臺灣博碩士論文加值系統

全世界大約有百分之十至十五比例的夫婦受到不孕症的影響，其中男性與女性的因素各占了一半。在不孕男性的Y染色體長臂之AZFc區域上，無精症缺失基因的缺失最為常見，其位於三號體染色體的同源基因類無精症缺失基因也同樣專一表現於生殖細胞，而且這兩者的蛋白質產物都具有高度保留性的RNA-binding motif以及特定的DAZ repeat。類無精症缺失基因在生殖細胞的分化過程中扮演重要的角色，研究指出在基因剔除的小鼠中，造成生殖細胞的缺失；以及影響受質是在轉譯調控的階段。我們在之前的研究中發現，類無精症缺失基因轉錄子在造精過程有所缺陷的男性中，表現量較低。為了闡明類無精症缺失基因在人類造精過程中扮演的角色，我們從二百三十一位患有嚴重寡精蟲症與非輸精管阻塞無精蟲症之台灣不孕男性中篩選變異，找到二個單一核苷酸多型性，分別在核苷酸260 (T12A)與386 (T54A)位置由adenine轉變成guanine，和二個突變點，分別在核苷酸382 (D52E)與427 (K67N)位置由thymine轉變成guanine和adenine轉變成cytosine。其中386位置單一核苷酸多型性在病人較為盛行，為了闡明單一核苷酸多型性和突變點在功能上的意義，我們試圖找出與人類類無精症缺失基因作用的mRNA，經由tether function assay分析，發現SDAD1、絲胺酸酥胺酸激酶蛋白質31、睾丸特異蛋白質1、MVH為類無精症缺失基因的受質。其中T12A和T54A變異點對於降低睾丸特異蛋白質1轉譯效率有關；T12A、D52E、K67N變異點則是對於降低絲胺酸酥胺酸激酶蛋白質31的表現有關；而所有的變異點都與降低MVH蛋白質表現有關，這個結果暗示著不同變異點在造精過程中作用於不同基因時，擁有不同的影響。除此之外，利用RNA-EMSA的實驗，確認睾丸特異蛋白質1和絲胺酸酥胺酸激酶蛋白質31為類無精症缺失基因的受質，更進一步定義出在睾丸特異蛋白質1三端未轉譯區上核苷酸第1-101與第301-414為類無精症缺失基因結合的區域，然而變異點的蛋白質也同樣可以與睾丸特異蛋白質1和絲胺酸酥胺酸激酶蛋白質31結合，這個研究發現變異的蛋白質影響受質並非是經由結合與否，而是可能影響到其它參與在類無精症缺失基因變異點致病機轉的蛋白質。

Infertility affects 10–15% of all couples, and impaired sperm production accounts for about half of these cases. Deleted in Azoospermia (DAZ) gene was originally isolated from the AZFc region on the long arm of the human Y chromosome that is frequently deleted in infertile men. DAZ-like (DAZL), an autosomal homolog located on chromosome 3p24, are expressed exclusively in the germ cells, and their protein products contain a highly conserved RNA-binding motif and a unique DAZ repeat. DAZL gene is essential for the differentiation of germ cells. Loss of germ cells was observed in Dazl knockout mice. And the effect of DAZL regulate substrate is on the translational control. We have previously found that the DAZL transcript amounts were lower in men with spermatogenic failure. To investigate the role of the DAZL gene in human spermatogenesis, DAZL was genotyped in 231 infertile Taiwanese men presenting with severe oligozoospermia and nonobstructive azoospermia. We identified two polymorphisms－A to G transition at nucleotide 386 in exon 3 (T54A), and nucleotide 260 in exon 2 (T12A); as well as two mutations: T to G transversion at nucleotide 382 in exon 3 (D52E) and A to C transversion at nucleotide 427 in exon 3 (K67N). The frequency of T54A allele was found to be more prevalent in patients than in controls (P=0.0004). To characterize the functional significance of DAZL polymorphisms or mutations, we identified the mRNA substrates that are specifically bound by human DAZL protein. We found that SDAD1, STK31, TPX1, MVH were the substrates of DAZL by the tether function assay analysis. Meanwhile, T12A and T54A were associated with decreased translational efficiency for TXP1. T12A, D52E, and K67N of DAZL were associated with decreased protein expression of STK31. All polymorphisms and mutations decreased protein expression of MVH. These results suggest that different polymorphisms may exert different effects on different genes during spermatogenesis. By using RNA-EMSA, we confirmed that TPX1 and STK31 were indeed DAZL substrates. In addition, by using RNA-EMSA, we define 3’UTR 1bp-101bp and 301-414bp of TPX1 interacts with DAZL. However, the variant DAZL proteins also bind to TPX1 and STK31. These findings suggest mutant proteins regulate substrates translation not directly through their binding activity. Other DAZL interacting partners may be involved in pathogenesis of these genetic variants and polymorphisms.

ABSTRACT IN CHINESE..................................i
ABSTRACT IN ENGLISH..................................iii
ACKNOWLEDGMENTS......................................v
TABLE OF CONTENTS....................................vi
LIST OF TABLES.......................................xii
LIST OF FIGURES......................................xiii
LIST OF ABBREVIATIONS................................xv
1. INTRODUCTION......................................1
1.1 Pathogenesis of male infertility.................1
1.2 Process of mammalian spermatogenesis.............1
1.3 Histology of primary spermatogenesis impairment..3
1.4 Sterile genes....................................3
1.5 Y chromosome and male infertility................4
1.6 The roles of DAZ and DAZL gene in spermatogenesis4
1.7 Functions of Dazl................................5
1.7.1 Several substrates are elucidate by diverse approach.............................................5
1.7.1.1 Cdc25C...................................6
1.7.1.2 Cdc25A...................................6
1.7.1.3 Tpx1.....................................6
1.7.1.4 Sdad1....................................7
1.7.1.5 Mvh......................................8
1.7.1.6 Stk31....................................8
1.7.1.7 Sycp3....................................9
1.7.2 Consensus binding sequence of DAZL...........9
1.7.3 The phenotype of Dazl mutant animals.........10
1.7.4 Dazl regulates substrates on the translational level................................................10
1.8 Objectives of this study.........................10

2 MATERIALS AND METHODS..............................12
2.1 Cell culture.....................................12
2.1.1 Maintenance and subculture...................12
2.1.2 Cell stock...................................12
2.2 Genotyping for infertile men.....................12
2.2.1 Subjects.....................................12
2.2.2 Genotyping...................................14
2.3 Total RNA isolation..............................15
2.3.1 Materials....................................15
2.3.2 Method.......................................15
2.4 Reverse Transcriptase-Polymerase Chain Reaction,
RT-PCR...............................................16
2.4.1 Materials....................................16
2.4.2 Method.......................................16
2.5 Real-Time PCR....................................18
2.5.1 Materials....................................18
2.5.2 Method.......................................18
2.6 SDS-polyacrylamide Gel Electrophoresis and Western Blot.................................................19
2.6.1 Materials....................................19
2.6.2 Method.......................................21
2.7 Plasmid DNA preparation and DNA purification.....22
2.7.1 PCR clean up.................................22
2.7.2 Gel Extraction...............................23
2.7.3 Minipreparation of Plasmid DNA...............24
2.7.4 Midi/Maxipreparation of plasmid DNA..........25
2.8 Generation of constructs.........................26
2.8.1 Construction of pcDNA3.1 V5 (B)-MS2-hDAZL His
tag plasmid........................................27
2.8.1.1 Vector...................................27
2.8.1.2 Primer design............................27
2.8.2 Construction of GST-hDAZL plasmid............27
2.8.2.1 Vector...................................27
2.8.2.2 Primer design............................27
2.8.3 Construction of reporter plasmid with 3’UTR
-MS2 hairpin of candidate substrates...............28
2.8.3.1 Vector...................................28
2.8.3.2 Primer design............................28
2.8.4 Construction of series deletion pGEM-T easy-
TPX1 3’UTR plasmid................................29
2.8.4.1 Vector...................................29
2.8.4.2 Primer design ...........................29
2.8.5 Construction of truncated pGEM-T easy-STK31
3’UTR plasmid.....................................29
2.8.5.1 Vector...................................30
2.8.5.2 Primer design............................30
2.8.6 Restriction enzyme digestion30
2.8.7 Ligation.....................................30
2.8.8 Transformation...............................31
2.9 Site-directed mutagenesis........................31
2.9.1 Primer design................................32
2.10 Tether function assay (Transient transfection and luciferase assay)....................................33
2.10.1 Transient transfection......................33
2.10.2 Dual luciferase assay.......................34
2.11 Statistical analysis............................35
2.11.1 Materials...................................35
2.11.2 Method......................................35
2.12 Gene expression and protein purification........35
2.12.1 Gene expression.............................35
2.12.2 Protein purification........................35
2.13 RNA-electrophoretic mobility shift assay
(RNA-EMSA)...........................................36
2.13.1 In vitro transcription and 32P isotope label
RNA probe............................................36
2.13.2 Electrophoresis.............................37
2.14 Biotin pull-down assay..........................38
2.14.1 In vitro transcription of biotin probe........38
2.14.2 Biotin probe pull-down assay..................39
2.15 Computational analysis..........................40
2.15.1 Restriction enzyme site prediction.........40
2.15.2 Multiple sequence alignment................40

3 RESULTS............................................41
3.1 Genotyping for DAZL in the fertile and infertile men..................................................41
3.2 Verify DAZL substrates in human..................41
3.3 TPX1 was specifically bound by DAZL..............42
3.4 Effect of genetic variants on TPX1...............43
3.5 STK31 was specifically bound by DAZL.............43
3.6 Effect of genetic variants on STK31..............44
3.7 Functional analysis of DAZL SNP for MVH..........44
4 DISCUSSIONS........................................45
4.1 The role of DAZL in spermatogenesis..............45
4.2 The possible role of DAZL variant in spermatogenesis......................................45
4.2.1 The amino acid conservation of DAZL variants during evolution.....................................46
4.2.2 The possible protein modification at amino
acid of variant......................................46
4.3 How DAZL variants regulate TPX1..................48
4.4 How DAZL variants regulate STK31.................49
4.5 Conclusion.......................................49

REFERENCES...........................................51 TABLE................................................63
FIGURE...............................................65
APPENDIX.............................................82
CURRICULUM VITAE.....................................85

Agulnik AI, Zharkikh A, Boettger-Tong H, Bourgeron T, McElreavey K, Bishop CE. Evolution of the DAZ gene family suggests that Y-linked DAZ plays little, or a limited, role in spermatogenesis but underlines a recent African origin for human populations. Hum Mol Genet 1998; 7:1371–1377.

Asano M, Furukawa K, Kido M, Matsumoto S, Umesaki Y, Kochibe N, Iwakura Y. Growth retardation and early death of beta-1,4-galactosyltransferase knockout mice with augmented proliferation and abnormal differentiation of epithelial cells. EMBO J 1997; 16:1850-1857.

Baker J, Hardy MP, Zhou J, Bondy C, Lupu F, Bellve AR, Efstratiadis A. Effects of an Igf1 gene null mutation on mouse reproduction. Molecular Endocrinology 1996; 10:903-918.

Baarends WM, Roest HP, Grootegoed JA. The ubiquitin system in gametogenesis. Mol Cell Endocrinol 1999; 151:5-16.

Buscemi G, Saracino F, Masnada D, Carbone ML. The Saccharomyces cerevisiae SDA1 gene is required for actin cytoskeleton organization and cell cycle progression. J Cell Sci 2000; 113:1199-1211.

Busso D, Cohen DJ, Hayashi M, Kasahara M, Cuasnicú PS. Human testicular protein TPX1/CRISP-2: localization in spermatozoa, fate after capacitation and relevance for gamete interaction. Mol Hum Reprod 2005; 11:299-305.

Busso D, Goldweic NM, Hayashi M, Kasahara M, Cuasnicú PS. Evidence for the involvement of testicular protein CRISP2 in mouse sperm-egg fusion. Biology of Reproduction 2007; 76:701–708.

Chai NN, Phillips A, Fernandez A, Yen PH. A putative human male infertility gene DAZLA: genomic structure and methylation status. Mol Hum Reprod 1997; 3:705–708.

Collier B, Gorgoni B, Loveridge C, Cooke HJ, Gray NK. The DAZL family proteins are PABP-binding proteins that regulate translation in germ cells. EMBO J 2005; 24:2656-2566.

Cooke HJ, Lee M, Kerr S, Ruggiu M. A murine homologue of the human DAZ gene is autosomal and expressed only in the male and female gonads. Hum Mol Genet 1996; 5:513–516.

Damjanov I. Pathology of infertility. First edition. Mosby-Year Book, Inc., St. Louis, 1993; 18-21.

Dai T, Vera Y, Salido EC, Yen PH. Characterization of the mouse Dazap1 gene encoding an RNA-binding protein that interacts with infertility factors DAZ and DAZL. BMC Genomics 2001; 2:1471-2164.

De Kretser DM. Male infertility. Lancet 1997; 349:787-90.

De Kretser DM, Burger HG. The Y chromosome and spermatogenesis. N Engl J Med 1997; 336:576-578.

De Kretser DM, Baker HW. Infertility in Men: Recent Advances and Continuing Controversies. The Journal of Clinical Endocrinology & Metabolism 1999; 84:3443-3450.

Du Y, Huang X, Li J, Hu Y, Zhou Z, Sha J. Human testis specific protein 1 expression in human spermatogenesis and involvement in the pathogenesis of male infertility. Fertil Steril 2006; 85:1852-1854.

Eberhart CG, Maines JZ, Wasserman SA. Meiotic cell cycle requirement for a fly homologue of human deleted in azoospermia. Nature 1996; 81:783–785.

Eddy EM, Washburn TF, Bunch DO, Goulding EH, Gladen BC, Lubahn DB, Korach KS. Targeted disruption of the estrogen receptor gene in male mice causes alteration of spermatogenesis and infertility. Endocrinology 1996; 137:4796-4805.

Edelmann W, Cohen PE, Kane M, Lau K, Morrow B, Bennett S, Umar A. Meiotic pachytene arrest in MLH1-deficient mice. Cell 1996; 85:1125-1134.

Foresta C, Moro E, Ferlin A. Y Chromosome Microdeletions and Alterations of Spermatogenesis. Endocrine Reviews 2001; 22:226–239.

Foster JA, Gerton GL. Autoantigen 1 of the guinea pig sperm acrosome is the homologue of mouse Tpx-1 and human TPX1 and is a member of the cysteine-rich secretory protein (CRISP) family. Mol Reprod Dev 1996; 44:221–229.

Fox M, Urano J, Reijo Pera RA. Identification and characterization of RNA sequences to which human PUMILIO-2 (PUM2) and deleted in Azoospermia-like (DAZL) bind. Genomics 2005; 85:92-105.

Gianotten J, Westerveld GH, Leschot NJ, Tanck MW, Lilford RJ, Lombardi MP, van der Veen F. Familial clustering of impaired spermatogenesis: no evidence for a common genetic inheritance pattern. Hum Reprod 2004; 19:71-76.

Golling G, Amsterdam A, Sun Z, Antonelli M, Maldonado E, Chen W, Burgess S, Haldi M, Artzt K, Farrington S, Lin SY, Nissen RM, Hopkins N. Insertional mutagenesis in zebrafish rapidly identifies genes essential for early vertebrate development. Nat Genet 2002; 31:135-140.

Haglund K, Dikic I. Ubiquitylation and cell signaling. EMBO J 2005; 24:3353-3359. Review.

Houston DW, Zhang J, Maines JZ, Wasserman SA, King ML. A Xenopus DAZ-like gene encodes an RNA component of germ plasm and is a functional homologue of Drosophlila boule. Development 1998; 125:171–180.

Houston DW, King ML. A critical role for Xdazl, a germ plasm-localized RNA, in the differentiation of primordial germ cells in Xenopus. Development 2000; 127:447-456.

Jiao X, Trifillis P and Kiledjian M. Identification of target messenger RNA substrates for the murine deleted in azoospermia-like RNA-binding protein. Biol Reprod 2002; 66:475–485.

Kasahara M, Gutknecht J, Brew K, Spurr N, Goodfellow PN. Cloning and mapping of a testis-specific gene with sequence similarity to a sperm-coating glycoprotein gene. Genomics 1989; 5:527-534.

Kierszenbaum AL. Tyrosine protein kinases and spermatogenesis: truncation matters. Mol Reprod Dev 2006; 73:399-403. Review.

Knudson CM, Tung KS, Tourtellotte WG, Brown GA, Korsmeyer SJ. Bax-deficient mice with lymphoid hyperplasia and male germ cell death. Science 1995; 270:96-99.

Larry I. Lipshultz, Stuart S. Infertility in the male. Third edition. Mosby-Year Book, Inc., St. Louis, 1997; 173-174.

Lee KH, Lee S, Kim B, Chang S, Kim SW, Paick JS, Rhee K. Dazl can bind to dynein motor complex and may play a role in transport of specific mRNAs. EMBO J 2006; 25:4263-4270.

Lin YM, Chen CW, Sun HS, Hsu CC, Chen JM, Lin SJ, Lin JS, Kuo PL. Y-chromosome microdeletion and its effect on reproductive decisions in Taiwanese patients presenting with nonobstructive azoospermia. Urology 2000; 56:1041–1046.

a. Lin YM, Chen CW, Sun HS, Tsai SJ, Hsu CC, Teng YN, Lin JS, Kuo PL. Expression patterns and transcript concentrations of the autosomal DAZL gene in testes of azoospermic men. Mol Hum Reprod 2001; 7:1015-1022.

b. Lin YM, Teng YN, Lee PC, Lin YH, Hsu CC, Lin JS, Kuo PL. AZFa candidate gene deletions in Taiwanese patients with spermatogenic failure. J Formos Med Assoc 2001; 100:592–597.

Lin YM, Lin YH, Teng YN, Hsu CC, Lin JSN, Kuo PL. Gene-based screening for Y-chromosome deletions in Taiwanese men presenting with spermatogenic failure. Fertil Steril 2002; 5:897–903.

Maeda T, Sakashita M, Ohba Y, and Nakanishi Y. Molecular cloning of the rat Tpx-1 responsible for the interaction between spermatogenic and sertoli cells. Biochemical and biophysical research communications 1998; 248:140–146.

Maeda T, Nishida J, Nakanishi Y. Expression pattern, subcellular localization and structure-function relationship of rat Tpx-1, a spermatogenic cell adhesion molecule responsible for association with Sertoli cells. Dev Growth Differ 1999; 41:715–722.

Maegawa S, Yamashita M, Yasuda K, Inoue K. Zebrafish DAZ-like protein controls translation via the sequence 'GUUC'. Genes Cells 2002; 7:971-984.

Ma K, Sharkey A, Kirsch S, Vogt P, Keil R, Hargreave TB, McBeath S, Chandley AC. Towards the molecular localization of the AZF locus: mapping of microdeletions in azoospermic men within 14 subintervals of interval 6 of the human Y chromosome. Human Molecular Genetics 1992; 1:29-33.

Marchler-Bauer A, Anderson JB, Derbyshire MK, DeWeese-Scott C, Gonzales NR, Gwadz M, Hao L, He S, Hurwitz DI, Jackson JD, Ke Z, Krylov D, Lanczycki CJ, Liebert CA, Liu C, Lu F, Lu S, Marchler GH, Mullokandov M, Song JS, Thanki N, Yamashita RA, Yin JJ, Zhang D, Bryant SH. A conserved domain database for interactive domain family analysis. Nucleic Acids Res 2007; 35:237-240.

Miyamoto T, Hasuike S, Yogev L, Maduro MR, Ishikawa M, Westphal H, Lamb DJ. Azoospermia in patients heterozygous for a mutation in SYCP3. Lancet 2003; 362:1714-1719.

O’Bryan MK, Loveland KL, Herszfeld D, McFarlane JR, Hearn MTW, de Kretser DE. Identification of a rat testis-specific gene encoding a potential rat outer dense fibre protein. Mol Reprod Dev 1998; 50:313–322.

Olesen C, Hansen C, Bendsen E, Byskov AG, Schwinger E, Lopez-Pajares I, Jensen PK, Kristoffersson U, Schubert R, Van Assche E, Wahlstroem J, Lespinasse J, Tommerup N. Identification of human candidate genes for male infertility by digital differential display. Mol Hum Reprod 2001; 7:11-20.

Page DC. Mouse autosomal homolog of DAZ, a candidate male sterility gene in humans, is expressed in male germ cells before and after puberty. Genomics 1996; 35:346–352.

Padmanabhan K, Richter JD. Regulated Pumilio-2 binding controls RINGO/Spy mRNA translation and CPEB activation. Genes Dev 2006; 20:199-209.

Reijo R, Seligman J, Dinulos MB, Jaffe T, Brown LG, Disteche CM, Russell LD, Ettlin RA, Sinha Hikim AP, Clegg ED. Histological and Histopathological: evaluation of the testis. 1st ed. Cache River Press, St. Louis 1990; 1-58.

Reijo R, Lee TY, Salo P, Alagappan R, Brown LG, Rosenberg M, Rozen S, Jaffe T, Straus D, Hovatta O, Chapelle A, Silber S, Page DC. Diverse spermatogenic defects in humans caused by Y chromosome deletions encompassing a novel RNA-binding protein gene. Nature Genetics 1995; 10: 383-393.

Reynolds N, Collier B, Maratou K, Bingham V, Speed RM, Taggart M, Semple CA, Gray NK, Cooke HJ. Dazl binds in vivo to specific transcripts and can regulate the pre-meiotic translation of Mvh in germ cells. Hum Mol Genet 2005; 14:3899–3909.

R R, Speed R, Taggart M, Cooke HJ. Spermatogenesis in testes of Dazl null mice after transplantation of wild-type germ cells. Reproduction 2003; 126:599-604.

Ruggiu M, Speed R, Taggart M, McKay SJ, Kilanowski F, Saunders P, Dorin J, Cooke HJ. The mouse Dazla gene encodes cytoplasmic protein essential for gametogenesis. Nature 1997; 389:73–77.

Ruggiu M, Cooke HJ. In vivo and in vitro analysis of homodimerisation activity of the mouse Dazl1 protein. Gene 2000; 252:119-126.

Saxena R, Brown LG, Hawkins T, Alagappan RK, Skaletsky H, Reeve MP, Reijo R, Rozen S, Dinulos MB, Disteche CM, Page DC. The DAZ gene cluster on the human Y chromosome arose from an autosomal gene that was transported, repeatedly amplified and pruned. Nat Genet 1996 14:292–299.

Schrans-Stassen BH, Saunders PT, Cooke HJ, de Rooij DG. Nature of the spermatogenic arrest in Dazl -/- mice. Biol Reprod 2001; 65:771-776.

Seshagiri PB, Thomas M, Sreekumar A, Ray P, Mariappa D. Pentoxifylline induced signalling events during capacitation of hamster spermatozoa: significance of protein tyrosine phosphorylation. Cell Mol Biol 2003; 49:371-80.

Shan Z, Hirschmann P, Seebacher T, Edelmann A, Jauch A, Morell J, Urbitsch P, Vogt PH. A SPGY copy homologous to the mouse gene Dazla and the Drosophila gene boule is autosomal and expressed only in the human male gonad. Hum Mol Genet 1996 5:2005–2011.

Slee R, Grimes B, Speed RM, Taggart M, Maguire SM, Ross A, McGill NI, Saunders PT, Cooke HJ. A human DAZ transgene confers partial rescue of the mouse Dazl null phenotype. Proc Natl Acad Sci USA 1999; 96:8040–8045.

Tanaka SS, Toyooka Y, Akasu R, Katoh-Fukui Y, Nakahara Y, Suzuki R, Yokoyama M, Noce T. The mouse homolog of Drosophila Vasa is required for the development of male germ cells. Genes Dev 2000; 14:841-853.

Teng YN, Lin YM, Lin YH, Tsao SY, Hsu CC, Lin SJ, Tsai WC, Kuo PL. Association of a single-nucleotide polymorphism of the deleted-in-azoospermia-like gene with susceptibility to spermatogenic failure. J Clin Endocrinol Metab 2002; 87:5258-5264.

Tiepolo L, Zuffardi O. Localization of factors controlling spermatogenesis in the nonfluorescent portion of the human Y chromosome long arm. Human Genetics 1976; 34:119-124.

Toscani A, Mettus RV, Coupland R, Simpkins H, Litvin J, Orth J, Hatton KS. Arrest of spermatogenesis and defective breast development in mice lacking A-myb. Nature 1997; 386:713-717.

Toyooka Y, Tsunekawa N, Takahashi Y, Matsui Y, Satoh M, Noce T. Expression and intracellular localization of mouse Vasa-homologue protein during germ cell development. Mech Dev 2000; 93:139-149.

Tuerlings JH, Kremer JA, Meuleman EJ. The practical application of genetics in the male infertility clinic. J Androl 1997; 18:576-581.

Venables JP, Ruggiu M, and Cooke HJ. The RNA-binding specificity of the mouse Dazl protein. Nucleic Acids Res 2001; 29:2479–2483.

Wang P, McCarrey J, Yang F, Page D. An abundance of X-linked genes expressed in spermatogonia. Nat Genet 2001; 27:422– 426.

Weger S, Hammer E, Engstler M. The DNA topoisomerase I binding protein topors as a novel cellular target for SUMO-1 modification: characterization of domains necessary for subcellular localization and sumolation. Exp Cell Res 2003; 290:13-27.

Yen PH, Chai NN, Salido EC. The human autosomal gene DAZLA: testis specificity and a candidate for male infertility. Hum Mol Genet 1996 5:2013–2017.

Yuan L, Liu JG, Zhao J, Brundell E, Daneholt B, Hoog C. The murine SCP3 gene is required for synaptonemal complex assembly, chromosome synapsis, and male fertility. Mol Cell 2000; 5:73–83.

Zimmerman ZA, Kellogg DR. The Sda1 protein is required for passage through start. Mol Biol Cell 2001; 12:201-219.