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SCIENCE CHINA Life Sciences, Volume 62 , Issue 4 : 544-552(2019) https://doi.org/10.1007/s11427-018-9469-4

The deubiquitinating gene Usp29 is dispensable for fertility in male mice

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  • ReceivedNov 10, 2018
  • AcceptedDec 1, 2018
  • PublishedMar 22, 2019

Abstract

The balanced actions between ubiquitination and deubiquitination precisely control the levels of various proteins vital for spermatogenesis. Ubiquitin-specific processing proteases (USPs) are the largest family of deubiquitinatingenzymes(DUBs), containing more than 50 members. So far, the functions of only a few USPs in male fertility have been studied, the roles of the majority are yet unknown. The present study aimed to explore the function of Usp29 (ubiquitin-specific protease 29) in male fertility. We found that Usp29 showed predominant expression in mouse testis, and its mRNA expression started to increase at 14 days postpartum (dpp), with a peak at 28 and 35 dpp. Using CRISPR/Cas9 technology, we generated Usp29 knockout mice (Usp29/). Usp29/ mice exhibited no overt developmental anomalies. Further examination revealed that Usp29/ mice had normal fertility and showed no detectable difference in the testis/body weight ratio, testicular and epididymal histology as well as epididymal sperm count from the wild-type littermates. Moreover, Usp29 is not a pseudogene in mice. Taken together, our study first reported that though Usp29 is predominantly expressed in the testis, it is not essential for male fertility in mice.


Funded by

the National Key Research and Developmental Program of China(2018YFC1004700,2016YFC1000600)

the Strategic Priority Research Program of the Chinese Academy of Sciences(XDB19000000)

the National Natural Science Foundation of China(31401953,31630050,31890780,31871514,81571495)

Major Program of Development Foundation of Hefei Centre for Physical Science and Technology(2018ZYFX005)


Acknowledgment

We thank Prof. Cheng Deng from Nanjing Normal University and Prof. Zhuqing Shao from Nanjing University for comments. This work was supported by the National Key Research and Developmental Program of China (2018YFC1004700 and 2016YFC1000600), the Strategic Priority Research Program of the Chinese Academy of Sciences (XDB19000000), the National Natural Science Foundation of China (31401953, 31630050, 31890780, 31871514 and 81571495) and Major Program of Development Foundation of Hefei Centre for Physical Science and Technology (2018ZYFX005).


Interest statement

The author(s) declare that they have no conflict of interest.


Supplement

SUPPORTING INFORMATION

Table S1ƒPrimers used in PCR

The supporting information is available online at http://life.scichina.com and https://link.springer.com. The supporting materials are published as submitted, without typesetting or editing. The responsibility for scientific accuracy and content remains entirely with the authors.


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  • Figure 1

    Expression of Usp29. A, RT-PCR using cDNA from various tissues. Rpl17 is used as the internal control. B, RT-PCR using cDNA from different ages. Rpl17 is used as the internal control. M, marker; dpp, day post-partum. *, P<0.05, compared with 3 dpp. Student’s t-test.

  • Figure 2

    Knockout strategy and genotyping of Usp29/ mice. A, Schematic strategy for Usp29/ mice generation. A fragment of 10 base pairs (bp) (c.252–261del) from the exon of Usp29 gene was deleted. B, RT-PCR using DNA from wild-type (WT), Usp29+/ and Usp29/ mice verify the successful disruption of Usp29 gene in Usp29/ mice. C, Sanger sequencing from WT, Usp29+/ and Usp29/ mice further verify the deletion of the fragment (c.252–261del). C, cytosine; G, guanine; T, thymine; A, adinine; nucleotides written in red and red line are targeted nucleotides.

  • Figure 3

    Spermatogenesis and fertility analyses of Usp29/ mice. A, Representative images of testes from 10-week-old wild-type (WT) and Usp29/ mice. Scale bar, 5 mm. B, Testis/body weight ratio of 10-week-old WT and Usp29/ mice. C, Hemotoxylin and eosin (H&E staining) of testes and epididymides from 10-week-old WT and Usp29/mice. Scale bar, 100 μm. D, Total sperm per epididymis from 10-week-old WT and Usp29/mice. E, The number of pups per litter from WT and Usp29/mice. Dots and squares represent the number of pups of each litter for WT and Usp29/mice, respectively. Student’s t-test was performed using graphpad software for all the statistical analysis. n, the number of animals; NS, no significant difference. The data shown are representative images from at least three separate experiments.

  • Figure 4

    Evolutionary and expression analysis of Usp29. A, The phylogenetic relationship between the USP29 and two closely related branches, USP26 and USP37. Red branch is the USP29 branch. B, The Pa and Ps values of Usp29 from 15 mouse genomes were calculated with DnaSP5 software. C, Complementary expression pattern of Usp29 and Usp26 in different tissues.

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