci USA 2014; 111(24): 8826–8831 https://doi.org/10.1073/pnas.1401837111
pmid: 24879440
42
X Hou, W Zhang, Z Xiao, H Gan, X Lin, S Liao, C Han. Mining and characterization of ubiquitin E3 ligases expressed in the mouse testis. BMC Genomics 2012; 13(1): 495 https://doi.org/10.1186/1471-2164-13-495
pmid: 22992278
43
C Yu, YL Zhang, WW Pan, XM Li, ZW Wang, ZJ Ge, JJ Zhou, Y Cang, C Tong, QY Sun, HY Fan. CRL4 complex regulates mammalian oocyte survival and reprogramming by activation of TET proteins. Science 2013; 342(6165): 1518–1521 https://doi.org/10.1126/science.1244587
pmid: 24357321
44
C Yu, YW Xu, QQ Sha, HY Fan. CRL4DCAF1 is required in activated oocytes for follicle maintenance and ovulation. Mol Hum Reprod 2015; 21(2): 195–205 https://doi.org/10.1093/molehr/gau103
pmid: 25371539
45
C Yu, SY Ji, QQ Sha, QY Sun, HY Fan. CRL4-DCAF1 ubiquitin E3 ligase directs protein phosphatase 2A degradation to control oocyte meiotic maturation. Nat Commun 2015; 6(1): 8017 https://doi.org/10.1038/ncomms9017
pmid: 26281983
KA Sutton, MK Jungnickel, HM Florman. A polycystin-1 controls postcopulatory reproductive selection in mice. Proc Natl Acad Sci USA 2008; 105(25): 8661–8666 https://doi.org/10.1073/pnas.0800603105
pmid: 18562295
48
J Zhang, YL Zhang, LW Zhao, JX Guo, JL Yu, SY Ji, LR Cao, SY Zhang, L Shen, XH Ou, HY Fan. Mammalian nucleolar protein DCAF13 is essential for ovarian follicle maintenance and oocyte growth by mediating rRNA processing. Cell Death Differ 2019; 26(7): 1251–1266 https://doi.org/10.1038/s41418-018-0203-7
pmid: 30283081